phosphonate-peptide nucleic acid: Topics by WorldWideScience.org

Sample records for phosphonate-peptide nucleic acid

Peptide Nucleic Acids

DEFF Research Database (Denmark)

2003-01-01

A novel class of compounds, known as peptide nucleic acids, bind complementary ssDNA and RNA strands more strongly than a corresponding DNA. The peptide nucleic acids generally comprise ligands such as naturally occurring DNA bases attached to a peptide backbone through a suitable linker....
Peptide Nucleic Acids

DEFF Research Database (Denmark)

1998-01-01

A novel class of compounds, known as peptide nucleic acids, bind complementary ssDNA and RNA strands more strongly than a corresponding DNA. The peptide nucleic acids generally comprise ligands such as naturally occurring DNA bases attached to a peptide backbone through a suitable linker....
Double-Stranded Peptide Nucleic Acids

DEFF Research Database (Denmark)

2001-01-01

A novel class of compounds, known as peptide nucleic acids, form double-stranded structures with one another and with ssDNA. The peptide nucleic acids generally comprise ligands such as naturally occurring DNA bases attached to a peptide backbone through a suitable linker.......A novel class of compounds, known as peptide nucleic acids, form double-stranded structures with one another and with ssDNA. The peptide nucleic acids generally comprise ligands such as naturally occurring DNA bases attached to a peptide backbone through a suitable linker....
Peptide Nucleic Acids

DEFF Research Database (Denmark)

2004-01-01

A novel class of compounds known as peptide nucleic acids, bind complementary DNA and RNA strands, and generally do so more strongly than the corresponding DNA or RNA strands while exhibiting increased sequence specificity and solubility. The peptide nucleic acids comprise ligands selected from...
Peptide Nucleic Acids (PNA)

DEFF Research Database (Denmark)

2002-01-01

A novel class of compounds, known as peptide nucleic acids, bind complementary ssDNA and RNA strands more strongly than a corresponding DNA. The peptide nucleic acids generally comprise ligands such as naturally occurring DNA bases attached to a peptide backbone through a suitable linker....
Peptide Nucleic Acid Synthons

DEFF Research Database (Denmark)

2004-01-01

A novel class of compounds, known as peptide nucleic acids, bind complementary ssDNA and RNA strands more strongly than a corresponding DNA. The peptide nucleic acids generally comprise ligands such as naturally occurring DNA bases attached to a peptide backbone through a suitable linker....
Histidine-Containing Peptide Nucleic Acids

DEFF Research Database (Denmark)

2000-01-01

Peptide nucleic acids containing histidine moieties are provided. These compounds have applications including diagnostics, research and potential therapeutics.......Peptide nucleic acids containing histidine moieties are provided. These compounds have applications including diagnostics, research and potential therapeutics....
Cleaving Double-Stranded DNA with Peptide Nucleic Acids

DEFF Research Database (Denmark)

1997-01-01

Peptide nucleic acids and analogues of peptide nucleic acids are used to form duplex, triplex, and other structures with nucleic acids and to modify nucleic acids. The peptide nucleic acids and analogues thereof also are used to modulate protein activity through, for example, transcription arrest......, transcription initiation, and site specific cleavage of nucleic acids....
Synthetic Procedures for Peptide Nucleic Acids

DEFF Research Database (Denmark)

2004-01-01

A novel class of compounds, known as peptide nucleic acids, bind complementary ssDNA and RNA strands more strongly than a corresponding DNA. The peptide nucleic acids generally comprise ligands such as naturally occurring DNA bases attached to a peptide backbone through a suitable linker....
Peptide Nucleic Acids Having Amino Acid Side Chains

DEFF Research Database (Denmark)

1998-01-01

A novel class of compounds, known as peptide nucleic acids, bind complementary DNA and RNA strands more strongly than the corresponding DNA or RNA strands, and exhibit increased sequence specificity and solubility. The peptide nucleic acids comprise ligands selected from a group consisting...
Peptide Nucleic Acids Having 2,6-Diaminopurine Nucleobases

DEFF Research Database (Denmark)

2003-01-01

A novel class of compounds, known as peptide nucleic acids, bind complementary DNA and RNA strands more strongly than a corresponding DNA strand, and exhibit increased sequence specificity and binding affinity. The peptide nucleic acids of the invention comprise ligands selected from a group...
Histidine-lysine peptides as carriers of nucleic acids.

Science.gov (United States)

Leng, Qixin; Goldgeier, Lisa; Zhu, Jingsong; Cambell, Patricia; Ambulos, Nicholas; Mixson, A James

2007-03-01

With their biodegradability and diversity of permutations, peptides have significant potential as carriers of nucleic acids. This review will focus on the sequence and branching patterns of peptide carriers composed primarily of histidines and lysines. While lysines within peptides are important for binding to the negatively charged phosphates, histidines are critical for endosomal lysis enabling nucleic acids to reach the cytosol. Histidine-lysine (HK) polymers by either covalent or ionic bonds with liposomes augment transfection compared to liposome carriers alone. More recently, we have examined peptides as sole carriers of nucleic acids because of their intrinsic advantages compared to the bipartite HK/liposome carriers. With a protocol change and addition of a histidine-rich tail, HK peptides as sole carriers were more effective than liposomes alone in several cell lines. While four-branched polymers with a primary repeating sequence pattern of -HHK- were more effective as carriers of plasmids, eight-branched polymers with a sequence pattern of -HHHK- were more effective as carriers of siRNA. Compared to polyethylenimine, HK carriers of siRNA and plasmids had reduced toxicity. When injected intravenously, HK polymers in complex with plasmids encoding antiangiogenic proteins significantly decreased tumor growth. Furthermore, modification of HK polymers with polyethylene glycol and vascular-specific ligands increased specificity of the polyplex to the tumor by more than 40-fold. Together with further development and insight on the structure of HK polyplexes, HK peptides may prove to be useful as carriers of different forms of nucleic acids both in vitro and in vivo.
Gene Targeting and Expression Modulation by Peptide Nucleic Acids (PNA)

DEFF Research Database (Denmark)

Nielsen, Peter E

2010-01-01

Peptide nucleic acids (PNA) are artificial structural mimics of nucleic acids capable of sequence specific hybridization to both RNA and DNA. Thus they have obvious potential as gene targeting agents for drug discovery approaches. An overview with emphasis on recent progress on RNA "interference...
Recent Developments in Peptide-Based Nucleic Acid Delivery

Directory of Open Access Journals (Sweden)

Tobias Restle

2008-07-01

Full Text Available Despite the fact that non-viral nucleic acid delivery systems are generally considered to be less efficient than viral vectors, they have gained much interest in recent years due to their superior safety profile compared to their viral counterpart. Among these synthetic vectors are cationic polymers, branched dendrimers, cationic liposomes and cellpenetrating peptides (CPPs. The latter represent an assortment of fairly unrelated sequences essentially characterised by a high content of basic amino acids and a length of 10-30 residues. CPPs are capable of mediating the cellular uptake of hydrophilic macromolecules like peptides and nucleic acids (e.g. siRNAs, aptamers and antisenseoligonucleotides, which are internalised by cells at a very low rate when applied alone. Up to now, numerous sequences have been reported to show cell-penetrating properties and many of them have been used to successfully transport a variety of different cargos into mammalian cells. In recent years, it has become apparent that endocytosis is a major route of internalisation even though the mechanisms underlying the cellular translocation of CPPs are poorly understood and still subject to controversial discussions. In this review, we will summarise the latest developments in peptide-based cellular delivery of nucleic acid cargos. We will discuss different mechanisms of entry, the intracellular fate of the cargo, correlation studies of uptake versus biological activity of the cargo as well as technical problems and pitfalls.
Peptides, polypeptides and peptide-polymer hybrids as nucleic acid carriers.

Science.gov (United States)

Ahmed, Marya

2017-10-24

Cell penetrating peptides (CPPs), and protein transduction domains (PTDs) of viruses and other natural proteins serve as a template for the development of efficient peptide based gene delivery vectors. PTDs are sequences of acidic or basic amphipathic amino acids, with superior membrane trespassing efficacies. Gene delivery vectors derived from these natural, cationic and cationic amphipathic peptides, however, offer little flexibility in tailoring the physicochemical properties of single chain peptide based systems. Owing to significant advances in the field of peptide chemistry, synthetic mimics of natural peptides are often prepared and have been evaluated for their gene expression, as a function of amino acid functionalities, architecture and net cationic content of peptide chains. Moreover, chimeric single polypeptide chains are prepared by a combination of multiple small natural or synthetic peptides, which imparts distinct physiological properties to peptide based gene delivery therapeutics. In order to obtain multivalency and improve the gene delivery efficacies of low molecular weight cationic peptides, bioactive peptides are often incorporated into a polymeric architecture to obtain novel 'polymer-peptide hybrids' with improved gene delivery efficacies. Peptide modified polymers prepared by physical or chemical modifications exhibit enhanced endosomal escape, stimuli responsive degradation and targeting efficacies, as a function of physicochemical and biological activities of peptides attached onto a polymeric scaffold. The focus of this review is to provide comprehensive and step-wise progress in major natural and synthetic peptides, chimeric polypeptides, and peptide-polymer hybrids for nucleic acid delivery applications.
Peptide nucleic acids and their potential applications in biotechnology

DEFF Research Database (Denmark)

Buchardt, O.; Egholm, M.; Berg, R.H.

1993-01-01

Peptide nucleic acids (PNAs) are novel DNA mimics in which the sugar-phosphate backbone has been replaced with a backbone based on amino acids1-3. PNAs exhibit sequence-specific binding to DNA and RNA with higher affinities and specificities than unmodified DNA. They,are resistant to nuclease...
Construction of tunable peptide nucleic acid junctions.

Science.gov (United States)

Duan, Tanghui; He, Liu; Tokura, Yu; Liu, Xin; Wu, Yuzhou; Shi, Zhengshuang

2018-03-15

We report here the construction of 3-way and 4-way peptide nucleic acid (PNA) junctions as basic structural units for PNA nanostructuring. The incorporation of amino acid residues into PNA chains makes PNA nanostructures with more structural complexity and architectural flexibility possible, as exemplified by building 3-way PNA junctions with tunable nanopores. Given that PNA nanostructures have good thermal and enzymatic stabilities, they are expected to have broad potential applications in biosensing, drug delivery and bioengineering.
Role of Cell-Penetrating Peptides in Intracellular Delivery of Peptide Nucleic Acids Targeting Hepadnaviral Replication

DEFF Research Database (Denmark)

Ndeboko, Benedicte; Ramamurthy, Narayan; Lemamy, Guy Joseph

2017-01-01

Peptide nucleic acids (PNAs) are potentially attractive antisense agents against hepatitis B virus (HBV), although poor cellular uptake limits their therapeutic application. In the duck HBV (DHBV) model, we evaluated different cell-penetrating peptides (CPPs) for delivery to hepatocytes of a PNA...
Efficacy of peptide nucleic acid and selected conjugates against specific cellular pathologies of amyotrophic lateral sclerosis.

Science.gov (United States)

Browne, Elisse C; Parakh, Sonam; Duncan, Luke F; Langford, Steven J; Atkin, Julie D; Abbott, Belinda M

2016-04-01

Cellular studies have been undertaken on a nonamer peptide nucleic acid (PNA) sequence, which binds to mRNA encoding superoxide dismutase 1, and a series of peptide nucleic acids conjugated to synthetic lipophilic vitamin analogs including a recently prepared menadione (vitamin K) analog. Reduction of both mutant superoxide dismutase 1 inclusion formation and endoplasmic reticulum stress, two of the key cellular pathological hallmarks in amyotrophic lateral sclerosis, by two of the prepared PNA oligomers is reported for the first time. Crown Copyright © 2016. Published by Elsevier Ltd. All rights reserved.
Improved cellular activity of antisense peptide nucleic acids by conjugation to a cationic peptide-lipid (CatLip) domain

DEFF Research Database (Denmark)

Koppelhus, Uffe; Shiraishi, Takehiko; Zachar, Vladimir

2008-01-01

Conjugation to cationic cell penetrating peptides (such as Tat, Penetratin, or oligo arginines) efficiently improves the cellular uptake of large hydrophilic molecules such as oligonucleotides and peptide nucleic acids, but the cellular uptake is predominantly via an unproductive endosomal pathway...... for future in vivo applications. We find that simply conjugating a lipid domain (fatty acid) to the cationic peptide (a CatLip conjugate) increases the biological effect of the corresponding PNA (CatLip) conjugates in a luciferase cellular antisense assay up to 2 orders of magnitude. The effect increases...... with increasing length of the fatty acid (C8-C16) but in parallel also results in increased cellular toxicity, with decanoic acid being optimal. Furthermore, the relative enhancement is significantly higher for Tat peptide compared to oligoarginine. Confocal microscopy and chloroquine enhancement indicates...

Implementation of antimicrobial peptides for sample preparation prior to nucleic acid amplification in point-of-care settings.

Science.gov (United States)

Krõlov, Katrin; Uusna, Julia; Grellier, Tiia; Andresen, Liis; Jevtuševskaja, Jekaterina; Tulp, Indrek; Langel, Ülo

2017-12-01

A variety of sample preparation techniques are used prior to nucleic acid amplification. However, their efficiency is not always sufficient and nucleic acid purification remains the preferred method for template preparation. Purification is difficult and costly to apply in point-of-care (POC) settings and there is a strong need for more robust, rapid, and efficient biological sample preparation techniques in molecular diagnostics. Here, the authors applied antimicrobial peptides (AMPs) for urine sample preparation prior to isothermal loop-mediated amplification (LAMP). AMPs bind to many microorganisms such as bacteria, fungi, protozoa and viruses causing disruption of their membrane integrity and facilitate nucleic acid release. The authors show that incubation of E. coli with antimicrobial peptide cecropin P1 for 5 min had a significant effect on the availability of template DNA compared with untreated or even heat treated samples resulting in up to six times increase of the amplification efficiency. These results show that AMPs treatment is a very efficient sample preparation technique that is suitable for application prior to nucleic acid amplification directly within biological samples. Furthermore, the entire process of AMPs treatment was performed at room temperature for 5 min thereby making it a good candidate for use in POC applications.
Enhanced peptide nucleic acid binding to supercoiled DNA: possible implications for DNA "breathing" dynamics

DEFF Research Database (Denmark)

Bentin, T; Nielsen, Peter E.

1996-01-01

The influence of DNA topology on peptide nucleic acid (PNA) binding was studied. Formation of sequence-specific PNA2/dsDNA (double-stranded DNA) complexes was monitored by a potassium permanganate probing/primer extension assay. At low ionic strengths, the binding of PNA was 2-3 times more...
Subnanomolar antisense activity of phosphonate-peptide nucleic acid (PNA) conjugates delivered by cationic lipids to HeLa cells

DEFF Research Database (Denmark)

Shiraishi, Takehiko; Hamzavi, Ramin; Nielsen, Peter E

2008-01-01

oligomer. This modification of the PNA does not interfere with the nucleic acid target binding affinity based on thermal stability of the PNA/RNA duplexes. When delivered to cultured HeLa pLuc705 cells by Lipofectamine, the PNAs showed dose-dependent nuclear antisense activity in the nanomolar range...
Light-emitting self-assembled peptide nucleic acids exhibit both stacking interactions and Watson-Crick base pairing.

Science.gov (United States)

Berger, Or; Adler-Abramovich, Lihi; Levy-Sakin, Michal; Grunwald, Assaf; Liebes-Peer, Yael; Bachar, Mor; Buzhansky, Ludmila; Mossou, Estelle; Forsyth, V Trevor; Schwartz, Tal; Ebenstein, Yuval; Frolow, Felix; Shimon, Linda J W; Patolsky, Fernando; Gazit, Ehud

2015-04-01

The two main branches of bionanotechnology involve the self-assembly of either peptides or DNA. Peptide scaffolds offer chemical versatility, architectural flexibility and structural complexity, but they lack the precise base pairing and molecular recognition available with nucleic acid assemblies. Here, inspired by the ability of aromatic dipeptides to form ordered nanostructures with unique physical properties, we explore the assembly of peptide nucleic acids (PNAs), which are short DNA mimics that have an amide backbone. All 16 combinations of the very short di-PNA building blocks were synthesized and assayed for their ability to self-associate. Only three guanine-containing di-PNAs-CG, GC and GG-could form ordered assemblies, as observed by electron microscopy, and these di-PNAs efficiently assembled into discrete architectures within a few minutes. The X-ray crystal structure of the GC di-PNA showed the occurrence of both stacking interactions and Watson-Crick base pairing. The assemblies were also found to exhibit optical properties including voltage-dependent electroluminescence and wide-range excitation-dependent fluorescence in the visible region.
Targeted correction of a thalassemia-associated beta-globin mutation induced by pseudo-complementary peptide nucleic acids

DEFF Research Database (Denmark)

Lonkar, Pallavi; Kim, Ki-Hyun; Kuan, Jean Y

2009-01-01

Beta-thalassemia is a genetic disorder caused by mutations in the beta-globin gene. Triplex-forming oligonucleotides and triplex-forming peptide nucleic acids (PNAs) have been shown to stimulate recombination in mammalian cells via site-specific binding and creation of altered helical structures...
Calcium ions effectively enhance the effect of antisense peptide nucleic acids conjugated to cationic tat and oligoarginine peptides

DEFF Research Database (Denmark)

Shiraishi, Takehiko; Pankratova, Stanislava; Nielsen, Peter E

2005-01-01

Cell-penetrating peptides have been widely used to improve cellular delivery of a variety of proteins and antisense agents. However, recent studies indicate that such cationic peptides are predominantly entering cells via an endosomal pathway. We now show that the nuclear antisense effect in He......La cells of a variety of peptide nucleic acid (PNA) peptide conjugates is significantly enhanced by addition of 6 mM Ca(2+) (as well as by the lysosomotrophic agent chloroquine). In particular, the antisense activities of Tat(48-60) and heptaarginine-conjugated PNAs were increased 44-fold and 8.5-fold......, respectively. Evidence is presented that the mechanism involves endosomal release. The present results show that Ca(2+) can be used as an effective enhancer for in vitro cellular delivery of cationic peptide-conjugated PNA oligomers, and also emphasize the significance of the endosomal escape route...
Alkyl phosphonic acids and sulfonic acids in the Murchison meteorite

Science.gov (United States)

Cooper, George W.; Onwo, Wilfred M.; Cronin, John R.

1992-01-01

Homologous series of alkyl phosphonic acids and alkyl sulfonic acids, along with inorganic orthophosphate and sulfate, are identified in water extracts of the Murchison meteorite after conversion to their t-butyl dimethylsilyl derivatives. The methyl, ethyl, propyl, and butyl compounds are observed in both series. Five of the eight possible alkyl phosphonic acids and seven of the eight possible alkyl sulfonic acids through C4 are identified. Abundances decrease with increasing carbon number as observed of other homologous series indigenous to Murchison. Concentrations range downward from approximately 380 nmol/gram in the alkyl sulfonic acid series, and from 9 nmol/gram in the alkyl phosphonic acid series.
Cell number and transfection volume dependent peptide nucleic acid antisense activity by cationic delivery methods

DEFF Research Database (Denmark)

Llovera Nadal, Laia; Berthold, Peter; Nielsen, Peter E

2012-01-01

have now quantitatively compared the cellular activity (in the pLuc705 HeLa cell splice correction system) of PNA antisense oligomers using lipoplex delivery of cholesterol- and bisphosphonate-PNA conjugates, polyplex delivery via a PNA-polyethyleneimine conjugate and CPP delivery via a PNA......Efficient intracellular delivery is essential for high activity of nucleic acids based therapeutics, including antisense agents. Several strategies have been developed and practically all rely on auxiliary transfection reagents such as cationic lipids, cationic polymers and cell penetrating...... peptides as complexing agents and carriers of the nucleic acids. However, uptake mechanisms remain rather poorly understood, and protocols always require optimization of transfection parameters. Considering that cationic transfection complexes bind to and thus may up-concentrate on the cell surface, we...
DNA-like double helix formed by peptide nucleic acid

DEFF Research Database (Denmark)

Wittung, P; Nielsen, Peter E.; Buchardt, O

1994-01-01

Although the importance of the nucleobases in the DNA double helix is well understood, the evolutionary significance of the deoxyribose phosphate backbone and the contribution of this chemical entity to the overall helical structure and stability of the double helix is not so clear. Peptide nucleic...
End-labeling of peptide nucleic acid with osmium complex. Voltammetry at carbon and mercury electrodes

Czech Academy of Sciences Publication Activity Database

Paleček, Emil; Trefulka, Mojmír; Fojta, Miroslav

2009-01-01

Roč. 11, č. 2 (2009), s. 359-362 ISSN 1388-2481 R&D Projects: GA AV ČR(CZ) KAN400310651; GA MŠk(CZ) LC06035 Institutional research plan: CEZ:AV0Z50040507; CEZ:AV0Z50040702 Keywords : peptide nucleic acid end-labeling * osmium tetroxide complexes * electroactive labels Subject RIV: BO - Biophysics Impact factor: 4.243, year: 2009
Reactions of acetylenes with hypophosphorous and phosphonous acids

International Nuclear Information System (INIS)

Nifant'ev, E.E.; Solovetskaya, L.A.; Maslennikova, V.I.; Magdeeva, R.K.; Sergeev, N.M.

1986-01-01

Alkenylphosphonous, alkenylphosphinic, and alkylenebisphosphinic acids were prepared for the first time. The course of the hydrophosphorylation of 1-alkynes depends on the form of the reagent used. The homolytic hydrophosphorylation of acetylenes with hypophosphorous acid proceeds mainly with the participation of one molecule of the acid. The addition of phosphonous acids to acetylenic hydrocarbons leads to both mono- and di-phosphorylated adducts. The nature of the phosphonous acid taken has a substantial influence on the readiness with which the alkylenebisphosphinic acids are formed. Study of the stereoselectivity of the reactions showed that in the case of the monoaddition of hypophosphorous and phosphonous acids to acetylenes mixtures of cis and trans isomeric products are always formed. Phosphorous 31 and carbon 13 NMR spectra are analyzed
Self-ordered Porous Alumina Fabricated via Phosphonic Acid Anodizing

OpenAIRE

Akiya, Shunta; Kikuchi, Tatsuya; Natsui, Shungo; Sakaguchi, Norihito; Suzuki, Ryosuke O.

2016-01-01

Self-ordered periodic porous alumina with an undiscovered cell diameter was fabricated via electrochemical anodizing in a new electrolyte, phosphonic acid (H3PO3). High-purity aluminum plates were anodized in phosphonic acid solution under various operating conditions of voltage, temperature, concentration, and anodizing time. Phosphonic acid anodizing at 150-180 V caused the self-ordering behavior of porous alumina, and an ideal honeycomb nanostructure measuring 370-440 nm in cell diameter w...
Peptide nucleic acid (PNA) antisense effects in Escherichia coli

DEFF Research Database (Denmark)

Good, L; Nielsen, P E

1999-01-01

Antisense peptide nucleic acid (PNA) can be used to control cell growth, gene expression and growth phenotypes in the bacteria Escherichia coli. PNAs targeted to the RNA components of the ribosome can inhibit translation and cell growth, and PNAs targeted to mRNA can limit gene expression with gene...... and sequence specificity. In an E. coli cell extract, efficient inhibition is observed when using PNA concentrations in the nanomolar range, whereas micromolar concentrations are required for inhibition in growing cells. A mutant strain of E. coli that is more permeable to antibiotics also is more susceptible...... to antisense PNAs than the wild type. This chapter details methods for testing the antisense activities of PNA in E. coli. As an example of the specific antisense inhibition possible, we show the effects of an anti-beta-galactosidase PNA in comparison to control PNAs. With improvements in cell uptake...
Peptide nucleic acid probe for protein affinity purification based on biotin-streptavidin interaction and peptide nucleic acid strand hybridization.

Science.gov (United States)

Tse, Jenny; Wang, Yuanyuan; Zengeya, Thomas; Rozners, Eriks; Tan-Wilson, Anna

2015-02-01

We describe a new method for protein affinity purification that capitalizes on the high affinity of streptavidin for biotin but does not require dissociation of the biotin-streptavidin complex for protein retrieval. Conventional reagents place both the selectively reacting group (the "warhead") and the biotin on the same molecule. We place the warhead and the biotin on separate molecules, each linked to a short strand of peptide nucleic acid (PNA), synthetic polymers that use the same bases as DNA but attached to a backbone that is resistant to attack by proteases and nucleases. As in DNA, PNA strands with complementary base sequences hybridize. In conditions that favor PNA duplex formation, the warhead strand (carrying the tagged protein) and the biotin strand form a complex that is held onto immobilized streptavidin. As in DNA, the PNA duplex dissociates at moderately elevated temperature; therefore, retrieval of the tagged protein is accomplished by a brief exposure to heat. Using iodoacetate as the warhead, 8-base PNA strands, biotin, and streptavidin-coated magnetic beads, we demonstrate retrieval of the cysteine protease papain. We were also able to use our iodoacetyl-PNA:PNA-biotin probe for retrieval and identification of a thiol reductase and a glutathione transferase from soybean seedling cotyledons. Copyright © 2014 Elsevier Inc. All rights reserved.
Summarization on the synthesis and radionuclide-labeling of peptide nucleic acid for an oligonucleotide analogue

International Nuclear Information System (INIS)

Song, Hongtao; Zhang, Huaming; Gao, Hui

2009-04-01

Peptide nucleic acid (PNA), which is one kind of antisense nucleic acid compounds and an oligonucleotide analogue that binds strongly to DNA and RNA in a sequence specific manner, has its unique advantages in the field of molecular diagnostics and treatment of diseases. Now, people gradually attach more importance to PNA. To optimize the application of PNA in genetic re- search and therapy, a great number of backbone modifications on the newly- type structures of PNA were synthesized to improve its physicochemical proper- ties, such as hybridization speciality, solubility in biofluid, or cell permeability. The modified PNA labeled with radionuclides, which can obtain the aim at specific target and minimal non-target trauma, has important role in research and application of tumorous genitherapy. Here a review on the basic synthesis idea and several primary synthetic methods of PNA analogs was given, and also correlative studies and expectation on the compounds belonging to PNA series labeled with radionuclides were included. (authors)
Promotion of double-duplex invasion of peptide nucleic acids through conjugation with nuclear localization signal peptide.

Science.gov (United States)

Aiba, Yuichiro; Honda, Yuta; Komiyama, Makoto

2015-03-02

Pseudo-complementary peptide nucleic acid (pcPNA), as one of the most widely used synthetic DNA analogues, invades double-stranded DNA according to Watson-Crick rules to form invasion complexes. This unique mode of DNA recognition induces structural changes at the invasion site and can be used for a range of applications. In this paper, pcPNA is conjugated with a nuclear localization signal (NLS) peptide, and its invading activity is notably promoted both thermodynamically and kinetically. Thus, the double-duplex invasion complex is formed promptly at low pcPNA concentrations under high salt conditions, where the invasion otherwise never occurs. Furthermore, NLS-modified pcPNA is successfully employed for site-selective DNA scission, and the targeted DNA is selectively cleaved under conditions that are not conducive for DNA cutters using unmodified pcPNAs. This strategy of pcPNA modification is expected to be advantageous and promising for a range of in vitro and in vivo applications. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.
Thermodynamics and kinetics of thermal decomposition of dibutylalkyl and dipentylalkyl phosphonate-nitric acid systems

International Nuclear Information System (INIS)

Chandran, K.; Brahmananda, C.V.S.; Anthonysamy, S.; Srinivasan, T.G.; Ganesan, V.

2013-01-01

Tributyl phosphate (TBP) is used in the reprocessing industry for the separation of uranium and plutonium from the spent nuclear fuels by PUREX process. A variety of dialkylalky/phosphonates were synthesised and their extraction behaviour on actinides was studied in this laboratory. The superior extraction characteristics of phosphonates make them useful for several metal recovery applications in the nuclear fuel cycle. However, higher solubility of phosphonates in aqueous phase as compared to TBP, introduces higher chances of the formation of thermally active 'red oil' like substances during the evaporation of aqueous streams. Thermal stability of neat and nitric acid solvated homologues of four dibutylalkyl phosphonates namely, dibutylpropyl phosphonate (DBPrP), dibutylbutyl phosphonate (DBBP), dibutylpentyl phosphonate (DBPP), dibutylhexyl phosphonate (DBHeP), and two dipentylalkyl phosphonates viz. dipentylbutyl phosphonate (DPBP), dipentylhexyl phosphonate (DPHeP) were studied. Experiments were conducted using an adiabatic calorimeter in heat-wait-search mode. Various stoichiometry of nitric acid-solvated DBalP and DPalP were prepared by equilibrating the respective organic with 4-15.6 M nitric acid followed by separation. Neat DBalP and DPalP are stable up to the temperature of ∼ 555 K while their acid-solvates decomposed in the temperature range 380-403 K. The results also indicated that the presence of nitric acid accelerated the decomposition of phosphonates. Decomposition of these acid solvated phosphonates is found to be exothermic and follows first order kinetics. The onset temperature, adiabatic temperature rise, pressure rise, decomposition enthalpy and activation energy were found to strongly depend on the nitric acid content of the acid-solvates. The studies also indicate that dibutylalkyl and dipentylalkyl phosphonates are also prone to form 'red oil' like substances under extreme conditions and can lead to run-away reactions
Label-Free Potentiometry for Detecting DNA Hybridization Using Peptide Nucleic Acid and DNA Probes

Science.gov (United States)

Goda, Tatsuro; Singi, Ankit Balram; Maeda, Yasuhiro; Matsumoto, Akira; Torimura, Masaki; Aoki, Hiroshi; Miyahara, Yuji

2013-01-01

Peptide nucleic acid (PNA) has outstanding affinity over DNA for complementary nucleic acid sequences by forming a PNA-DNA heterodimer upon hybridization via Watson-Crick base-pairing. To verify whether PNA probes on an electrode surface enhance sensitivity for potentiometric DNA detection or not, we conducted a comparative study on the hybridization of PNA and DNA probes on the surface of a 10-channel gold electrodes microarray. Changes in the charge density as a result of hybridization at the solution/electrode interface on the self-assembled monolayer (SAM)-formed microelectrodes were directly transformed into potentiometric signals using a high input impedance electrometer. The charge readout allows label-free, reagent-less, and multi-parallel detection of target oligonucleotides without any optical assistance. The differences in the probe lengths between 15- to 22-mer dramatically influenced on the sensitivity of the PNA and DNA sensors. Molecular type of the capturing probe did not affect the degree of potential shift. Theoretical model for charged rod-like duplex using the Gouy-Chapman equation indicates the dominant effect of electrostatic attractive forces between anionic DNA and underlying electrode at the electrolyte/electrode interface in the potentiometry. PMID:23435052
Label-Free Potentiometry for Detecting DNA Hybridization Using Peptide Nucleic Acid and DNA Probes

Directory of Open Access Journals (Sweden)

Yuji Miyahara

2013-02-01

Full Text Available Peptide nucleic acid (PNA has outstanding affinity over DNA for complementary nucleic acid sequences by forming a PNA-DNA heterodimer upon hybridization via Watson-Crick base-pairing. To verify whether PNA probes on an electrode surface enhance sensitivity for potentiometric DNA detection or not, we conducted a comparative study on the hybridization of PNA and DNA probes on the surface of a 10-channel gold electrodes microarray. Changes in the charge density as a result of hybridization at the solution/electrode interface on the self-assembled monolayer (SAM-formed microelectrodes were directly transformed into potentiometric signals using a high input impedance electrometer. The charge readout allows label-free, reagent-less, and multi-parallel detection of target oligonucleotides without any optical assistance. The differences in the probe lengths between 15- to 22-mer dramatically influenced on the sensitivity of the PNA and DNA sensors. Molecular type of the capturing probe did not affect the degree of potential shift. Theoretical model for charged rod-like duplex using the Gouy-Chapman equation indicates the dominant effect of electrostatic attractive forces between anionic DNA and underlying electrode at the electrolyte/electrode interface in the potentiometry.
Extraction of uranium from aqueous solution by phosphonic acid-imbedded polyurethane foam

International Nuclear Information System (INIS)

Katragadda, S.; Gesser, H.D.; Chow, A.

1997-01-01

Phenylphosphonic acid was imbedded into the matrix of the polyurethane foam during the fabrication process of the polymer. The extraction of uranium by phosphonic acid-imbedded polyurethane foam and blank polyurethane (i.e., foam without phosphonic acid functional groups) was investigated. Phosphonic acid-imbedded foam showed superior extractability of uranium from solutions with pH = 7.0 ± 1.5 over a wide range of temperatures. (author)

Modulation of mdm2 pre-mRNA splicing by 9-aminoacridine-PNA (peptide nucleic acid) conjugates targeting intron-exon junctions

DEFF Research Database (Denmark)

Shiraishi, Takehiko; Eysturskard, Jonhard; Nielsen, Peter E

2010-01-01

ABSTRACT: BACKGROUND: Modulation of pre-mRNA splicing by antisense molecules is a promising mechanism of action for gene therapeutic drugs. In this study, we have examined the potential of peptide nucleic acid (PNA) 9-aminoacridine conjugates to modulate the pre-mRNA splicing of the mdm2 human ca...
Rapid identification of Staphylococcus aureus and methicillin resistance by flow cytometry using a peptide nucleic acid probe.

Science.gov (United States)

Shrestha, Nabin K; Scalera, Nikole M; Wilson, Deborah A; Brehm-Stecher, Byron; Procop, Gary W

2011-09-01

A total of 56 Staphylococcus aureus isolates incubated for 2 h in the presence or absence of oxacillin were analyzed by flow cytometry after labeling with an S. aureus-specific peptide nucleic acid (PNA) probe. Two defined ratios, the paired signal count ratio (PSCR) and the gate signal count ratio (GSCR), differentiated methicillin-resistant S. aureus (MRSA) and methicillin-susceptible S. aureus (MSSA) with sensitivities of 100% each and specificities of 96% and 100%, respectively.
Nanocomposites of phosphonic-acid-functionalized polyethylenes with inorganic quantum dots.

Science.gov (United States)

Rünzi, Thomas; Baier, Moritz C; Negele, Carla; Krumova, Marina; Mecking, Stefan

2015-01-01

Insertion of diethyl vinyl phosphonates and free vinyl phosphonic acid, respectively, into [(P^O)Pd(Me)(dmso)] ((P^O) = κ(2)-P,O-Ar2PC6H4SO2O with Ar = 2-MeOC6H4) (1-dmso) occurs in a 2,1- as well as 1,2-fashion, to form a four-and a five-membered chelate [(P^O)Pd{κ(2)-C,O-CH(P(O)(OR)2)CH2CH3}] and [(P^O)Pd{κ(2)-C,O-CH2CH(P(O)(OR)2)CH3}] (R = H, Et). No decomposition or other reactions of 1 by free phosphonic acid moieties occur. Copolymerization in a pressure reactor by 1-dmso yields linear random poly(ethylene-co-diethyl vinyl phosphonate) and poly(ethylene-co-vinyl phosphonic acid). In these copolymerizations, reversible coordination of the phosphonate moieties of free monomer as well as chelate formation by incorporated monomer retards chain growth as also evidenced by relative binding studies of diethyl phosphonate towards 1. Post-polymerization emulsification of poly(ethylene-co-vinyl phosphonic acid) together with CdSe/CdS quantum dots (QDs) yields submicron (ca. 50 nm from dynamic light scattering (DLS) and transmission electron microscopy (TEM)) polymer particles with the QDs embedded in the functionalized polyethylene in a nonaggregated fashion. This embedding benefits the fluorescence behavior in terms of continuous emission and life-time as revealed by wide-field fluorescence measurements. These composite particle dispersions are employed as a ″masterbatch" together with an aqueous high density polyethylene (HDPE) dispersion to generate thin films (by spin-coating) and bulk materials (from the melt), respectively, in which the inorganic nanoparticles remain highly disperse. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.
Isosteric phosphonate pyrrolidine-based dinucleoside monophosphate analogues

Czech Academy of Sciences Publication Activity Database

Vaněk, Václav; Buděšínský, Miloš; Kavenová, Ivana; Rinnová, Markéta; Rosenberg, Ivan

2003-01-01

Roč. 22, 5/8 (2003), s. 1065-1067 ISSN 1525-7770. [International Roundtable Nucleosides, Nucleotides and Nucleic Acids /15./. Leuven, 10.09.2002-14.09.2002] R&D Projects: GA ČR GA203/01/1166; GA AV ČR IAA4055101 Institutional research plan: CEZ:AV0Z4055905 Keywords : pyrrolidine-based phosphonate nucleotides * ApA analogues * triplex Subject RIV: CC - Organic Chemistry Impact factor: 0.813, year: 2003
Cathepsin B-sensitive polymers for compartment-specific degradation and nucleic acid release.

Science.gov (United States)

Chu, David S H; Johnson, Russell N; Pun, Suzie H

2012-02-10

Degradable cationic polymers are desirable for in vivo nucleic acid delivery because they offer significantly decreased toxicity over non-degradable counterparts. Peptide linkers provide chemical stability and high specificity for particular endopeptidases but have not been extensively studied for nucleic acid delivery applications. In this work, enzymatically degradable peptide-HPMA copolymers were synthesized by RAFT polymerization of HPMA with methacrylamido-terminated peptide macromonomers, resulting in polymers with low polydispersity and near quantitative incorporation of peptides. Three peptide-HPMA copolymers were evaluated: (i) pHCathK(10), containing peptides composed of the linker phe-lys-phe-leu (FKFL), a substrate of the endosomal/lysosomal endopeptidase cathepsin B, connected to oligo-(L)-lysine for nucleic acid binding, (ii) pHCath(D)K(10), containing the FKFL linker with oligo-(D)-lysine, and (iii) pH(D)Cath(D)K(10), containing all (D) amino acids. Cathepsin B degraded copolymers pHCathK(10) and pHCath(D)K(10) within 1 h while no degradation of pH(D)Cath(D)K(10) was observed. Polyplexes formed with pHCathK(10) copolymers show DNA release by 4 h of treatment with cathepsin B; comparatively, polyplexes formed with pHCath(D)K(10) and pH(D)Cath(D)K(10) show no DNA release within 8 h. Transfection efficiency in HeLa and NIH/3T3 cells were comparable between the copolymers but pHCathK(10) was less toxic. This work demonstrates the successful application of peptide linkers for degradable cationic polymers and DNA release. Copyright Â© 2011 Elsevier B.V. All rights reserved.
Nucleic acids, proteins, and chirality

Science.gov (United States)

Usher, D. A.; Profy, A. T.; Walstrum, S. A.; Needels, M. C.; Bulack, S. C.; Lo, K. M.

1984-01-01

The present investigation is concerned with experimental results related, in one case, to the chirality of nucleotides, and, in another case, to the possibility of a link between the chirality of nucleic acids, and that of peptides. It has been found that aminoacylation of the 'internal' hydroxyl group of a dinucleoside monophosphate can occur stereoselectively. However, this reaction has not yet been made a part of a working peptide synthesis scheme. The formation and cleavage of oligonucleotides is considered. In the event of the formation of a helical complex between the oligonucleotide and the polymer, 1-prime,5-prime-bonds in the oligomer are found to become more resistant towards cleavage. The conditions required for peptide bond formation are examined, taking into account the known structures of RNA and possible mechanisms for prebiotic peptide bond formation. The possibility is considered that the 2-prime,5-prime-internucleotide linkage could have played an important part in the early days of biological peptide synthesis.
Synthesis and optical properties of pyrrolidinyl peptide nucleic acid carrying a clicked Nile red label

Directory of Open Access Journals (Sweden)

Nattawut Yotapan

2014-09-01

Full Text Available DNA or its analogues with an environment-sensitive fluorescent label are potentially useful as a probe for studying the structure and dynamics of nucleic acids. In this work, pyrrolidinyl peptide nucleic acid (acpcPNA was labeled at its backbone with Nile red, a solvatochromic benzophenoxazine dye, by means of click chemistry. The optical properties of the Nile red-labeled acpcPNA were investigated by UV–vis and fluorescence spectroscopy in the absence and in the presence of DNA. In contrast to the usual quenching observed in Nile red-labeled DNA, the hybridization with DNA resulted in blue shifting and an enhanced fluorescence regardless of the neighboring bases. More pronounced blue shifts and fluorescence enhancements were observed when the DNA target carried a base insertion in close proximity to the Nile red label. The results indicate that the Nile red label is located in a more hydrophobic environment in acpcPNA–DNA duplexes than in the single-stranded acpcPNA. The different fluorescence properties of the acpcPNA hybrids of complementary DNA and DNA carrying a base insertion are suggestive of different interactions between the Nile red label and the duplexes.
Sequence-specific inhibition of duck hepatitis B virus reverse transcription by peptide nucleic acids (PNA)

DEFF Research Database (Denmark)

Robaczewska, Magdalena; Narayan, Ramamurthy; Seigneres, Beatrice

2005-01-01

BACKGROUND/AIMS: Peptide nucleic acids (PNAs) appear as promising new antisense agents, that have not yet been examined as hepatitis B virus (HBV) inhibitors. Our aim was to study the ability of PNAs targeting the duck HBV (DHBV) encapsidation signal epsilon to inhibit reverse transcription (RT...... in primary duck hepatocytes (PDH). RESULTS: Both PNAs reproducibly inhibited DHBV RT in a dose-dependent manner with IC(50) of 10nM, whereas up to 600-fold higher concentration of S-ODNs was required for similar inhibition. The PNA targeting the bulge and upper stem of epsilon appeared as more efficient RT...
On the stereochemistry of C3'-O-P-CH2-O-C4'' phosphonate internucleotide bond, a phosphate isostere

Czech Academy of Sciences Publication Activity Database

Točík, Zdeněk; Buděšínský, Miloš; Barvík Jr., I.; Rosenberg, Ivan

-, č. 52 (2008), s. 427-428 ISSN 0261-3166. [Joint Symposium of the International Roundtable on Nucleosides, Nucleotides and Nucleic Acids /18./ and the International Symposium on Nucleic Acid Chemistry /35./. Kyoto, 08.09.2008-12.09.2008] R&D Projects: GA MŠk(CZ) LC06061 Institutional research plan: CEZ:AV0Z40550506 Keywords : phosphonate oligonucleotide stereochemistry Subject RIV: CC - Organic Chemistry
Tethered particle analysis of supercoiled circular DNA using peptide nucleic acid handles

DEFF Research Database (Denmark)

Norregaard, Kamilla; Andersson, Magnus; Nielsen, Peter Eigil

2014-01-01

This protocol describes how to monitor individual naturally supercoiled circular DNA plasmids bound via peptide nucleic acid (PNA) handles between a bead and a surface. The protocol was developed for single-molecule investigation of the dynamics of supercoiled DNA, and it allows the investigation...... of both the dynamics of the molecule itself and of its interactions with a regulatory protein. Two bis-PNA clamps designed to bind with extremely high affinity to predetermined homopurine sequence sites in supercoiled DNA are prepared: one conjugated with digoxigenin for attachment to an anti......-digoxigenin-coated glass cover slide, and one conjugated with biotin for attachment to a submicron-sized streptavidin-coated polystyrene bead. Plasmids are constructed, purified and incubated with the PNA handles. The dynamics of the construct is analyzed by tracking the tethered bead using video microscopy: less...
Rapid Identification of Staphylococcus aureus and Methicillin Resistance by Flow Cytometry Using a Peptide Nucleic Acid Probe ▿

Science.gov (United States)

Shrestha, Nabin K.; Scalera, Nikole M.; Wilson, Deborah A.; Brehm-Stecher, Byron; Procop, Gary W.

2011-01-01

A total of 56 Staphylococcus aureus isolates incubated for 2 h in the presence or absence of oxacillin were analyzed by flow cytometry after labeling with an S. aureus-specific peptide nucleic acid (PNA) probe. Two defined ratios, the paired signal count ratio (PSCR) and the gate signal count ratio (GSCR), differentiated methicillin-resistant S. aureus (MRSA) and methicillin-susceptible S. aureus (MSSA) with sensitivities of 100% each and specificities of 96% and 100%, respectively. PMID:21795508
Reaction of acid esters of methylenebis(phosphonous acid) with carbonyl compounds

International Nuclear Information System (INIS)

Novikova, Z.S.; Odinets, I.L.; Lutsenko, I.F.

1987-01-01

The reaction of methylenebis(phosphonites) containing two hydrophosphoryl groupings with aliphatic and aromatic aldehydes and ketones in the presence of alkali metal fluorides leads to methylenebis(α-hydroxyalkylphosphinates). The reaction of methylenebis(phosphonites) containing one hydrophosphoryl groupings with carbonyl compounds in the presence of alkali metal fluorides proceeds with the formation of a new type of heterocyclic phosphorus compound, viz., 1,2λ 3 ,4λ 5 -oxadiphospholanes. The reaction of acid esters of methylenebis(phosphonous) acid with carbonyl compounds in the presence of alkali metal alkoxides or a tertiary amine is accompanied by phosphinate-phosphonate rearrangement of the intermediately formed α-hydroxylalkylphosphinates
Anionic magnetite nanoparticle conjugated with pyrrolidinyl peptide nucleic acid for DNA base discrimination

International Nuclear Information System (INIS)

Khadsai, Sudarat; Rutnakornpituk, Boonjira; Vilaivan, Tirayut; Nakkuntod, Maliwan; Rutnakornpituk, Metha

2016-01-01

Magnetite nanoparticles (MNPs) were surface modified with anionic poly(N-acryloyl glycine) (PNAG) and streptavidin for specific interaction with biotin-conjugated pyrrolidinyl peptide nucleic acid (PNA). Hydrodynamic size (D h ) of PNAG-grafted MNPs varied from 334 to 496 nm depending on the loading ratio of the MNP to NAG in the reaction. UV–visible and fluorescence spectrophotometries were used to confirm the successful immobilization of streptavidin and PNA on the MNPs. About 291 pmol of the PNA/mg MNP was immobilized on the particle surface. The PNA-functionalized MNPs were effectively used as solid supports to differentiate between fully complementary and non-complementary/single-base mismatch DNA using the PNA probe. These novel anionic MNPs can be efficiently applicable for use as a magnetically guidable support for DNA base discrimination.Graphical Abstract
The HIV-1 transcriptional activator Tat has potent nucleic acid chaperoning activities in vitro.

Science.gov (United States)

Kuciak, Monika; Gabus, Caroline; Ivanyi-Nagy, Roland; Semrad, Katharina; Storchak, Roman; Chaloin, Olivier; Muller, Sylviane; Mély, Yves; Darlix, Jean-Luc

2008-06-01

The human immunodeficiency virus type 1 (HIV-1) is a primate lentivirus that causes the acquired immunodeficiency syndrome (AIDS). In addition to the virion structural proteins and enzyme precursors, that are Gag, Env and Pol, HIV-1 encodes several regulatory proteins, notably a small nuclear transcriptional activator named Tat. The Tat protein is absolutely required for virus replication since it controls proviral DNA transcription to generate the full-length viral mRNA. Tat can also regulate mRNA capping and splicing and was recently found to interfere with the cellular mi- and siRNA machinery. Because of its extensive interplay with nucleic acids, and its basic and disordered nature we speculated that Tat had nucleic acid-chaperoning properties. This prompted us to examine in vitro the nucleic acid-chaperoning activities of Tat and Tat peptides made by chemical synthesis. Here we report that Tat has potent nucleic acid-chaperoning activities according to the standard DNA annealing, DNA and RNA strand exchange, RNA ribozyme cleavage and trans-splicing assays. The active Tat(44-61) peptide identified here corresponds to the smallest known sequence with DNA/RNA chaperoning properties.
Anionic magnetite nanoparticle conjugated with pyrrolidinyl peptide nucleic acid for DNA base discrimination

Energy Technology Data Exchange (ETDEWEB)

Khadsai, Sudarat; Rutnakornpituk, Boonjira [Naresuan University, Department of Chemistry and Center of Excellence in Biomaterials, Faculty of Science (Thailand); Vilaivan, Tirayut [Chulalongkorn University, Department of Chemistry, Organic Synthesis Research Unit, Faculty of Science (Thailand); Nakkuntod, Maliwan [Naresuan University, Department of Biology, Faculty of Science (Thailand); Rutnakornpituk, Metha, E-mail: methar@nu.ac.th [Naresuan University, Department of Chemistry and Center of Excellence in Biomaterials, Faculty of Science (Thailand)

2016-09-15

Magnetite nanoparticles (MNPs) were surface modified with anionic poly(N-acryloyl glycine) (PNAG) and streptavidin for specific interaction with biotin-conjugated pyrrolidinyl peptide nucleic acid (PNA). Hydrodynamic size (D{sub h}) of PNAG-grafted MNPs varied from 334 to 496 nm depending on the loading ratio of the MNP to NAG in the reaction. UV–visible and fluorescence spectrophotometries were used to confirm the successful immobilization of streptavidin and PNA on the MNPs. About 291 pmol of the PNA/mg MNP was immobilized on the particle surface. The PNA-functionalized MNPs were effectively used as solid supports to differentiate between fully complementary and non-complementary/single-base mismatch DNA using the PNA probe. These novel anionic MNPs can be efficiently applicable for use as a magnetically guidable support for DNA base discrimination.Graphical Abstract.
Portable nucleic acid thermocyclers.

Science.gov (United States)

Almassian, David R; Cockrell, Lisa M; Nelson, William M

2013-11-21

A nucleic acid thermal cycler is considered to be portable if it is under ten pounds, easily carried by one individual, and battery powered. Nucleic acid amplification includes both polymerase chain reaction (e.g. PCR, RT-PCR) and isothermal amplification (e.g. RPA, HDA, LAMP, NASBA, RCA, ICAN, SMART, SDA). There are valuable applications for portable nucleic acid thermocyclers in fields that include clinical diagnostics, biothreat detection, and veterinary testing. A system that is portable allows for the distributed detection of targets at the point of care and a reduction of the time from sample to answer. The designer of a portable nucleic acid thermocycler must carefully consider both thermal control and the detection of amplification. In addition to thermal control and detection, the designer may consider the integration of a sample preparation subsystem with the nucleic acid thermocycler. There are a variety of technologies that can achieve accurate thermal control and the detection of nucleic acid amplification. Important evaluation criteria for each technology include maturity, power requirements, cost, sensitivity, speed, and manufacturability. Ultimately the needs of a particular market will lead to user requirements that drive the decision between available technologies.
Locked nucleic acid

DEFF Research Database (Denmark)

Jepsen, Jan Stenvang; Sørensen, Mads D; Wengel, Jesper

2004-01-01

Locked nucleic acid (LNA) is a class of nucleic acid analogs possessing very high affinity and excellent specificity toward complementary DNA and RNA, and LNA oligonucleotides have been applied as antisense molecules both in vitro and in vivo. In this review, we briefly describe the basic...
Fischer carbene mediated covalent grafting of a peptide nucleic acid on gold surfaces and IR optical detection of DNA hybridization with a transition metalcarbonyl label

Czech Academy of Sciences Publication Activity Database

Srivastava, P.; Ghasemi, M.; Ray, N.; Sarkar, A.; Kocábová, Jana; Lachmanová, Štěpánka; Hromadová, Magdaléna; Boujday, S.; Cauteruccio, S.; Thakare, P.; Licandro, E.; Fosse, C.; Salmain, M.

2016-01-01

Roč. 385, NOV 2016 (2016), s. 47-55 ISSN 0169-4332 R&D Projects: GA ČR(CZ) GA14-05180S Institutional support: RVO:61388955 Keywords : peptide nucleic acid * self-assembled monolayer * genosensor Subject RIV: CG - Electrochemistry Impact factor: 3.387, year: 2016
Correction of a splice-site mutation in the beta-globin gene stimulated by triplex-forming peptide nucleic acids

DEFF Research Database (Denmark)

Chin, Joanna Y; Kuan, Jean Y; Lonkar, Pallavi S

2008-01-01

Splice-site mutations in the beta-globin gene can lead to aberrant transcripts and decreased functional beta-globin, causing beta-thalassemia. Triplex-forming DNA oligonucleotides (TFOs) and peptide nucleic acids (PNAs) have been shown to stimulate recombination in reporter gene loci in mammalian...... DNA fragments, can promote single base-pair modification at the start of the second intron of the beta-globin gene, the site of a common thalassemia-associated mutation. This single base pair change was detected by the restoration of proper splicing of transcripts produced from a green fluorescent...
Molecular Structure of Nucleic Acids

Indian Academy of Sciences (India)

Molecular Structure of Nucleic Acids. A Structure for Deoxyribose Nucleic Acid. J. D. Watson and F. H. C. Crick. Medical Research Council Unit for the Study of the Molecular Structure of Biological. Systems, Cavendish Laboratory, Cambridge. April 2. We wish to suggest a structure for the salt of deoxyribose nucleic acid ...

Quantitative thermodynamic predication of interactions between nucleic acid and non-nucleic acid species using Microsoft excel.

Science.gov (United States)

Zou, Jiaqi; Li, Na

2013-09-01

Proper design of nucleic acid sequences is crucial for many applications. We have previously established a thermodynamics-based quantitative model to help design aptamer-based nucleic acid probes by predicting equilibrium concentrations of all interacting species. To facilitate customization of this thermodynamic model for different applications, here we present a generic and easy-to-use platform to implement the algorithm of the model with Microsoft(®) Excel formulas and VBA (Visual Basic for Applications) macros. Two Excel spreadsheets have been developed: one for the applications involving only nucleic acid species, the other for the applications involving both nucleic acid and non-nucleic acid species. The spreadsheets take the nucleic acid sequences and the initial concentrations of all species as input, guide the user to retrieve the necessary thermodynamic constants, and finally calculate equilibrium concentrations for all species in various bound and unbound conformations. The validity of both spreadsheets has been verified by comparing the modeling results with the experimental results on nucleic acid sequences reported in the literature. This Excel-based platform described here will allow biomedical researchers to rationalize the sequence design of nucleic acid probes using the thermodynamics-based modeling even without relevant theoretical and computational skills. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.
The solid-state structures of organic salts formed by calix[4]arene dihydroxyphosphonic acid with nucleic bases cations: adeninium, cytosinium, guaninium and uracilium

KAUST Repository

Shkurenko, Aleksander

2018-02-19

Calix[4]arene dihydroxyphosphonic acid has been demonstrated to possess an interesting range of biological properties, including atypical anti-cancer activity. The robustness of calix[4]arene dihydroxyphosphonic acid and its ubiquitous dimeric motif offers perspectives for pre-defined solid state complexation with small molecules. In the current article we describe co-crystals (organic salts) of calix[4]arene dihydroxyphosphonic acid with four nucleic base cations: adeninium, cytosinium, guaninium and uracilium. A number of characteristic interactions between the components in the four co-crystals are pointed out also using the Hirshfeld surface analysis. All the four co-crystals are based on layers of calix[4]arene dimers, alternating with layers of nucleic acid molecules. Two of the reported crystal structures (cytosinium and guaninium) are 1D channel-type structures, while the two others (adeninium and uracilium) represent 2D channel-type structures. In three out of four reported structures, interactions between the cations of nucleic bases are present generating 1D chains of cations. A constant motif is that the nucleic base is present in a type of cavity formed by one aromatic ring and a phosphonic acid moiety.
Application of Peptide Nucleic Acid-based Assays Toward Detection of Somatic Mosaicism

Directory of Open Access Journals (Sweden)

Christopher S Hong

2016-01-01

Full Text Available Peptide nucleic acids (PNAs are synthetic oligonucleotides with many applications. Compared with DNA, PNAs bind their complementary DNA strand with higher specificity and strength, an attribute that can make it an effective polymerase chain reaction clamp. A growing body of work has demonstrated the utility of PNAs in detecting low levels of mutant DNA, particularly in the detection of circulating mutated tumor cells in the peripheral blood. The PNA-based assay has greater sensitivity than direct sequencing and is significantly more affordable and rapid than next-generation deep sequencing. We have previously demonstrated that PNAs can successfully detect somatic mosaicism in patients with suspected disease phenotypes. In this report, we detail our methodology behind PNA design and application. We describe our protocol for optimizing the PNA for sequencing use and for determining the sensitivity of the PNA-based assay. Lastly, we discuss the potential applications of our assay for future laboratory and clinical purposes and highlight the role of PNAs in the detection of somatic mosaicism.
Preparation, characterization and application of N-methylene phosphonic acid chitosan grafted magnesia–zirconia stationary phase

Energy Technology Data Exchange (ETDEWEB)

Wang, Qing; Chen, Jie; Huang, Kun; Zhang, Xin; Xu, Li [Tongji School of Pharmacy, Huazhong University of Science and Technology, Wuhan 430030 (China); Shi, Zhi-guo, E-mail: shizg@whu.edu.cn [Department of Chemistry, Wuhan University, Wuhan 430072 (China)

2015-01-07

Highlights: • N-methylene phosphonic acid chitosan grafted MgO–ZrO{sub 2} was prepared. • It exhibited superior HILIC chromatographic performance to the bare MgO–ZrO{sub 2}. • Monosaccharides, phospholipids and peptides were successfully separated. • It was a promising HILIC stationary phase. - Abstract: A hydrophilic stationary phase (SP) was prepared through grafting N-methylene phosphonic acid chitosan on magnesia–zirconia particles (P-CTS-MgO–ZrO{sub 2}) via Lewis acid–base interaction. The resulting material was characterized by thermogravimetric analysis, Fourier transform infrared spectroscopy, scanning electron microscope and nitrogen adsorption analysis. The chromatographic performance of P-CTS-MgO–ZrO{sub 2} was systemically evaluated by studying effect of acetonitrile content, pH and buffer concentration in the mobile phase. The results demonstrated that the novel SP provided hydrophilic, electrostatic-repulsion and ion-exchange interactions. Compared to the bare MgO–ZrO{sub 2}, P-CTS-MgO–ZrO{sub 2} exhibited superior peak shape, reasonable resolution and reduced analysis time in separation of basic analytes. Besides, remarkable resolving power of acids, i.e. six non-steroidal anti-inflammatory drugs which failed to be eluted from the bare MgO–ZrO{sub 2}, was obtained with the theoretical plate number (N/m) of 4653–31313, asymmetry factor <1.21 and the resolution of 1.6–3.4. Finally, P-CTS-MgO–ZrO{sub 2} SP was applied to separate monosaccharides, phospholipids and peptides. P-CTS-MgO–ZrO{sub 2} was a promising hydrophilic SP for wide applications.
Identifying a base in a nucleic acid

Science.gov (United States)

Fodor, Stephen P. A.; Lipshutz, Robert J.; Huang, Xiaohua

2005-02-08

Devices and techniques for hybridization of nucleic acids and for determining the sequence of nucleic acids. Arrays of nucleic acids are formed by techniques, preferably high resolution, light-directed techniques. Positions of hybridization of a target nucleic acid are determined by, e.g., epifluorescence microscopy. Devices and techniques are proposed to determine the sequence of a target nucleic acid more efficiently and more quickly through such synthesis and detection techniques.
Cellular delivery and antisense effects of peptide nucleic acid conjugated to polyethyleneimine via disulfide linkers

DEFF Research Database (Denmark)

Berthold, Peter R; Shiraishi, Takehiko; Nielsen, Peter E

2010-01-01

Peptide nucleic acid (PNA) is potentially an attractive antisense and antigene agent for which more efficient cellular delivery systems are still warranted. The cationic polymer polyethylenimine (PEI) is commonly used for cellular transfection of DNA and RNA complexes, but is not readily applicable...... moiety) and further reacted this with a cysteine PNA. The level of modification was determined spectrophotometrically with high accuracy, and the PNA transfection efficiency of the conjugates was evaluated in an antisense luciferase splice-correction assay using HeLa pLuc705 cells. We find that PEI...... is an efficient vector for PNA delivery yielding significantly higher (up to 10-fold) antisense activity than an analogous PNA-octaarginine conjugate, even in the presence of chloroquine, which only slightly enhances the PEI-PNA activity. The PEI-PEG conjugates are preferred due to lower acute cellular toxicity...
Triptycene: A Nucleic Acid Three-Way Junction Binder Scaffold

Science.gov (United States)

Yoon, Ina

Nucleic acids play a critical role in many biological processes such as gene regulation and replication. The development of small molecules that modulate nucleic acids with sequence or structure specificity would provide new strategies for regulating disease states at the nucleic acid level. However, this remains challenging mainly because of the nonspecific interactions between nucleic acids and small molecules. Three-way junctions are critical structural elements of nucleic acids. They are present in many important targets such as trinucleotide repeat junctions related to Huntington's disease, a temperature sensor sigma32 in E. coli, Dengue virus, and HIV. Triptycene-derived small molecules have been shown to bind to nucleic acid three-way junctions, resulting from their shape complementary. To develop a better understanding of designing molecules for targeting different junctions, a rapid screening of triptycene-based small molecules is needed. We envisioned that the installation of a linker at C9 position of the bicyclic core would allow for a rapid solid phase diversification. To achieve this aim, we synthesized 9-substituted triptycene scaffolds by using two different synthetic routes. The first synthetic route installed the linker from the amidation reaction between carboxylic acid at C9 position of the triptycene and an amine linker, beta-alanine ethyl ester. This new 9-substituted triptycene scaffold was then attached to a 2-chlorotrityl chloride resin for solid-phase diversification. This enabled a rapid diversification and an easy purification of mono-, di-, and tri-peptide triptycene derivatives. The binding affinities of these compounds were investigated towards a (CAG)˙(CTG) trinucleotide repeat junction. In the modified second synthetic route, we utilized a combined Heck coupling/benzyne Diels-Alder strategy. This improved synthetic strategy reduced the number of steps and total reaction times, increased the overall yield, improved solubilities of
Inducible Alkylation of DNA by a Quinone Methide-Peptide Nucleic Acid Conjugate†

Science.gov (United States)

Liu, Yang; Rokita, Steven E.

2012-01-01

The reversibility of alkylation by a quinone methide intermediate (QM) avoids the irreversible consumption that plagues most reagents based on covalent chemistry and allows for site specific reaction that is controlled by the thermodynamics rather than kinetics of target association. This characteristic was originally examined with an oligonucleotide QM conjugate but broad application depends on alternative derivatives that are compatible with a cellular environment. Now, a peptide nucleic acid (PNA) derivative has been constructed and shown to exhibit an equivalent ability to delivery the reactive QM in a controlled manner. This new conjugate demonstrates high selectivity for a complementary sequence of DNA even when challenged with an alternative sequence containing a single T/T mismatch. Alkylation of non-complementary sequences is only possible when a template strand is present to co-localize the conjugate and its target. For efficient alkylation in this example, a single-stranded region of the target is required adjacent to the QM conjugate. Most importantly, the intrastrand self adducts formed between the PNA and its attached QM remained active and reversible over more than eight days in aqueous solution prior to reaction with a chosen target added subsequently. PMID:22243337
Preliminary assessment of developmental toxicity of Perfluorinated Phosphonic Acid in mice

Science.gov (United States)

Perfluorinated phosphonic acids (PFPAs) are a third member of the perfluoroalkyl acid (PFAA) family, and are structurally similar to the perfluoroalkyl sulfonates and perfluoroalkyl carboxylates. These emerging chemicals have recently been detected in the environment, particularl...
Nucleic acid interactions with pyrite surfaces

International Nuclear Information System (INIS)

Mateo-Marti, E.; Briones, C.; Rogero, C.; Gomez-Navarro, C.; Methivier, Ch.; Pradier, C.M.; Martin-Gago, J.A.

2008-01-01

The study of the interaction of nucleic acid molecules with mineral surfaces is a field of growing interest in organic chemistry, origin of life, material science and biotechnology. We have characterized the adsorption of single-stranded peptide nucleic acid (ssPNA) on a natural pyrite surface, as well as the further adsorption of ssDNA on a PNA-modified pyrite surface. The characterization has been performed by means of reflection absorption infrared spectroscopy (RAIRS), atomic force microscopy (AFM) and X-ray photoemission spectroscopy (XPS) techniques. The N(1s) and S(2p) XPS core level peaks of PNA and PNA + DNA have been decomposed in curve-components that we have assigned to different chemical species. RAIRS spectra recorded for different concentrations show the presence of positive and negative adsorption bands, related to the semiconducting nature of the surface. The combination of the information gathered by these techniques confirms that PNA adsorbs on pyrite surface, interacting through nitrogen-containing groups of the nucleobases and the iron atoms of the surface, instead of the thiol group of the molecule. The strong PNA/pyrite interaction inhibits further hybridization of PNA with complementary ssDNA, contrary to the behavior reported on gold surfaces
Nucleic acid interactions with pyrite surfaces

Energy Technology Data Exchange (ETDEWEB)

Mateo-Marti, E. [Centro de Astrobiologia (CSIC-INTA), Ctra. Ajalvir, Km. 4, 28850-Torrejon de Ardoz, Madrid (Spain)], E-mail: mateome@inta.es; Briones, C.; Rogero, C. [Centro de Astrobiologia (CSIC-INTA), Ctra. Ajalvir, Km. 4, 28850-Torrejon de Ardoz, Madrid (Spain); Gomez-Navarro, C. [Instituto de Ciencia de Materiales de Madrid (CSIC), Cantoblanco, 28049-Madrid (Spain); Methivier, Ch.; Pradier, C.M. [Laboratoire de Reactivite de Surface, UMR CNRS 7609. Universite Pierre et Marie Curie, 4, Pl Jussieu, 75005-Paris (France); Martin-Gago, J.A. [Centro de Astrobiologia (CSIC-INTA), Ctra. Ajalvir, Km. 4, 28850-Torrejon de Ardoz, Madrid (Spain); Instituto de Ciencia de Materiales de Madrid (CSIC), Cantoblanco, 28049-Madrid (Spain)

2008-09-03

The study of the interaction of nucleic acid molecules with mineral surfaces is a field of growing interest in organic chemistry, origin of life, material science and biotechnology. We have characterized the adsorption of single-stranded peptide nucleic acid (ssPNA) on a natural pyrite surface, as well as the further adsorption of ssDNA on a PNA-modified pyrite surface. The characterization has been performed by means of reflection absorption infrared spectroscopy (RAIRS), atomic force microscopy (AFM) and X-ray photoemission spectroscopy (XPS) techniques. The N(1s) and S(2p) XPS core level peaks of PNA and PNA + DNA have been decomposed in curve-components that we have assigned to different chemical species. RAIRS spectra recorded for different concentrations show the presence of positive and negative adsorption bands, related to the semiconducting nature of the surface. The combination of the information gathered by these techniques confirms that PNA adsorbs on pyrite surface, interacting through nitrogen-containing groups of the nucleobases and the iron atoms of the surface, instead of the thiol group of the molecule. The strong PNA/pyrite interaction inhibits further hybridization of PNA with complementary ssDNA, contrary to the behavior reported on gold surfaces.
Selective extraction of hydrocarbons, phosphonates and phosphonic acids from soils by successive supercritical fluid and pressurized liquid extractions.

Science.gov (United States)

Chaudot, X; Tambuté, A; Caude, M

2000-01-14

Hydrocarbons, dialkyl alkylphosphonates and alkyl alkylphosphonic acids are selectively extracted from spiked soils by successive implementation of supercritical carbon dioxide, supercritical methanol-modified carbon dioxide and pressurized water. More than 95% of hydrocarbons are extracted during the first step (pure supercritical carbon dioxide extraction) whereas no organophosphorus compound is evidenced in this first extract. A quantitative extraction of phosphonates is achieved during the second step (methanol-modified supercritical carbon dioxide extraction). Polar phosphonic acids are extracted during a third step (pressurized water extraction) and analyzed by gas chromatography under methylated derivatives (diazomethane derivatization). Global recoveries for these compounds are close to 80%, a loss of about 20% occurring during the derivatization process (co-evaporation with solvent). The developed selective extraction method was successfully applied to a soil sample during an international collaborative exercise.
Nucleic acid-binding glycoproteins which solubilize nucleic acids in dilute acid: re-examination of the Ustilago maydis glycoproteins

Energy Technology Data Exchange (ETDEWEB)

Unrau, P.; Champ, D.R.; Young, J.L.; Grant, C.E.

1980-01-01

Holloman reported the isolation from Ustilago maydis of a glycoprotein which prevented the precipitation of nucleic acids in cold 5% trichloroacetic acid. Two glycoprotein fractions from U. maydis with this nucleic acid-solubilizing activity were isolated in our laboratory using improved purification procedures. The activity was not due to nuclease contamination. The glycoproteins are distinguished by: their ability to bind to concanavalin A-Sepharose; their differential binding to double- and single-stranded deoxyribonucleic acid, and to ribonucleic acid; their molecular weights (46,000 and 69,000); and the relative amounts present in growing versus nongrowing cells. Both fractions required sulfhydryl-reducing conditions for optimal yields, specific activity, and stability. Nucleic acid binding was cooperative, the minimum number of glycoproteins required to make a native T7 DNA molecule soluble in dilute acid being estimated at 2 and 15, respectively.
High temperature proton exchange membranes prepared from epoxycyclohexylethyltrimethoxysilane and amino trimethylene phosphonic acid as anhydrous proton conductors

International Nuclear Information System (INIS)

Chen, Cheng; Shen, Chunhui; Kong, Gengjin; Gao, Shanjun

2013-01-01

High temperature anhydrous proton exchange membranes based on phosphonic acid were prepared from epoxycyclohexylethyltrimethoxysilane (EHTMS) and amino trimethylene phosphonic acid (ATMP) by sol–gel process. The structures and properties of membranes with different phosphonic acid content were extensively characterized by FTIR, TG-DSC and XRD. Their proton conductivity under dry condition was also investigated under different temperature. The results show that the proton conductivity of the prepared membranes strongly depends on temperature, and the proton conductivity ranges from 8.81 × 10 −5 S cm −1 at 20 °C to 4.65 × 10 −2 S cm −1 at 140 °C under anhydrous condition. It indicates that the increasing temperature is favorable for congregating of the grafted–PO 3 H 2 and increasing of the proton mobility. In addition, from the results of AFM images, it was confirmed that the continuous distribution of phosphonic acid groups is favorable for the formation of the proton transport channel, which can significantly enhance the proton conductivity of the membranes. Highlights: ► Hybrid membranes of Epoxycyclohexylethyltrimethoxysilane and Amino trimethylene phosphonic acid. ► The proton conductivity is 4.65 × 10 −2 S cm −1 at 140 °C under anhydrous condition. ► Continuous uniform distributions of phosphonic acid groups can be observed by AFM. ► There could be hydrogen bond network within high temperature membranes
High temperature proton exchange membranes prepared from epoxycyclohexylethyltrimethoxysilane and amino trimethylene phosphonic acid as anhydrous proton conductors

Energy Technology Data Exchange (ETDEWEB)

Chen, Cheng [Department of Polymer Materials and Engineering, School of Material Science and Engineering, Wuhan University of Technology, No. 122 Luoshi Road, Wuhan 430070 (China); Shen, Chunhui, E-mail: shenchunhui@whut.edu.cn [Department of Polymer Materials and Engineering, School of Material Science and Engineering, Wuhan University of Technology, No. 122 Luoshi Road, Wuhan 430070 (China); Kong, Gengjin; Gao, Shanjun [Department of Polymer Materials and Engineering, School of Material Science and Engineering, Wuhan University of Technology, No. 122 Luoshi Road, Wuhan 430070 (China)

2013-06-15

High temperature anhydrous proton exchange membranes based on phosphonic acid were prepared from epoxycyclohexylethyltrimethoxysilane (EHTMS) and amino trimethylene phosphonic acid (ATMP) by sol–gel process. The structures and properties of membranes with different phosphonic acid content were extensively characterized by FTIR, TG-DSC and XRD. Their proton conductivity under dry condition was also investigated under different temperature. The results show that the proton conductivity of the prepared membranes strongly depends on temperature, and the proton conductivity ranges from 8.81 × 10{sup −5} S cm{sup −1} at 20 °C to 4.65 × 10{sup −2} S cm{sup −1} at 140 °C under anhydrous condition. It indicates that the increasing temperature is favorable for congregating of the grafted–PO{sub 3}H{sub 2} and increasing of the proton mobility. In addition, from the results of AFM images, it was confirmed that the continuous distribution of phosphonic acid groups is favorable for the formation of the proton transport channel, which can significantly enhance the proton conductivity of the membranes. Highlights: ► Hybrid membranes of Epoxycyclohexylethyltrimethoxysilane and Amino trimethylene phosphonic acid. ► The proton conductivity is 4.65 × 10{sup −2} S cm{sup −1} at 140 °C under anhydrous condition. ► Continuous uniform distributions of phosphonic acid groups can be observed by AFM. ► There could be hydrogen bond network within high temperature membranes.
Tethered particle analysis of supercoiled circular DNA using peptide nucleic acid handles.

Science.gov (United States)

Norregaard, Kamilla; Andersson, Magnus; Nielsen, Peter Eigil; Brown, Stanley; Oddershede, Lene B

2014-09-01

This protocol describes how to monitor individual naturally supercoiled circular DNA plasmids bound via peptide nucleic acid (PNA) handles between a bead and a surface. The protocol was developed for single-molecule investigation of the dynamics of supercoiled DNA, and it allows the investigation of both the dynamics of the molecule itself and of its interactions with a regulatory protein. Two bis-PNA clamps designed to bind with extremely high affinity to predetermined homopurine sequence sites in supercoiled DNA are prepared: one conjugated with digoxigenin for attachment to an anti-digoxigenin-coated glass cover slide, and one conjugated with biotin for attachment to a submicron-sized streptavidin-coated polystyrene bead. Plasmids are constructed, purified and incubated with the PNA handles. The dynamics of the construct is analyzed by tracking the tethered bead using video microscopy: less supercoiling results in more movement, and more supercoiling results in less movement. In contrast to other single-molecule methodologies, the current methodology allows for studying DNA in its naturally supercoiled state with constant linking number and constant writhe. The protocol has potential for use in studying the influence of supercoils on the dynamics of DNA and its associated proteins, e.g., topoisomerase. The procedure takes ~4 weeks.
Efficiency of cellular delivery of antisense peptide nucleic acid by electroporation depends on charge and electroporation geometry

DEFF Research Database (Denmark)

Joergensen, Mette; Agerholm-Larsen, Birgit; Nielsen, Peter E

2011-01-01

Electroporation is potentially a very powerful technique for both in vitro cellular and in vivo drug delivery, particularly relating to oligonucleotides and their analogs for genetic therapy. Using a sensitive and quantitative HeLa cell luciferase RNA interference mRNA splice correction assay...... with a functional luciferase readout, we demonstrate that parameters such as peptide nucleic acid (PNA) charge and the method of electroporation have dramatic influence on the efficiency of productive delivery. In a suspended cell electroporation system (cuvettes), a positively charged PNA (+8) was most efficiently...... transferred, whereas charge neutral PNA was more effective in a microtiter plate electrotransfer system for monolayer cells. Surprisingly, a negatively charged (-23) PNA did not show appreciable activity in either system. Findings from the functional assay were corroborated by pulse parameter variations...
A Tat-grafted anti-nucleic acid antibody acquires nuclear-localization property and a preference for TAR RNA

International Nuclear Information System (INIS)

Jeong, Jong-Geun; Kim, Dong-Sik; Kim, Yong-Sung; Kwon, Myung-Hee

2011-01-01

Highlights: → We generate ' H3 Tat-3D8' by grafting Tat 48-60 peptide to VH CDR of 3D8 scFv antibody. → H3 Tat-3D8 antibody retains nucleic acid binding and hydrolyzing activities. → H3 Tat-3D8 acquires a preference for TAR RNA structure. → Properties of Tat 48-60 is transferred to an antibody via Tat-grafting into a CDR. -- Abstract: The 3D8 single chain variable fragment (3D8 scFv) is an anti-nucleic acid antibody that can hydrolyze nucleic acids and enter the cytosol of cells without reaching the nucleus. The Tat peptide, derived from the basic region of the HIV-1 Tat protein, translocates to cell nuclei and has TAR RNA binding activity. In this study, we generated a Tat-grafted antibody ( H3 Tat-3D8) by replacing complementarity-determining region 3 (CDR3) within the VH domain of the 3D8 scFv with a Tat 48-60 peptide (GRKKRRQRRRPPQ). H3 Tat-3D8 retained the DNA-binding and DNA-hydrolyzing activity of the scFv, and translocated to the nuclei of HeLa cells and preferentially recognized TAR RNA. Thus, the properties associated with the Tat peptide were transferred to the antibody via Tat-grafting without loss of the intrinsic DNA-binding and hydrolyzing activities of the 3D8 scFv antibody.
Use of Nucleic Acid Analogs for the Study of Nucleic Acid Interactions

Directory of Open Access Journals (Sweden)

Shu-ichi Nakano

2011-01-01

Full Text Available Unnatural nucleosides have been explored to expand the properties and the applications of oligonucleotides. This paper briefly summarizes nucleic acid analogs in which the base is modified or replaced by an unnatural stacking group for the study of nucleic acid interactions. We also describe the nucleoside analogs of a base pair-mimic structure that we have examined. Although the base pair-mimic nucleosides possess a simplified stacking moiety of a phenyl or naphthyl group, they can be used as a structural analog of Watson-Crick base pairs. Remarkably, they can adopt two different conformations responding to their interaction energies, and one of them is the stacking conformation of the nonpolar aromatic group causing the site-selective flipping of the opposite base in a DNA double helix. The base pair-mimic nucleosides can be used to study the mechanism responsible for the base stacking and the flipping of bases out of a nucleic acid duplex.
Synthesis of phosphonic acid derivatized bipyridine ligands and their ruthenium complexes.

Science.gov (United States)

Norris, Michael R; Concepcion, Javier J; Glasson, Christopher R K; Fang, Zhen; Lapides, Alexander M; Ashford, Dennis L; Templeton, Joseph L; Meyer, Thomas J

2013-11-04

Water-stable, surface-bound chromophores, catalysts, and assemblies are an essential element in dye-sensitized photoelectrosynthesis cells for the generation of solar fuels by water splitting and CO2 reduction to CO, other oxygenates, or hydrocarbons. Phosphonic acid derivatives provide a basis for stable chemical binding on metal oxide surfaces. We report here the efficient synthesis of 4,4'-bis(diethylphosphonomethyl)-2,2'-bipyridine and 4,4'-bis(diethylphosphonate)-2,2'-bipyridine, as well as the mono-, bis-, and tris-substituted ruthenium complexes, [Ru(bpy)2(Pbpy)](2+), [Ru(bpy)(Pbpy)2](2+), [Ru(Pbpy)3](2+), [Ru(bpy)2(CPbpy)](2+), [Ru(bpy)(CPbpy)2](2+), and [Ru(CPbpy)3](2+) [bpy = 2,2'-bipyridine; Pbpy = 4,4'-bis(phosphonic acid)-2,2'-bipyridine; CPbpy = 4,4'-bis(methylphosphonic acid)-2,2'-bipyridine].

Multicenter Evaluation of a New Shortened Peptide Nucleic Acid Fluorescence In Situ Hybridization Procedure for Species Identification of Select Gram-Negative Bacilli from Blood Cultures▿

Science.gov (United States)

Morgan, Margie; Marlowe, Elizabeth; Della-Latta, Phyllis; Salimnia, Hossein; Novak-Weekley, Susan; Wu, Fann; Crystal, Benjamin S.

2010-01-01

A shortened protocol for two peptide nucleic acid fluorescence in situ hybridization (PNA FISH) assays for the detection of Gram-negative bacilli from positive blood cultures was evaluated in a multicenter trial. There was 100% concordance between the two protocols for each assay (368 of 368 and 370 of 370 results) and 99.7% (367 of 368 and 369 of 370 results) agreement with routine laboratory techniques. PMID:20357212
Method of Identifying a Base in a Nucleic Acid

Science.gov (United States)

Fodor, Stephen P. A.; Lipshutz, Robert J.; Huang, Xiaohua

1999-01-01

Devices and techniques for hybridization of nucleic acids and for determining the sequence of nucleic acids. Arrays of nucleic acids are formed by techniques, preferably high resolution, light-directed techniques. Positions of hybridization of a target nucleic acid are determined by, e.g., epifluorescence microscopy. Devices and techniques are proposed to determine the sequence of a target nucleic acid more efficiently and more quickly through such synthesis and detection techniques.
Detection of nucleic acid sequences by invader-directed cleavage

Science.gov (United States)

Brow, Mary Ann D.; Hall, Jeff Steven Grotelueschen; Lyamichev, Victor; Olive, David Michael; Prudent, James Robert

1999-01-01

The present invention relates to means for the detection and characterization of nucleic acid sequences, as well as variations in nucleic acid sequences. The present invention also relates to methods for forming a nucleic acid cleavage structure on a target sequence and cleaving the nucleic acid cleavage structure in a site-specific manner. The 5' nuclease activity of a variety of enzymes is used to cleave the target-dependent cleavage structure, thereby indicating the presence of specific nucleic acid sequences or specific variations thereof. The present invention further relates to methods and devices for the separation of nucleic acid molecules based by charge.
Enhanced anti-HIV-1 activity of G-quadruplexes comprising locked nucleic acids and intercalating nucleic acids

DEFF Research Database (Denmark)

Pedersen, Erik Bjerregaard; Nielsen, Jakob Toudahl; Nielsen, Claus

2011-01-01

Two G-quadruplex forming sequences, 50-TGGGAG and the 17-mer sequence T30177, which exhibit anti-HIV-1 activity on cell lines, were modified using either locked nucleic acids (LNA) or via insertions of (R)-1-O-(pyren-1-ylmethyl)glycerol (intercalating nucleic acid, INA) or (R)-1-O-[4-(1......-pyrenylethynyl)phenylmethyl]glycerol (twisted intercalating nucleic acid, TINA). Incorporation of LNA or INA/TINA monomers provide as much as 8-fold improvement of anti-HIV-1 activity. We demonstrate for the first time a detailed analysis of the effect the incorporation of INA/TINA monomers in quadruplex forming...
Synthesis of novel castor oil phosphonates

Science.gov (United States)

Castor oil has served as a versatile hydroxy fatty acid (HFA); its principle component, ricinoleic acid, can be isolated from castor oil and has been modified extensively for a number of applications. Additionally, phosphonates and their corresponding phosphonic acids are a functional moiety that ha...
Nucleic acid binding and other biomedical properties of artificial oligolysines

Directory of Open Access Journals (Sweden)

Roviello GN

2016-11-01

Full Text Available Giovanni N Roviello,1 Caterina Vicidomini,1 Vincenzo Costanzo,1 Valentina Roviello2 1CNR Istituto di Biostrutture e Bioimmagini, Via Mezzocannone site and Headquarters, 2Centro Regionale di Competenza (CRdC Tecnologie, Via Nuova Agnano, Napoli, Italy Abstract: In the present study, we report the interaction of an artificial oligolysine (referred to as AOL realized in our laboratory with targets of biomedical importance. These included polyinosinic acid (poly rI and its complex with polycytidylic acid (poly I:C, RNAs with well-known interferon-inducing ability, and double-stranded (ds DNA. The ability of the peptide to bind both single-stranded poly rI and ds poly I:C RNAs emerged from our circular dichroism (CD and ultraviolet (UV studies. In addition, we found that AOL forms complexes with dsDNA, as shown by spectroscopic binding assays and UV thermal denaturation experiments. These findings are encouraging for the possible use of AOL in biomedicine for nucleic acid targeting and oligonucleotide condensation, with the latter being a key step preceding their clinical application. Moreover, we tested the ability of AOL to bind to proteins, using serum albumin as a model protein. We demonstrated the oligolysine–protein binding by CD experiments which suggested that AOL, positively charged under physiological conditions, binds to the protein regions rich in anionic residues. Finally, the morphology characterization of the solid oligolysine, performed by scanning electron microscopy, showed different crystal forms including cubic-shaped crystals confirming the high purity of AOL. Keywords: nucleic acid binding, polyinosinic acid, double-stranded nucleic acids, oligolysine, circular dichroism
Spectroscopic Study of the Binding of Netropsin and Hoechst 33258 to Nucleic Acids

Science.gov (United States)

Vardevanyan, P. O.; Parsadanyan, M. A.; Antonyan, A. P.; Sahakyan, V. G.

2018-05-01

The interaction of groove binding compounds — peptide antibiotic (polyamide) netropsin and fluorescent dye (bisbenzimidazole) Hoechst 33258 — with the double-stranded DNA and synthetic double-stranded polynucleotide poly(rA)-poly(rU) has been studied by spectrophotometry. Absorption spectra of these ligand complexes with nucleic acids have been obtained. Spectral changes at the complexation of individual ligands with the mentioned nucleic acids reveal the similarity of binding of each of these ligands with both DNA and RNA. Based on the spectroscopic measurements, the binding parameters of netropsin and Hoechst 33258 binding to DNA and poly(rA)-poly(rU) - K and n, as well as the thermodynamic parameters ΔS, ΔG, and ΔH have been determined. It was found that the binding of Hoechst 33258 to both nucleic acids is accompanied by a positive change in enthalpy, while in the case of netropsin the change in enthalpy is negative. Moreover, the contribution of entropy to the formation of the complexes is more pronounced in the case of Hoechst 33258.
Intracellular delivery of peptide nucleic acid and organic molecules using zeolite-L nanocrystals.

Science.gov (United States)

Bertucci, Alessandro; Lülf, Henning; Septiadi, Dedy; Manicardi, Alex; Corradini, Roberto; De Cola, Luisa

2014-11-01

The design and synthesis of smart nanomaterials can provide interesting potential applications for biomedical purposes from bioimaging to drug delivery. Manufacturing multifunctional systems in a way to carry bioactive molecules, like peptide nucleic acids able to recognize specific targets in living cells, represents an achievement towards the development of highly selective tools for both diagnosis and therapeutics. This work describes a very first example of the use of zeolite nanocrystals as multifunctional nanocarriers to deliver simultaneously PNA and organic molecules into living cells. Zeolite-L nanocrystals are functionalized by covalently attaching the PNA probes onto the surface, while the channel system is filled with fluorescent guest molecules. The cellular uptake of the PNA/Zeolite-L hybrid material is then significantly increased by coating the whole system with a thin layer of biodegradable poly-L-lysine. The delivery of DAPI as a model drug molecule, inserted into the zeolite pores, is also demonstrated to occur in the cells, proving the multifunctional ability of the system. Using this zeolite nanosystem carrying PNA probes designed to target specific RNA sequences of interest in living cells could open new possibilities for theranostic and gene therapy applications. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.
Multiplexed microfluidic approach for nucleic acid enrichment

Science.gov (United States)

VanderNoot, Victoria A.; Langevin, Stanley Alan; Bent, Zachary; Renzi, Ronald F.; Ferko, Scott M.; Van De Vreugde, James L.; Lane, Todd; Patel, Kamlesh; Branda, Steven

2016-04-26

A system for enhancing a nucleic acid sample may include a one pump, a denaturing chamber; a microfluidic hydroxyapatite chromatography device configured for performing hydroxyapatite chromatography on the nucleic acid sample, a sample collector, and tubing connecting the pump with the denaturing chamber, the hydroxyapatite chromatography device and the sample collector such that the pump may be used to move the nucleic acid sample from the denaturing chamber to the hydroxyapatite chromatography device and then to the sample collector.
Hybridization and sequencing of nucleic acids using base pair mismatches

Science.gov (United States)

Fodor, Stephen P. A.; Lipshutz, Robert J.; Huang, Xiaohua

2001-01-01

Devices and techniques for hybridization of nucleic acids and for determining the sequence of nucleic acids. Arrays of nucleic acids are formed by techniques, preferably high resolution, light-directed techniques. Positions of hybridization of a target nucleic acid are determined by, e.g., epifluorescence microscopy. Devices and techniques are proposed to determine the sequence of a target nucleic acid more efficiently and more quickly through such synthesis and detection techniques.
Probe kit for identifying a base in a nucleic acid

Science.gov (United States)

Fodor, Stephen P. A.; Lipshutz, Robert J.; Huang, Xiaohua

2001-01-01

Devices and techniques for hybridization of nucleic acids and for determining the sequence of nucleic acids. Arrays of nucleic acids are formed by techniques, preferably high resolution, light-directed techniques. Positions of hybridization of a target nucleic acid are determined by, e.g., epifluorescence microscopy. Devices and techniques are proposed to determine the sequence of a target nucleic acid more efficiently and more quickly through such synthesis and detection techniques.
Nucleic acid drugs: a novel approach

African Journals Online (AJOL)

Administrator

Nucleic acid base sequence of proteins plays a crucial role in the expression of gene. The gene is responsible for the synthesis of proteins and these proteins, which are synthesized, are responsible for the biological process and also for dreadful diseases as well. Once if the nucleic acid sequence is altered, we would be ...
Electrochemical paper-based peptide nucleic acid biosensor for detecting human papillomavirus

Energy Technology Data Exchange (ETDEWEB)

Teengam, Prinjaporn [Program in Petrochemistry, Faculty of Science, Chulalongkorn University, Pathumwan, Bangkok, 10330 (Thailand); Siangproh, Weena [Department of Chemistry, Faculty of Science, Srinakharinwirot University, Bangkok, 10110 (Thailand); Tuantranont, Adisorn [Nanoelectronics and MEMS Laboratory, National Electronics and Computer Technology Center, Pathumthani, 12120 (Thailand); Henry, Charles S. [Department of Chemistry, Colorado State University, Fort Collins, CO, 80523 (United States); Vilaivan, Tirayut [Organic Synthesis Research Unit, Department of Chemistry, Faculty of Science, Chulalongkorn University, Pathumwan, Bangkok, 10330 (Thailand); Chailapakul, Orawon, E-mail: corawon@chula.ac.th [Electrochemistry and Optical Spectroscopy Research Unit, Department of Chemistry, Chulalongkorn University, Pathumwan, Bangkok, 10330 (Thailand); Nanotec-CU Center of Excellence on Food and Agriculture, Bangkok, 10330 (Thailand)

2017-02-01

A novel paper-based electrochemical biosensor was developed using an anthraquinone-labeled pyrrolidinyl peptide nucleic acid (acpcPNA) probe (AQ-PNA) and graphene-polyaniline (G-PANI) modified electrode to detect human papillomavirus (HPV). An inkjet printing technique was employed to prepare the paper-based G-PANI-modified working electrode. The AQ-PNA probe baring a negatively charged amino acid at the N-terminus was immobilized onto the electrode surface through electrostatic attraction. Electrochemical impedance spectroscopy (EIS) was used to verify the AQ-PNA immobilization. The paper-based electrochemical DNA biosensor was used to detect a synthetic 14-base oligonucleotide target with a sequence corresponding to human papillomavirus (HPV) type 16 DNA by measuring the electrochemical signal response of the AQ label using square-wave voltammetry before and after hybridization. It was determined that the current signal significantly decreased after the addition of target DNA. This phenomenon is explained by the rigidity of PNA-DNA duplexes, which obstructs the accessibility of electron transfer from the AQ label to the electrode surface. Under optimal conditions, the detection limit of HPV type 16 DNA was found to be 2.3 nM with a linear range of 10–200 nM. The performance of this biosensor on real DNA samples was tested with the detection of PCR-amplified DNA samples from the SiHa cell line. The new method employs an inexpensive and disposable device, which easily incinerated after use and is promising for the screening and monitoring of the amount of HPV-DNA type 16 to identify the primary stages of cervical cancer. - Highlights: • A paper-based DNA biosensor using AQ-PNA probe and G-PANI modified electrode was first developed. • This developed DNA biosensor was highly specific over the non-complementary DNA. • This sensor was successfully applied to detect the HPV-DNA type 16 obtained from cancer cell lines. • This sensor is inexpensive and
Analysis of nucleic acid chaperoning by the prion protein and its inhibition by oligonucleotides.

Science.gov (United States)

Guichard, Cécile; Ivanyi-Nagy, Roland; Sharma, Kamal Kant; Gabus, Caroline; Marc, Daniel; Mély, Yves; Darlix, Jean-Luc

2011-10-01

Prion diseases are unique neurodegenerative illnesses associated with the conversion of the cellular prion protein (PrP(C)) into the aggregated misfolded scrapie isoform, named PrP(Sc). Recent studies on the physiological role of PrP(C) revealed that this protein has probably multiple functions, notably in cell-cell adhesion and signal transduction, and in assisting nucleic acid folding. In fact, in vitro findings indicated that the human PrP (huPrP) possesses nucleic acid binding and annealing activities, similarly to nucleic acid chaperone proteins that play essential roles in cellular DNA and RNA metabolism. Here, we show that a peptide, representing the N-terminal domain of huPrP, facilitates nucleic acid annealing by two parallel pathways nucleated through the stem termini. We also show that PrP of human or ovine origin facilitates DNA strand exchange, ribozyme-directed cleavage of an RNA template and RNA trans-splicing in a manner similar to the nucleocapsid protein of HIV-1. In an attempt to characterize inhibitors of PrP-chaperoning in vitro we discovered that the thioaptamer 5'-GACACAAGCCGA-3' was extensively inhibiting the PrP chaperoning activities. At the same time a recently characterized methylated oligoribonucleotide inhibiting the chaperoning activity of the HIV-1 nucleocapsid protein was poorly impairing the PrP chaperoning activities.
Quantitative structure-property relationship (correlation analysis) of phosphonic acid-based chelates in design of MRI contrast agent.

Science.gov (United States)

Tiwari, Anjani K; Ojha, Himanshu; Kaul, Ankur; Dutta, Anupama; Srivastava, Pooja; Shukla, Gauri; Srivastava, Rakesh; Mishra, Anil K

2009-07-01

Nuclear magnetic resonance imaging is a very useful tool in modern medical diagnostics, especially when gadolinium (III)-based contrast agents are administered to the patient with the aim of increasing the image contrast between normal and diseased tissues. With the use of soft modelling techniques such as quantitative structure-activity relationship/quantitative structure-property relationship after a suitable description of their molecular structure, we have studied a series of phosphonic acid for designing new MRI contrast agent. Quantitative structure-property relationship studies with multiple linear regression analysis were applied to find correlation between different calculated molecular descriptors of the phosphonic acid-based chelating agent and their stability constants. The final quantitative structure-property relationship mathematical models were found as--quantitative structure-property relationship Model for phosphonic acid series (Model 1)--log K(ML) = {5.00243(+/-0.7102)}- MR {0.0263(+/-0.540)}n = 12 l r l = 0.942 s = 0.183 F = 99.165 quantitative structure-property relationship Model for phosphonic acid series (Model 2)--log K(ML) = {5.06280(+/-0.3418)}- MR {0.0252(+/- .198)}n = 12 l r l = 0.956 s = 0.186 F = 99.256.
Soybean phytase and nucleic acid encoding the same

OpenAIRE

1999-01-01

Isolated soybean phytase polypeptides and isolated nucleic acids encoding soybean phytases are provided. The invention is also directed to nucleic acid expression constructs, vectors, and host cells comprising the isolated soybean phytase nucleic acids, as well as methods for producing recombinant and non-recombinant purified soybean phytase. The invention also relates to transgenic plants expressing the soybean phytase, particularly expression under seed-specific expression control elements.
Nucleic Acid-Based Nanoconstructs

Science.gov (United States)

Focuses on the design, synthesis, characterization, and development of spherical nucleic acid constructs as effective nanotherapeutic, single-entity agents for the treatment of glioblastoma multiforme and prostate cancers.
Nucleic acid-functionalized transition metal nanosheets for biosensing applications.

Science.gov (United States)

Mo, Liuting; Li, Juan; Liu, Qiaoling; Qiu, Liping; Tan, Weihong

2017-03-15

In clinical diagnostics, as well as food and environmental safety practices, biosensors are powerful tools for monitoring biological or biochemical processes. Two-dimensional (2D) transition metal nanomaterials, including transition metal chalcogenides (TMCs) and transition metal oxides (TMOs), are receiving growing interest for their use in biosensing applications based on such unique properties as high surface area and fluorescence quenching abilities. Meanwhile, nucleic acid probes based on Watson-Crick base-pairing rules are also being widely applied in biosensing based on their excellent recognition capability. In particular, the emergence of functional nucleic acids in the 1980s, especially aptamers, has substantially extended the recognition capability of nucleic acids to various targets, ranging from small organic molecules and metal ions to proteins and cells. Based on π-π stacking interaction between transition metal nanosheets and nucleic acids, biosensing systems can be easily assembled. Therefore, the combination of 2D transition metal nanomaterials and nucleic acids brings intriguing opportunities in bioanalysis and biomedicine. In this review, we summarize recent advances of nucleic acid-functionalized transition metal nanosheets in biosensing applications. The structure and properties of 2D transition metal nanomaterials are first discussed, emphasizing the interaction between transition metal nanosheets and nucleic acids. Then, the applications of nucleic acid-functionalized transition metal nanosheet-based biosensors are discussed in the context of different signal transducing mechanisms, including optical and electrochemical approaches. Finally, we provide our perspectives on the current challenges and opportunities in this promising field. Copyright © 2016 Elsevier B.V. All rights reserved.
EGVII endoglucanase and nucleic acids encoding the same

Science.gov (United States)

Dunn-Coleman, Nigel [Los Gatos, CA; Goedegebuur, Frits [Vlaardingen, NL; Ward, Michael [San Francisco, CA; Yao, Jian [Sunnyvale, CA

2009-05-05

The present invention provides an endoglucanase nucleic acid sequence, designated egl7, and the corresponding EGVII amino acid sequence. The invention also provides expression vectors and host cells comprising a nucleic acid sequence encoding EGVII, recombinant EGVII proteins and methods for producing the same.
Detection of group B streptococci in Lim broth by use of group B streptococcus peptide nucleic acid fluorescent in situ hybridization and selective and nonselective agars.

Science.gov (United States)

Montague, Naomi S; Cleary, Timothy J; Martinez, Octavio V; Procop, Gary W

2008-10-01

The sensitivity, specificity, and positive and negative predictive values for the detection of group B streptococci from Lim enrichment broth with sheep blood agar (SBA), with selective Streptococcus agar (SSA), and by a peptide nucleic acid fluorescent in situ hybridization (PNA FISH) assay were as follows: for culture on SBA, 68.4%, 100%, 100%, and 87.9%, respectively; for culture on SSA, 85.5%, 100%, 100%, and 94.1%, respectively; and for the PNA FISH assay, 97.4%, 98.3%, 96.1%, and 98.9%, respectively.

Peptide and nucleic acid-directed self-assembly of cationic nanovehicles through giant unilamellar vesicle modification

DEFF Research Database (Denmark)

Tagalakis, A. D.; Maeshima, R.; Yu-Wai-Man, C.

2017-01-01

into a neuroblastoma xenograft mouse model, nanovesicle complexes were found to distribute throughout the tumour interstitium, thus providing an alternative safer approach for future development of tumour-specific therapeutic nucleic acid interventions. On oropharyngeal instillation, nanovesicle complexes displayed...
Single-stranded nucleic acids promote SAMHD1 complex formation.

Science.gov (United States)

Tüngler, Victoria; Staroske, Wolfgang; Kind, Barbara; Dobrick, Manuela; Kretschmer, Stefanie; Schmidt, Franziska; Krug, Claudia; Lorenz, Mike; Chara, Osvaldo; Schwille, Petra; Lee-Kirsch, Min Ae

2013-06-01

SAM domain and HD domain-containing protein 1 (SAMHD1) is a dGTP-dependent triphosphohydrolase that degrades deoxyribonucleoside triphosphates (dNTPs) thereby limiting the intracellular dNTP pool. Mutations in SAMHD1 cause Aicardi-Goutières syndrome (AGS), an inflammatory encephalopathy that mimics congenital viral infection and that phenotypically overlaps with the autoimmune disease systemic lupus erythematosus. Both disorders are characterized by activation of the antiviral cytokine interferon-α initiated by immune recognition of self nucleic acids. Here we provide first direct evidence that SAMHD1 associates with endogenous nucleic acids in situ. Using fluorescence cross-correlation spectroscopy, we demonstrate that SAMHD1 specifically interacts with ssRNA and ssDNA and establish that nucleic acid-binding and formation of SAMHD1 complexes are mutually dependent. Interaction with nucleic acids and complex formation do not require the SAM domain, but are dependent on the HD domain and the C-terminal region of SAMHD1. We finally demonstrate that mutations associated with AGS exhibit both impaired nucleic acid-binding and complex formation implicating that interaction with nucleic acids is an integral aspect of SAMHD1 function.
Selenium Derivatization of Nucleic Acids for Phase and Structure Determination in Nucleic Acid X-ray Crystallography

Directory of Open Access Journals (Sweden)

Zhen Huang

2008-03-01

Full Text Available Selenium derivatization (via selenomethionine of proteins for crystal structure determination via MAD phasing has revolutionized protein X-ray crystallography. It is estimated that over two thirds of all new crystal structures of proteins have been determined via Se-Met derivatization. Similarly, selenium functionalities have also been successfully incorporated into nucleic acids to facilitate their structure studies and it has been proved that this Se-derivatization has advantages over halogen strategy, which was usually used as a traditional method in this field. This review reports the development of site-specific selenium derivatization of nucleic acids for phase determination since the year of 2001 (mainly focus on the 2Ã¢Â€Â™-position of the ribose. All the synthesis of 2Ã¢Â€Â™-SeMe modified phosphoramidite building blocks (U, C, T, A, G and the according oligonucleotides are included. In addition, several structures of selenium contained nucleic acid are also described in this paper.
Detection of Group B Streptococci in Lim Broth by Use of Group B Streptococcus Peptide Nucleic Acid Fluorescent In Situ Hybridization and Selective and Nonselective Agars▿

Science.gov (United States)

Montague, Naomi S.; Cleary, Timothy J.; Martinez, Octavio V.; Procop, Gary W.

2008-01-01

The sensitivity, specificity, and positive and negative predictive values for the detection of group B streptococci from Lim enrichment broth with sheep blood agar (SBA), with selective Streptococcus agar (SSA), and by a peptide nucleic acid fluorescent in situ hybridization (PNA FISH) assay were as follows: for culture on SBA, 68.4%, 100%, 100%, and 87.9%, respectively; for culture on SSA, 85.5%, 100%, 100%, and 94.1%, respectively; and for the PNA FISH assay, 97.4%, 98.3%, 96.1%, and 98.9%, respectively. PMID:18667597
Non-enzymatic Polymerization of Nucleic Acids from Monomers

DEFF Research Database (Denmark)

Dörr, Mark; Löffler, Philipp M. G.; Monnard, Pierre-Alain

2012-01-01

synthesis of long nucleic acid polymers or to sequence-specifically amplify nucleic acid polymers, respectively. Starting from molecular requirements, details of the polymerization mechanisms and strategies are first presented and then compared. Finally, we discuss the relevance of these strategies...
Metal-Free, Site-Selective Peptide Modification by Conversion of "Customizable" Units into β-Substituted Dehydroamino Acids.

Science.gov (United States)

Saavedra, Carlos J; Hernández, Dácil; Boto, Alicia

2018-01-12

Our site-selective modification of serine or threonine units in peptides allows the generation of β-substituted dehydroamino acids, which increase peptide resistance to hydrolysis and may improve their biological properties. Both the terminal and internal positions can be modified, and different customizable units can be activated separately. Remarkably, high Z selectivity is achieved, even at internal positions. The conversion involves a one-pot oxidative radical scission/phosphorylation process by using the low-toxicity (diacetoxyiodo)benzene/iodine system as the scission reagent. The resulting α-amino phosphonates undergo a Horner-Wadsworth-Emmons reaction to produce the dehydroamino acid derivatives (in a Z/E ratio of usually >98:2) under mild and metal-free conditions. © 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.
Accelerated digestion of nucleic acids by pepsin from the stomach of chicken.

Science.gov (United States)

Liu, Y; Zhang, Y; Guo, H; Wu, W; Dong, P; Liang, X

2016-10-01

Nucleic acids have become an important nutritional supplement in poultry feed; however, the digestion of nucleic acids in poultry is unclear. The objective of this study was to investigate the digestion of nucleic acids by chicken pepsin in vitro. The extracted pepsinogen from the stomach of the chicken was purified to homogeneity. Upon activation at pH 2.0, chicken pepsinogen was converted to its active form. Nucleic acids, including λ-DNA, salmon sperm DNA and single-strand DNA (ssDNA), can be used as substrates and digested into short-chain oligonucleotides by pepsin. Interestingly, the digestion of the nucleic acids was inhibited when pepsin was treated by alkaline solution (pH 8.0) or pepstatin A. Also, the digestion of the nucleic acids was not affected by the addition of haemoglobin or bovine serum albumin. The results suggested that nucleic acids could be digested by chicken pepsin. Thus pepsin may have a role in digesting nucleic acids in vivo. Nucleic acids added to poultry fed may be digested, starting from the stomach.
MEANS AND METHODS FOR CLONING NUCLEIC ACID SEQUENCES

NARCIS (Netherlands)

Geertsma, Eric Robin; Poolman, Berend

2008-01-01

The invention provides means and methods for efficiently cloning nucleic acid sequences of interest in micro-organisms that are less amenable to conventional nucleic acid manipulations, as compared to, for instance, E.coli. The present invention enables high-throughput cloning (and, preferably,
Selected topics in photochemistry of nucleic acids. Recent results and perspectives

International Nuclear Information System (INIS)

Loeber, G.; Kittler, L.

1977-01-01

Recent results on the following photoreactions of nucleic acids are reported: photochemistry of aza-bases and minor bases, formation of photoproducts of the non-cyclobutane type, formations of furocoumarin-pyrimidine photoadducts, fluorescence of dye-nucleic acid complexes and their role in chromosomal fluorescence staining, and mechanisms of the photochemical reaction. Results are discussed with respect to: (i) photobiological relevance of light-induced defects in nucleic acids; (ii) possibilities of achieving higher selectivity of light-induced defects in nucleic acids; (iii) the use of nucleic acid photochemistry to analyze genetic material. An extensive bibliography is included. (author)
Novel Biochip Platform for Nucleic Acid Analysis

Directory of Open Access Journals (Sweden)

Juan J. Diaz-Mochon

2012-06-01

Full Text Available This manuscript describes the use of a novel biochip platform for the rapid analysis/identification of nucleic acids, including DNA and microRNAs, with very high specificity. This approach combines a unique dynamic chemistry approach for nucleic acid testing and analysis developed by DestiNA Genomics with the STMicroelectronics In-Check platform, which comprises two microfluidic optimized and independent PCR reaction chambers, and a sequential microarray area for nucleic acid capture and identification by fluorescence. With its compact bench-top “footprint” requiring only a single technician to operate, the biochip system promises to transform and expand routine clinical diagnostic testing and screening for genetic diseases, cancers, drug toxicology and heart disease, as well as employment in the emerging companion diagnostics market.
Hydrophosphorylation of substituted alkynes by phosphonic acids

International Nuclear Information System (INIS)

Nifant'ev, E.F.; Solovetskaya, L.A.; Maslennikova, V.I.; Sergeev, N.M.

1987-01-01

Hydrophosphorylation of functionally substituted alkynes by phosphonic acids can be a convenient method for synthesis of functionally substituted mono- and diphosphine oxides. The ease of hydrophosphorylation is determined by the strength of the negative inductive effect of the substituents on the triple bond and the steric factor. The structure of the bis-adducts was confirmed by elementary analysis and the 31 P and 13 C NMR spectra. The 31 P NMR spectrum is an AB two-spin system. The values of the chemical shifts and spin-spin interaction constants 3 J/sub PP/ are in agreement with the data in the literature for similar compounds
Assembly of barcode-like nucleic acid nanostructures.

Science.gov (United States)

Wang, Pengfei; Tian, Cheng; Li, Xiang; Mao, Chengde

2014-10-15

Barcode-like (BC) nanopatterns from programmed self-assembly of nucleic acids (DNA and RNA) are reported. BC nanostructures are generated by the introduction of open spaces at selected sites to an otherwise closely packed, plain, rectangle nucleic acid nanostructure. This strategy is applied to nanostructures assembled from both origami approach and single stranded tile approach. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.
Novel sulfonated poly (ether ether ketone)/phosphonic acid-functionalized titania nanohybrid membrane by an in situ method for direct methanol fuel cells

Science.gov (United States)

Wu, Hong; Cao, Ying; Li, Zhen; He, Guangwei; Jiang, Zhongyi

2015-01-01

Sulfonated poly (ether ether ketone)/phosphonic acid-functionalized titania nanohybrid membranes are prepared by an in situ method using titanium tetrachloride (TiCl4) as inorganic precursor and amino trimethylene phosphonic acid (ATMP) as modifier. Phosphonic acid-functionalized titania nanoparticles with a uniform particle size of ∼50 nm are formed and dispersed homogeneously in the SPEEK matrix with good interfacial compatibility. Accordingly, the nanohybrid membranes display remarkably enhanced proton conduction property due to the incorporation of additional sites for proton transport and the formation of well-connected channels by bridging the hydrophilic domains in SPEEK matrix. The nanohybrid membrane with 6 wt. % of phosphonic acid-functionalized titania nanoparticles exhibits the highest proton conductivity of 0.334 S cm-1 at 65 °C and 100% RH, which is 63.7% higher than that of pristine SPEEK membrane. Furthermore, the as-prepared nanohybrid membranes also show elevated thermal and mechanical stabilities as well as decreased methanol permeability.
Identification of Escherichia coli, Klebsiella pneumoniae, and Pseudomonas aeruginosa in Blood Cultures: a Multicenter Performance Evaluation of a Three-Color Peptide Nucleic Acid Fluorescence In Situ Hybridization Assay▿

Science.gov (United States)

Della-Latta, Phyllis; Salimnia, Hossein; Painter, Theresa; Wu, Fann; Procop, Gary W.; Wilson, Deborah A.; Gillespie, Wendy; Mender, Alayna; Crystal, Benjamin S.

2011-01-01

A multicenter evaluation was undertaken to evaluate the performance of a new three-color peptide nucleic acid fluorescence in situ hybridization assay that identifies isolates directly from blood cultures positive for Gram-negative bacilli (GNB). The assay correctly identified 100% (186/186) of the Escherichia coli isolates, 99.1% (109/110) of the Klebsiella pneumoniae isolates, and 95.8% (46/48) of the Pseudomonas aeruginosa isolates in this study. Negative assay results were correctly obtained for 162 of 165 other GNB (specificity, 98.2%). PMID:21490185
Actinide phosphonate complexes in aqueous solutions

International Nuclear Information System (INIS)

Nash, K.L.

1993-01-01

Complexes formed by actinides with carboxylic acids, polycarboxylic acids, and aminopolycarboxylic acids play a central role in both the basic and process chemistry of the actinides. Recent studies of f-element complexes with phosphonic acid ligands indicate that new ligands incorporating doubly ionizable phosphonate groups (-PO 3 H 2 ) have many properties which are unique chemically, and promise more efficient separation processes for waste cleanup and environmental restoration. Simple diphosphonate ligands form much stronger complexes than isostructural carboxylates, often exhibiting higher solubility as well. In this manuscript recent studies of the thermodynamics and kinetics of f-element complexation by 1,1 and 1,2 diphosphonic acid ligands are described
Oxacillin sensitization of methicillin-resistant Staphylococcus aureus and methicillin-resistant Staphylococcus pseudintermedius by antisense peptide nucleic acids in vitro.

Science.gov (United States)

Goh, Shan; Loeffler, Anette; Lloyd, David H; Nair, Sean P; Good, Liam

2015-11-11

Antibiotic resistance genes can be targeted by antisense agents, which can reduce their expression and thus restore cellular susceptibility to existing antibiotics. Antisense inhibitors can be gene and pathogen specific, or designed to inhibit a group of bacteria having conserved sequences within resistance genes. Here, we aimed to develop antisense peptide nucleic acids (PNAs) that could be used to effectively restore susceptibility to β-lactams in methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-resistant Staphylococcus pseudintermedius (MRSP). Antisense PNAs specific for conserved regions of the mobilisable gene mecA, and the growth essential gene, ftsZ, were designed. Clinical MRSA and MRSP strains of high oxacillin resistance were treated with PNAs and assayed for reduction in colony forming units on oxacillin plates, reduction in target gene mRNA levels, and cell size. Anti-mecA PNA at 7.5 and 2.5 μM reduced mecA mRNA in MRSA and MRSP (p resistance in staphylococci. Further studies are warranted as clinical treatment alternatives are needed.
Room-Temperature Alternative to the Arbuzov Reaction: The Reductive Deoxygenation of Acyl Phosphonates

OpenAIRE

Kedrowski, Sean M. A.; Dougherty, Dennis A.

2010-01-01

The reductive deoxygenation of acyl phosphonates using a Wolff−Kishner-like sequence is described. This transformation allows direct access to alkyl phosphonates from acyl phosphonates at room temperature. The method can be combined with acyl phosphonate synthesis into a one pot, four-step procedure for the conversion of carboxylic acids into alkyl phosphonates. The methodology works well for a variety of aliphatic acids and shows a functional group tolerance similar to that of other hydrazon...
Universal nucleic acids sample preparation method for cells, spores and their mixture

Science.gov (United States)

Bavykin, Sergei [Darien, IL

2011-01-18

The present invention relates to a method for extracting nucleic acids from biological samples. More specifically the invention relates to a universal method for extracting nucleic acids from unidentified biological samples. An advantage of the presently invented method is its ability to effectively and efficiently extract nucleic acids from a variety of different cell types including but not limited to prokaryotic or eukaryotic cells and/or recalcitrant organisms (i.e. spores). Unlike prior art methods which are focused on extracting nucleic acids from vegetative cell or spores, the present invention effectively extracts nucleic acids from spores, multiple cell types or mixtures thereof using a single method. Important that the invented method has demonstrated an ability to extract nucleic acids from spores and vegetative bacterial cells with similar levels effectiveness. The invented method employs a multi-step protocol which erodes the cell structure of the biological sample, isolates, labels, fragments nucleic acids and purifies labeled samples from the excess of dye.
Nucleic acid and nucleotide-mediated synthesis of inorganic nanoparticles

Science.gov (United States)

Berti, Lorenzo; Burley, Glenn A.

2008-02-01

Since the advent of practical methods for achieving DNA metallization, the use of nucleic acids as templates for the synthesis of inorganic nanoparticles (NPs) has become an active area of study. It is now widely recognized that nucleic acids have the ability to control the growth and morphology of inorganic NPs. These biopolymers are particularly appealing as templating agents as their ease of synthesis in conjunction with the possibility of screening nucleotide composition, sequence and length, provides the means to modulate the physico-chemical properties of the resulting NPs. Several synthetic procedures leading to NPs with interesting photophysical properties as well as studies aimed at rationalizing the mechanism of nucleic acid-templated NP synthesis are now being reported. This progress article will outline the current understanding of the nucleic acid-templated process and provides an up to date reference in this nascent field.
Nucleic acid detection system and method for detecting influenza

Science.gov (United States)

Cai, Hong; Song, Jian

2015-03-17

The invention provides a rapid, sensitive and specific nucleic acid detection system which utilizes isothermal nucleic acid amplification in combination with a lateral flow chromatographic device, or DNA dipstick, for DNA-hybridization detection. The system of the invention requires no complex instrumentation or electronic hardware, and provides a low cost nucleic acid detection system suitable for highly sensitive pathogen detection. Hybridization to single-stranded DNA amplification products using the system of the invention provides a sensitive and specific means by which assays can be multiplexed for the detection of multiple target sequences.

Nucleic acids in circulation

Indian Academy of Sciences (India)

Elevated blood levels of extracellular nucleic acids have been reported in various disease conditions; such as ageing and age-related degenerative disorders, cancer; acute and chronic inflammatory conditions, severe trauma and autoimmune disorders. In addition to genomic DNA and nucleosomes, mitochondrial DNA is ...
Synthesis of highly phosphonic acid functionalized benzene-bridged periodic mesoporous organosilicas for use as efficient dye adsorbents

International Nuclear Information System (INIS)

Deka, Juti Rani; Liu, Chia-Ling; Wang, Tzu-Hua; Chang, Wei-Chieh; Kao, Hsien-Ming

2014-01-01

Highlights: • Synthesis of highly phosphonic acid functionalized benzene-bridged PMOs. • Phosphonic acid loaded PMOs as adsorbent for cationic and anionic dyes. • Due to electrostatic interaction the adsorbent has high dye adsorption capacity. • π–π stacking interaction between benzene and dye enhances adsorption capacity. • Intraparticle diffusion played a dominant role in the adsorption process. - Abstract: Periodic mesoporous organosilicas (PMOs) with benzene bridging groups in the silica wall were functionalized with a tunable content of phosphonic acid groups. These bifunctional materials were synthesized by co-condensation of two different organosilane precursors, that is, 1,4-bis(triethoxysilyl)benzene (BTEB) and sodium 3-(trihydroxysilyl)propyl methyl phosphate (SPMP), under acidic conditions using nonionic surfactant Brij-S10 as template. The materials exhibited well-ordered mesostructures and were characterized by X-ray diffraction, nitrogen sorption, TEM, TGA, FTIR, and solid-state NMR measurements. The materials thus obtained were employed as adsorbents to remove different types of dyes, for example, cationic dyes methylene blue and phenosafranine, anionic orange II, and amphoteric rhodamine B, from aqueous solutions. The materials exhibited a remarkably high adsorption capacity than activated carbon due to their ordered mesostructures, a large number of phosphonic acid groups, and high surface areas. The adsorption was mainly governed by electrostatic interaction, but also involved π–π stacking interaction as well as hydrogen bonding. The adsorption kinetics can be better fitted by the pseudo-second order model. The adsorption process was controlled by the mechanisms of external mass transfer and intraparticle diffusion. The materials retained more than 97% dye removal efficiency after use for five consecutive cycles
Electricity-free, sequential nucleic acid and protein isolation.

Science.gov (United States)

Pawlowski, David R; Karalus, Richard J

2012-05-15

Traditional and emerging pathogens such as Enterohemorrhagic Escherichia coli (EHEC), Yersinia pestis, or prion-based diseases are of significant concern for governments, industries and medical professionals worldwide. For example, EHECs, combined with Shigella, are responsible for the deaths of approximately 325,000 children each year and are particularly prevalent in the developing world where laboratory-based identification, common in the United States, is unavailable (1). The development and distribution of low cost, field-based, point-of-care tools to aid in the rapid identification and/or diagnosis of pathogens or disease markers could dramatically alter disease progression and patient prognosis. We have developed a tool to isolate nucleic acids and proteins from a sample by solid-phase extraction (SPE) without electricity or associated laboratory equipment (2). The isolated macromolecules can be used for diagnosis either in a forward lab or using field-based point-of-care platforms. Importantly, this method provides for the direct comparison of nucleic acid and protein data from an un-split sample, offering a confidence through corroboration of genomic and proteomic analysis. Our isolation tool utilizes the industry standard for solid-phase nucleic acid isolation, the BOOM technology, which isolates nucleic acids from a chaotropic salt solution, usually guanidine isothiocyanate, through binding to silica-based particles or filters (3). CUBRC's proprietary solid-phase extraction chemistry is used to purify protein from chaotropic salt solutions, in this case, from the waste or flow-thru following nucleic acid isolation(4). By packaging well-characterized chemistries into a small, inexpensive and simple platform, we have generated a portable system for nucleic acid and protein extraction that can be performed under a variety of conditions. The isolated nucleic acids are stable and can be transported to a position where power is available for PCR amplification
Nucleic acid-induced antiviral immunity in invertebrates: an evolutionary perspective.

Science.gov (United States)

Wang, Pei-Hui; Weng, Shao-Ping; He, Jian-Guo

2015-02-01

Nucleic acids derived from viral pathogens are typical pathogen associated molecular patterns (PAMPs). In mammals, the recognition of viral nucleic acids by pattern recognition receptors (PRRs), which include Toll-like receptors (TLRs) and retinoic acid-inducible gene (RIG)-I-like receptors (RLRs), induces the release of inflammatory cytokines and type I interferons (IFNs) through the activation of nuclear factor κB (NF-κB) and interferon regulatory factor (IRF) 3/7 pathways, triggering the host antiviral state. However, whether nucleic acids can induce similar antiviral immunity in invertebrates remains ambiguous. Several studies have reported that nucleic acid mimics, especially dsRNA mimic poly(I:C), can strongly induce non-specific antiviral immune responses in insects, shrimp, and oyster. This behavior shows multiple similarities to the hallmarks of mammalian IFN responses. In this review, we highlight the current understanding of nucleic acid-induced antiviral immunity in invertebrates. We also discuss the potential recognition and regulatory mechanisms that confer non-specific antiviral immunity on invertebrate hosts. Copyright © 2014 Elsevier Ltd. All rights reserved.
Geometric properties of nucleic acids with potential for autobuilding

International Nuclear Information System (INIS)

Gruene, Tim; Sheldrick, George M.

2011-01-01

Algorithms and geometrical properties are described for the automated building of nucleic acids in experimental electron density. Medium- to high-resolution X-ray structures of DNA and RNA molecules were investigated to find geometric properties useful for automated model building in crystallographic electron-density maps. We describe a simple method, starting from a list of electron-density ‘blobs’, for identifying backbone phosphates and nucleic acid bases based on properties of the local electron-density distribution. This knowledge should be useful for the automated building of nucleic acid models into electron-density maps. We show that the distances and angles involving C1′ and the P atoms, using the pseudo-torsion angles η' and θ' that describe the …P—C1′—P—C1′… chain, provide a promising basis for building the nucleic acid polymer. These quantities show reasonably narrow distributions with asymmetry that should allow the direction of the phosphate backbone to be established
Optimizing the specificity of nucleic acid hybridization.

Science.gov (United States)

Zhang, David Yu; Chen, Sherry Xi; Yin, Peng

2012-01-22

The specific hybridization of complementary sequences is an essential property of nucleic acids, enabling diverse biological and biotechnological reactions and functions. However, the specificity of nucleic acid hybridization is compromised for long strands, except near the melting temperature. Here, we analytically derived the thermodynamic properties of a hybridization probe that would enable near-optimal single-base discrimination and perform robustly across diverse temperature, salt and concentration conditions. We rationally designed 'toehold exchange' probes that approximate these properties, and comprehensively tested them against five different DNA targets and 55 spurious analogues with energetically representative single-base changes (replacements, deletions and insertions). These probes produced discrimination factors between 3 and 100+ (median, 26). Without retuning, our probes function robustly from 10 °C to 37 °C, from 1 mM Mg(2+) to 47 mM Mg(2+), and with nucleic acid concentrations from 1 nM to 5 µM. Experiments with RNA also showed effective single-base change discrimination.
Synthesis of Ethylene diamine tetra methylene phosphonic acid EDTMP

International Nuclear Information System (INIS)

Assaad, Th.

2009-06-01

Ethylenediamine tetramethylene phosphonic acid (EDTMP) is one of the most widely used ligands which forms stable complexes with various radionuclides all of which have shown high bone affinity and other favorable pharmacological characteristics in biodistribution studies. The EDTMP is very important precursor in the preparation of radiopharmaceutical kits which is used in the applications for bone diagnosis and therapy was synthesized in one step with Mannich type reaction. The obtained product was fully characterized by its multi-nuclear NMR and IR spectroscopy. The purity of the product was confirmed by HPLC. (author)
Photochemical reactions of nucleic acids and their constituents of photobiological relevance

International Nuclear Information System (INIS)

Saito, I.; Sugiyama, H.; Matsuura, T.

1983-01-01

A review is given of the papers published from 1977 to May 1983 on the UV-induced photochemical reactions of nucleic acids and their constituents of photobiological relevance where the structures of photoproducts have been fully characterized. Among the topics discussed are photoadditions relevant to nucleic acid-protein photocrosslinking, photoreactions with psoralens and nucleic acids and photochemical reactions of polynucleotides. (U.K.)
Integrated sample-to-detection chip for nucleic acid test assays.

Science.gov (United States)

Prakash, R; Pabbaraju, K; Wong, S; Tellier, R; Kaler, K V I S

2016-06-01

Nucleic acid based diagnostic techniques are routinely used for the detection of infectious agents. Most of these assays rely on nucleic acid extraction platforms for the extraction and purification of nucleic acids and a separate real-time PCR platform for quantitative nucleic acid amplification tests (NATs). Several microfluidic lab on chip (LOC) technologies have been developed, where mechanical and chemical methods are used for the extraction and purification of nucleic acids. Microfluidic technologies have also been effectively utilized for chip based real-time PCR assays. However, there are few examples of microfluidic systems which have successfully integrated these two key processes. In this study, we have implemented an electro-actuation based LOC micro-device that leverages multi-frequency actuation of samples and reagents droplets for chip based nucleic acid extraction and real-time, reverse transcription (RT) PCR (qRT-PCR) amplification from clinical samples. Our prototype micro-device combines chemical lysis with electric field assisted isolation of nucleic acid in a four channel parallel processing scheme. Furthermore, a four channel parallel qRT-PCR amplification and detection assay is integrated to deliver the sample-to-detection NAT chip. The NAT chip combines dielectrophoresis and electrostatic/electrowetting actuation methods with resistive micro-heaters and temperature sensors to perform chip based integrated NATs. The two chip modules have been validated using different panels of clinical samples and their performance compared with standard platforms. This study has established that our integrated NAT chip system has a sensitivity and specificity comparable to that of the standard platforms while providing up to 10 fold reduction in sample/reagent volumes.
[Development of a Novel Liposomal DDS by Manipulating Pharmacokinetics and Intracellular Trafficking for Drug Therapy and Nucleic Acid Medicine].

Science.gov (United States)

Hatakeyama, Hiroto

2018-01-01

　Nucleic acid therapy is expected to be a next generation medicine. We recently developed a multifunctional envelope-type nano device (MEND) for use as a novel delivery system. The modification of polyethylene glycol (PEG), i.e., PEGylation, is useful for achieving the delivery of MENDs to tumors via an enhanced permeability and retention (EPR) effect. However, PEGylation strongly inhibits the cellular uptake and endosomal escape of MEND, which results in significant loss of action, and therefore lost effectiveness, of the cargo therapeutic. For successful nucleic acid delivery in cancer treatment, the crucial problem associated with the use of PEG, known as the "PEG dilemma", must be solved. In this review, we describe the development and application of MEND in overcoming the PEG dilemma based on manipulating both the pharmacokinetics and intracellular trafficking of cellular uptake and endosomal release using a cleavable PEG lipid, a pH-sensitive fusogenic peptide, and a pH-sensitive cationic lipid. We also developed dual-ligand liposomes with a controlled diameter of around 300 nm, then modified these with a specific ligand and a cell penetrating peptide designed to target the neovasculature of tumors. Dual-ligand liposomes could induce an anti-tumor effect in drug resistant tumors by delivering drugs to tumor blood vessels, rather than to the cancer cells themselves. Here, we review our recent efforts to develop a novel liposomal drug delivery system (DDS) by manipulating pharmacokinetics and intracellular trafficking for drug therapy and nucleic acid medicine.
Nucleic Acid Therapy: from humble beginnings a dynamic technology

CSIR Research Space (South Africa)

Millroy, L

2011-02-01

Full Text Available The term “nucleic acid therapy” encompasses a wide range of technologies for the treatment of a range of plant and animal ailments. As the name implies, it makes use of nucleic acid (either DNA or RNA) as a therapeutic agent. There are six branches...
A specific scenario for the origin of life and the genetic code based on peptide/oligonucleotide interdependence.

Science.gov (United States)

Griffith, Robert W

2009-12-01

Among various scenarios that attempt to explain how life arose, the RNA world is currently the most widely accepted scientific hypothesis among biologists. However, the RNA world is logistically implausible and doesn't explain how translation arose and DNA became incorporated into living systems. Here I propose an alternative hypothesis for life's origin based on cooperation between simple nucleic acids, peptides and lipids. Organic matter that accumulated on the prebiotic Earth segregated into phases in the ocean based on density and solubility. Synthesis of complex organic monomers and polymerization reactions occurred within a surface hydrophilic layer and at its aqueous and atmospheric interfaces. Replication of nucleic acids and translation of peptides began at the emulsified interface between hydrophobic and aqueous layers. At the core of the protobiont was a family of short nucleic acids bearing arginine's codon and anticodon that added this amino acid to pre-formed peptides. In turn, the survival and replication of nucleic acid was aided by the peptides. The arginine-enriched peptides served to sequester and transfer phosphate bond energy and acted as cohesive agents, aggregating nucleic acids and keeping them at the interface.
Artificial Specific Binders Directly Recovered from Chemically Modified Nucleic Acid Libraries

Directory of Open Access Journals (Sweden)

Yuuya Kasahara

2012-01-01

Full Text Available Specific binders comprised of nucleic acids, that is, RNA/DNA aptamers, are attractive functional biopolymers owing to their potential broad application in medicine, food hygiene, environmental analysis, and biological research. Despite the large number of reports on selection of natural DNA/RNA aptamers, there are not many examples of direct screening of chemically modified nucleic acid aptamers. This is because of (i the inferior efficiency and accuracy of polymerase reactions involving transcription/reverse-transcription of modified nucleotides compared with those of natural nucleotides, (ii technical difficulties and additional time and effort required when using modified nucleic acid libraries, and (iii ambiguous efficacies of chemical modifications in binding properties until recently; in contrast, the effects of chemical modifications on biostability are well studied using various nucleotide analogs. Although reports on the direct screening of a modified nucleic acid library remain in the minority, chemical modifications would be essential when further functional expansion of nucleic acid aptamers, in particular for medical and biological uses, is considered. This paper focuses on enzymatic production of chemically modified nucleic acids and their application to random screenings. In addition, recent advances and possible future research are also described.
Artificial specific binders directly recovered from chemically modified nucleic acid libraries.

Science.gov (United States)

Kasahara, Yuuya; Kuwahara, Masayasu

2012-01-01

Specific binders comprised of nucleic acids, that is, RNA/DNA aptamers, are attractive functional biopolymers owing to their potential broad application in medicine, food hygiene, environmental analysis, and biological research. Despite the large number of reports on selection of natural DNA/RNA aptamers, there are not many examples of direct screening of chemically modified nucleic acid aptamers. This is because of (i) the inferior efficiency and accuracy of polymerase reactions involving transcription/reverse-transcription of modified nucleotides compared with those of natural nucleotides, (ii) technical difficulties and additional time and effort required when using modified nucleic acid libraries, and (iii) ambiguous efficacies of chemical modifications in binding properties until recently; in contrast, the effects of chemical modifications on biostability are well studied using various nucleotide analogs. Although reports on the direct screening of a modified nucleic acid library remain in the minority, chemical modifications would be essential when further functional expansion of nucleic acid aptamers, in particular for medical and biological uses, is considered. This paper focuses on enzymatic production of chemically modified nucleic acids and their application to random screenings. In addition, recent advances and possible future research are also described.
Ionizing radiation induced attachment reactions of nucleic acids and their components

International Nuclear Information System (INIS)

Myers, L.S. Jr.

1975-01-01

An extensive bibliographic review is given of experimental and theoretical data on radiation-induced attachment reactions of nucleic acids and their components. Mechanisms of these reactions are reviewed. The reactions with water, formate, and alcohols, with amines and other small molecules, and with radiation sensitizers and nucleic acid-nucleic acid reactions are discussed. Studies of the reaction mechanisms show that many of the reactions occur by radical-molecule reactions, but radical-radical reactions also occur. Radiation modifiers become attached to nucleic acids in vitro and in vivo and there are indications that attachment may be necessary for the action of some sensitizers. (U.S.)
Experimental and theoretical studies on extraction of actinides and lanthanides by alicyclic H-phosphonates

Energy Technology Data Exchange (ETDEWEB)

Annam, Suresh; Sivaramakrishna, Akella; Vijayakrishna, Kari [VIT Univ., Tamil Nadu (India). Dept. of Chemistry; Gopakumar, Gopinadhanpillai; Rao, C.V.S. Brahmmananda; Sivaraman, N. [Indira Gandhi Centre for Atomic Research (IGCAR), Tamil Nadu (India). Chemistry Group

2017-06-01

Three different alicyclic substituents H-phosphonates, namely, dicyclopentyl H-phosphonate, dicyclohexyl H-phosphonate and dimenthyl H-phosphonate were synthesized and used for liquid-liquid extraction of actinide elements (U, Am and Th) and lanthanide (Gd) in n-dodecane from nitric acid medium. The physicochemical properties of the extractants, such as density, viscosity, solubility were determined. At lower acidities, these H-phosphonates exhibit higher distribution values and the extraction following cation exchange mechanism through P-OH group of tri-coordinated phosphite form. At higher acidities (2N), the extraction is primarily via solvation mechanism through P=O group of penta-coordinated phosphonate form. Amongst the three H-phosphonates, examined dimenthyl H-phosphonate showed the best results for the actinide extraction. Density functional theory (DFT) calculations were applied to understand the electronic structure of the ligands and the metal complexes. The calculated large complexation energy of UO{sub 2}(NO{sub 3}){sub 2}.@2DMnHP is in agreement with the observed trend in experimental distribution ratio data.
BGL6 beta-glucosidase and nucleic acids encoding the same

Science.gov (United States)

Dunn-Coleman, Nigel [Los Gatos, CA; Ward, Michael [San Francisco, CA

2009-09-01

The present invention provides a novel .beta.-glucosidase nucleic acid sequence, designated bgl6, and the corresponding BGL6 amino acid sequence. The invention also provides expression vectors and host cells comprising a nucleic acid sequence encoding BGL6, recombinant BGL6 proteins and methods for producing the same.
Biological activity and biotechnological aspects of locked nucleic acids

DEFF Research Database (Denmark)

Lundin, Karin E; Højland, Torben; Hansen, Bo

2013-01-01

Locked nucleic acid (LNA) is one of the most promising new nucleic acid analogues that has been produced under the past two decades. In this chapter, we have tried to cover many of the different areas, where this molecule has been used to improve the function of synthetic oligonucleotides (ONs). ...
Biogenic and Synthetic Peptides with Oppositely Charged Amino Acids as Binding Sites for Mineralization.

Science.gov (United States)

Lemloh, Marie-Louise; Altintoprak, Klara; Wege, Christina; Weiss, Ingrid M; Rothenstein, Dirk

2017-01-28

Proteins regulate diverse biological processes by the specific interaction with, e.g., nucleic acids, proteins and inorganic molecules. The generation of inorganic hybrid materials, such as shell formation in mollusks, is a protein-controlled mineralization process. Moreover, inorganic-binding peptides are attractive for the bioinspired mineralization of non-natural inorganic functional materials for technical applications. However, it is still challenging to identify mineral-binding peptide motifs from biological systems as well as for technical systems. Here, three complementary approaches were combined to analyze protein motifs consisting of alternating positively and negatively charged amino acids: (i) the screening of natural biomineralization proteins; (ii) the selection of inorganic-binding peptides derived from phage display; and (iii) the mineralization of tobacco mosaic virus (TMV)-based templates. A respective peptide motif displayed on the TMV surface had a major impact on the SiO₂ mineralization. In addition, similar motifs were found in zinc oxide- and zirconia-binding peptides indicating a general binding feature. The comparative analysis presented here raises new questions regarding whether or not there is a common design principle based on acidic and basic amino acids for peptides interacting with minerals.
Miniaturized isothermal nucleic acid amplification, a review.

Science.gov (United States)

Asiello, Peter J; Baeumner, Antje J

2011-04-21

Micro-Total Analysis Systems (µTAS) for use in on-site rapid detection of DNA or RNA are increasingly being developed. Here, amplification of the target sequence is key to increasing sensitivity, enabling single-cell and few-copy nucleic acid detection. The several advantages to miniaturizing amplification reactions and coupling them with sample preparation and detection on the same chip are well known and include fewer manual steps, preventing contamination, and significantly reducing the volume of expensive reagents. To-date, the majority of miniaturized systems for nucleic acid analysis have used the polymerase chain reaction (PCR) for amplification and those systems are covered in previous reviews. This review provides a thorough overview of miniaturized analysis systems using alternatives to PCR, specifically isothermal amplification reactions. With no need for thermal cycling, isothermal microsystems can be designed to be simple and low-energy consuming and therefore may outperform PCR in portable, battery-operated detection systems in the future. The main isothermal methods as miniaturized systems reviewed here include nucleic acid sequence-based amplification (NASBA), loop-mediated isothermal amplification (LAMP), helicase-dependent amplification (HDA), rolling circle amplification (RCA), and strand displacement amplification (SDA). Also, important design criteria for the miniaturized devices are discussed. Finally, the potential of miniaturization of some new isothermal methods such as the exponential amplification reaction (EXPAR), isothermal and chimeric primer-initiated amplification of nucleic acids (ICANs), signal-mediated amplification of RNA technology (SMART) and others is presented.

Novel approaches to tumor imaging in mice: pre targeting with radiolabeled peptide nucleic acid

International Nuclear Information System (INIS)

Hnatowich, D.J.; Qu, T.; Chang, F.; Rusckowski, M.

1997-01-01

Full text.Since targeting of tumour by conventional methods is not consistently favorable, we have considered pre targeting with separate administrations of anti tumour antibody and radiolabel. As an alternative to streptavidin and biotin for this application, we earlier considered single stranded peptide nucleic acid (PNA) bound to an irrelevant protein administered first and allowed to diffuse non specifically into tumour. This was followed later by the administration of 99 m Tc labeled complementary PNA. We now report on the first studies with PNA conjugated anti tumour antibody to allow specific binding. PNA was conjugated to the NRLU-10 IgG antibody while the complementary PNA (amine derivatized) was labeled with ((m Tc using MAG3. LS174T tumour-bearing nude mice received IV 200 ug of the PNA-antibody conjugate and 20 h later, received IV 100 ug (130 uCl) of 99m Tc- complementary PNA. Animals were imaged and sacrificed 5 h later. Because of rapid clearance, at sacrifice all tissue levels of 99 m Tc were low, the highest being kidneys at about 4%ID/gm. Tumour uptake was 0.55%ID/gm for the study animals vs. 0. 13 for controls and tumour/muscle ratios were 9.8 vs. 3.6 respectively. These values represent a 2.5-fold improvement in localization over the nonspecific study. The whole body images also reflected the superior targeting of study vs. control animals. We conclude that single-stranded PNAs should be a useful alternative to streptavidin and biotin for pre targeting studies
BIOPHYSICAL PROPERTIES OF NUCLEIC ACIDS AT SURFACES RELEVANT TO MICROARRAY PERFORMANCE.

Science.gov (United States)

Rao, Archana N; Grainger, David W

2014-04-01

Both clinical and analytical metrics produced by microarray-based assay technology have recognized problems in reproducibility, reliability and analytical sensitivity. These issues are often attributed to poor understanding and control of nucleic acid behaviors and properties at solid-liquid interfaces. Nucleic acid hybridization, central to DNA and RNA microarray formats, depends on the properties and behaviors of single strand (ss) nucleic acids (e.g., probe oligomeric DNA) bound to surfaces. ssDNA's persistence length, radius of gyration, electrostatics, conformations on different surfaces and under various assay conditions, its chain flexibility and curvature, charging effects in ionic solutions, and fluorescent labeling all influence its physical chemistry and hybridization under assay conditions. Nucleic acid (e.g., both RNA and DNA) target interactions with immobilized ssDNA strands are highly impacted by these biophysical states. Furthermore, the kinetics, thermodynamics, and enthalpic and entropic contributions to DNA hybridization reflect global probe/target structures and interaction dynamics. Here we review several biophysical issues relevant to oligomeric nucleic acid molecular behaviors at surfaces and their influences on duplex formation that influence microarray assay performance. Correlation of biophysical aspects of single and double-stranded nucleic acids with their complexes in bulk solution is common. Such analysis at surfaces is not commonly reported, despite its importance to microarray assays. We seek to provide further insight into nucleic acid-surface challenges facing microarray diagnostic formats that have hindered their clinical adoption and compromise their research quality and value as genomics tools.
BIOPHYSICAL PROPERTIES OF NUCLEIC ACIDS AT SURFACES RELEVANT TO MICROARRAY PERFORMANCE

Science.gov (United States)

Rao, Archana N.; Grainger, David W.

2014-01-01

Both clinical and analytical metrics produced by microarray-based assay technology have recognized problems in reproducibility, reliability and analytical sensitivity. These issues are often attributed to poor understanding and control of nucleic acid behaviors and properties at solid-liquid interfaces. Nucleic acid hybridization, central to DNA and RNA microarray formats, depends on the properties and behaviors of single strand (ss) nucleic acids (e.g., probe oligomeric DNA) bound to surfaces. ssDNA’s persistence length, radius of gyration, electrostatics, conformations on different surfaces and under various assay conditions, its chain flexibility and curvature, charging effects in ionic solutions, and fluorescent labeling all influence its physical chemistry and hybridization under assay conditions. Nucleic acid (e.g., both RNA and DNA) target interactions with immobilized ssDNA strands are highly impacted by these biophysical states. Furthermore, the kinetics, thermodynamics, and enthalpic and entropic contributions to DNA hybridization reflect global probe/target structures and interaction dynamics. Here we review several biophysical issues relevant to oligomeric nucleic acid molecular behaviors at surfaces and their influences on duplex formation that influence microarray assay performance. Correlation of biophysical aspects of single and double-stranded nucleic acids with their complexes in bulk solution is common. Such analysis at surfaces is not commonly reported, despite its importance to microarray assays. We seek to provide further insight into nucleic acid-surface challenges facing microarray diagnostic formats that have hindered their clinical adoption and compromise their research quality and value as genomics tools. PMID:24765522
Prediction of molecular alignment of nucleic acids in aligned media

International Nuclear Information System (INIS)

Wu Bin; Petersen, Michael; Girard, Frederic; Tessari, Marco; Wijmenga, Sybren S.

2006-01-01

We demonstrate - using the data base of all deposited DNA and RNA structures aligned in Pf1-medium and RDC refined - that for nucleic acids in a Pf1-medium the electrostatic alignment tensor can be predicted reliably and accurately via a simple and fast calculation based on the gyration tensor spanned out by the phosphodiester atoms. The rhombicity is well predicted over its full range from 0 to 0.66, while the alignment tensor orientation is predicted correctly for rhombicities up to ca. 0.4, for larger rhombicities it appears to deviate somewhat more than expected based on structural noise and measurement error. This simple analytical approach is based on the Debye-Huckel approximation for the electrostatic interaction potential, valid at distances sufficiently far away from a poly-ionic charged surface, a condition naturally enforced when the charge of alignment medium and solute are of equal sign, as for nucleic acids in a Pf1-phage medium. For the usual salt strengths and nucleic acid sizes, the Debye-Huckel screening length is smaller than the nucleic acid size, but large enough for the collective of Debye-Huckel spheres to encompass the whole molecule. The molecular alignment is then purely electrostatic, but it's functional form is under these conditions similar to that for steric alignment. The proposed analytical expression allows for very fast calculation of the alignment tensor and hence RDCs from the conformation of the nucleic acid molecule. This information provides opportunities for improved structure determination of nucleic acids, including better assessment of dynamics in (multi-domain) nucleic acids and the possibility to incorporate alignment tensor prediction from shape directly into the structure calculation process. The procedures are incorporated into MATLAB scripts, which are available on request
On the Sensitivity of Peptide Nucleic Acid Duplex Formation and Crystal Dissolution to a Variation of Force-Field Parameters.

Science.gov (United States)

Bachmann, Stephan J; Lin, Zhixiong; Stafforst, Thorsten; van Gunsteren, Wilfred F; Dolenc, Jožica

2014-01-14

The technique of one-step perturbation to explore the relation between particular force-field parameters on the one hand and particular properties of a biomolecular system on the other hand from one or a few molecular dynamics simulations is applied to investigate the dependence of the free enthalpy of dimer formation and of crystal dissolution of a self-complementary fragment (H-CGTACG-NH2) of peptide nucleic acid, PNA, a mimic of DNA. The simulations show that PNA dimer formation in aqueous solution is favored by a decrease in the base charges with respect to values of the GROMOS 45A4 force field, while it is disfavored by a decrease in the backbone charges. In contrast, crystal dissolution of the PNA dimer is favored by a decrease in base charges, while a variation of backbone charges has a minor effect on this free enthalpy change. These opposite effects in a crystalline versus aqueous solution environment can be understood from the different water contents for these systems and have consequences for biomolecular force-field development.
Rapid Identification of Staphylococcus aureus Directly from Blood Cultures by Fluorescence In Situ Hybridization with Peptide Nucleic Acid Probes

Science.gov (United States)

Oliveira, Kenneth; Procop, Gary W.; Wilson, Deborah; Coull, James; Stender, Henrik

2002-01-01

A new fluorescence in situ hybridization (FISH) method with peptide nucleic acid (PNA) probes for identification of Staphylococcus aureus directly from positive blood culture bottles that contain gram-positive cocci in clusters (GPCC) is described. The test (the S. aureus PNA FISH assay) is based on a fluorescein-labeled PNA probe that targets a species-specific sequence of the 16S rRNA of S. aureus. Evaluations with 17 reference strains and 48 clinical isolates, including methicillin-resistant and methicillin-susceptible S. aureus species, coagulase-negative Staphylococcus species, and other clinically relevant and phylogenetically related bacteria and yeast species, showed that the assay had 100% sensitivity and 96% specificity. Clinical trials with 87 blood cultures positive for GPCC correctly identified 36 of 37 (97%) of the S. aureus-positive cultures identified by standard microbiological methods. The positive and negative predictive values were 100 and 98%, respectively. It is concluded that this rapid method (2.5 h) for identification of S. aureus directly from blood culture bottles that contain GPCC offers important information for optimal antibiotic therapy. PMID:11773123
BIOPHYSICAL PROPERTIES OF NUCLEIC ACIDS AT SURFACES RELEVANT TO MICROARRAY PERFORMANCE

OpenAIRE

Rao, Archana N.; Grainger, David W.

2014-01-01

Both clinical and analytical metrics produced by microarray-based assay technology have recognized problems in reproducibility, reliability and analytical sensitivity. These issues are often attributed to poor understanding and control of nucleic acid behaviors and properties at solid-liquid interfaces. Nucleic acid hybridization, central to DNA and RNA microarray formats, depends on the properties and behaviors of single strand (ss) nucleic acids (e.g., probe oligomeric DNA) bound to surface...
Soni-removal of nucleic acids from inclusion bodies.

Science.gov (United States)

Neerathilingam, Muniasamy; Mysore, Sumukh; Gandham, Sai Hari A

2014-05-23

Inclusion bodies (IBs) are commonly formed in Escherichia coli due to over expression of recombinant proteins in non-native state. Isolation, denaturation and refolding of these IBs is generally performed to obtain functional protein. However, during this process IBs tend to form non-specific interactions with sheared nucleic acids from the genome, thus getting carried over into downstream processes. This may hinder the refolding of IBs into their native state. To circumvent this, we demonstrate a methodology termed soni-removal which involves disruption of nucleic acid-inclusion body interaction using sonication; followed by solvent based separation. As opposed to conventional techniques that use enzymes and column-based separations, soni-removal is a cost effective alternative for complete elimination of buried and/or strongly bound short nucleic acid contaminants from IBs. Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.
"Clickable" LNA/DNA probes for fluorescence sensing of nucleic acids and autoimmune antibodies

DEFF Research Database (Denmark)

Jørgensen, Anna S; Gupta, Pankaj; Wengel, Jesper

2013-01-01

Herein we describe fluorescent oligonucleotides prepared by click chemistry between novel alkyne-modified locked nucleic acid (LNA) strands and a series of fluorescent azides for homogeneous (all-in-solution) detection of nucleic acids and autoimmune antibodies.......Herein we describe fluorescent oligonucleotides prepared by click chemistry between novel alkyne-modified locked nucleic acid (LNA) strands and a series of fluorescent azides for homogeneous (all-in-solution) detection of nucleic acids and autoimmune antibodies....
Revealing Nucleic Acid Mutations Using Förster Resonance Energy Transfer-Based Probes

Directory of Open Access Journals (Sweden)

Nina P. L. Junager

2016-07-01

Full Text Available Nucleic acid mutations are of tremendous importance in modern clinical work, biotechnology and in fundamental studies of nucleic acids. Therefore, rapid, cost-effective and reliable detection of mutations is an object of extensive research. Today, Förster resonance energy transfer (FRET probes are among the most often used tools for the detection of nucleic acids and in particular, for the detection of mutations. However, multiple parameters must be taken into account in order to create efficient FRET probes that are sensitive to nucleic acid mutations. In this review; we focus on the design principles for such probes and available computational methods that allow for their rational design. Applications of advanced, rationally designed FRET probes range from new insights into cellular heterogeneity to gaining new knowledge of nucleic acid structures directly in living cells.
Solving nucleic acid structures by molecular replacement: examples from group II intron studies

International Nuclear Information System (INIS)

Marcia, Marco; Humphris-Narayanan, Elisabeth; Keating, Kevin S.; Somarowthu, Srinivas; Rajashankar, Kanagalaghatta; Pyle, Anna Marie

2013-01-01

Strategies for phasing nucleic acid structures by molecular replacement, using both experimental and de novo designed models, are discussed. Structured RNA molecules are key players in ensuring cellular viability. It is now emerging that, like proteins, the functions of many nucleic acids are dictated by their tertiary folds. At the same time, the number of known crystal structures of nucleic acids is also increasing rapidly. In this context, molecular replacement will become an increasingly useful technique for phasing nucleic acid crystallographic data in the near future. Here, strategies to select, create and refine molecular-replacement search models for nucleic acids are discussed. Using examples taken primarily from research on group II introns, it is shown that nucleic acids are amenable to different and potentially more flexible and sophisticated molecular-replacement searches than proteins. These observations specifically aim to encourage future crystallographic studies on the newly discovered repertoire of noncoding transcripts
Antibacterial Peptide Nucleic Acid-Antimicrobial Peptide (PNA-AMP) Conjugates

DEFF Research Database (Denmark)

Hansen, Anna Mette; Bonke, Gitte; Larsen, Camilla Josephine

2016-01-01

. In the present study we show that antimicrobial peptides (AMPs) with an intracellular mode of action can be efficient vehicles for bacterial delivery of an antibacterial PNA targeting the essential acpP gene. The results demonstrate that buforin 2-A (BF2-A), drosocin, oncocin 10, Pep-1-K, KLW-9,13-a, (P59→W59...
Nucleic acids for the rational design of reaction circuits.

Science.gov (United States)

Padirac, Adrien; Fujii, Teruo; Rondelez, Yannick

2013-08-01

Nucleic acid-based circuits are rationally designed in vitro assemblies that can perform complex preencoded programs. They can be used to mimic in silico computations. Recent works emphasized the modularity and robustness of these circuits, which allow their scaling-up. Another new development has led to dynamic, time-responsive systems that can display emergent behaviors like oscillations. These are closely related to biological architectures and provide an in vitro model of in vivo information processing. Nucleic acid circuits have already been used to handle various processes for technological or biotechnological purposes. Future applications of these chemical smart systems will benefit from the rapidly growing ability to design, construct, and model nucleic acid circuits of increasing size. Copyright © 2012 Elsevier Ltd. All rights reserved.
Polymerase chain reaction system using magnetic beads for analyzing a sample that includes nucleic acid

Science.gov (United States)

Nasarabadi, Shanavaz [Livermore, CA

2011-01-11

A polymerase chain reaction system for analyzing a sample containing nucleic acid includes providing magnetic beads; providing a flow channel having a polymerase chain reaction chamber, a pre polymerase chain reaction magnet position adjacent the polymerase chain reaction chamber, and a post pre polymerase magnet position adjacent the polymerase chain reaction chamber. The nucleic acid is bound to the magnetic beads. The magnetic beads with the nucleic acid flow to the pre polymerase chain reaction magnet position in the flow channel. The magnetic beads and the nucleic acid are washed with ethanol. The nucleic acid in the polymerase chain reaction chamber is amplified. The magnetic beads and the nucleic acid are separated into a waste stream containing the magnetic beads and a post polymerase chain reaction mix containing the nucleic acid. The reaction mix containing the nucleic acid flows to an analysis unit in the channel for analysis.
Chirality as a tool in nucleic acid recognition: principles and relevance in biotechnology and in medicinal chemistry.

Science.gov (United States)

Corradini, Roberto; Sforza, Stefano; Tedeschi, Tullia; Marchelli, Rosangela

2007-05-05

The understanding of the interaction of chiral species with DNA or RNA is very important for the development of new tools in biology and of new drugs. Several cases in which chirality is a crucial point in determining the DNA binding mode are reviewed and discussed, with the aim of illustrating how chirality can be considered as a tool for improving the understanding of mechanisms and the effectiveness of nucleic acid recognition. The review is divided into two parts: the former describes examples of chiral species interacting with DNA: intercalators, metal complexes, and groove binders; the latter part is dedicated to chirality in DNA analogs, with discussion of phosphate stereochemistry and chirality of ribose substitutes, in particular of peptide nucleic acids (PNAs) for which a number of works have been published recently dealing with the effect of chirality in DNA recognition. The discussion is intended to show how enantiomeric recognition originates at the molecular level, by exploiting the enormous progresses recently achieved in the field of structural characterization of complexes formed by nucleic acid with their ligands by crystallographic and spectroscopic methods. Examples of application of the DNA binding molecules described and the role of chirality in DNA recognition relevant for biotechnology or medicinal chemistry are reported. (c) 2007 Wiley-Liss, Inc.
Extraction of plutonium and uranium from oxalate bearing solutions using phosphonic acid

International Nuclear Information System (INIS)

Godbole, A.G.; Mapara, P.M.; Swarup, Rajendra

1995-01-01

A feasibility study on the solvent extraction of plutonium and uranium from solutions containing oxalic and nitric acids using a phosphonic acid extractant (PC88A) was made to explore the possibility of recovering Pu from these solutions. Batch experiments on the extraction of Pu(IV) and U(VI) under different parameters were carried out using PC88A in dodecane. The results indicated that Pu could be extracted quantitatively by PC88A from these solutions. A good separation of Pu from U could be achieved at higher temperatures. (author). 6 refs., 3 tabs
Similarities and differences in the nucleic acid chaperone activity of HIV-2 and HIV-1 nucleocapsid proteins in vitro.

Science.gov (United States)

Pachulska-Wieczorek, Katarzyna; Stefaniak, Agnieszka K; Purzycka, Katarzyna J

2014-07-03

The nucleocapsid domain of Gag and mature nucleocapsid protein (NC) act as nucleic acid chaperones and facilitate folding of nucleic acids at critical steps of retroviral replication cycle. The basic N-terminus of HIV-1 NC protein was shown most important for the chaperone activity. The HIV-2 NC (NCp8) and HIV-1 NC (NCp7) proteins possess two highly conserved zinc fingers, flanked by basic residues. However, the NCp8 N-terminal domain is significantly shorter and contains less positively charged residues. This study characterizes previously unknown, nucleic acid chaperone activity of the HIV-2 NC protein. We have comparatively investigated the in vitro nucleic acid chaperone properties of the HIV-2 and HIV-1 NC proteins. Using substrates derived from the HIV-1 and HIV-2 genomes, we determined the ability of both proteins to chaperone nucleic acid aggregation, annealing and strand exchange in duplex structures. Both NC proteins displayed comparable, high annealing activity of HIV-1 TAR DNA and its complementary nucleic acid. Interesting differences between the two NC proteins were discovered when longer HIV substrates, particularly those derived from the HIV-2 genome, were used in chaperone assays. In contrast to NCp7, NCp8 weakly facilitates annealing of HIV-2 TAR RNA to its complementary TAR (-) DNA. NCp8 is also unable to efficiently stimulate tRNALys3 annealing to its respective HIV-2 PBS motif. Using truncated NCp8 peptide, we demonstrated that despite the fact that the N-terminus of NCp8 differs from that of NCp7, this domain is essential for NCp8 activity. Our data demonstrate that the HIV-2 NC protein displays reduced nucleic acid chaperone activity compared to that of HIV-1 NC. We found that NCp8 activity is limited by substrate length and stability to a greater degree than that of NCp7. This is especially interesting in light of the fact that the HIV-2 5'UTR is more structured than that of HIV-1. The reduced chaperone activity observed with NCp8 may
Helicase-dependent amplification of nucleic acids.

Science.gov (United States)

Cao, Yun; Kim, Hyun-Jin; Li, Ying; Kong, Huimin; Lemieux, Bertrand

2013-10-11

Helicase-dependent amplification (HDA) is a novel method for the isothermal in vitro amplification of nucleic acids. The HDA reaction selectively amplifies a target sequence by extension of two oligonucleotide primers. Unlike the polymerase chain reaction (PCR), HDA uses a helicase enzyme to separate the deoxyribonucleic acid (DNA) strands, rather than heat denaturation. This allows DNA amplification without the need for thermal cycling. The helicase used in HDA is a helicase super family II protein obtained from a thermophilic organism, Thermoanaerobacter tengcongensis (TteUvrD). This thermostable helicase is capable of unwinding blunt-end nucleic acid substrates at elevated temperatures (60° to 65°C). The HDA reaction can also be coupled with reverse transcription for ribonucleic acid (RNA) amplification. The products of this reaction can be detected during the reaction using fluorescent probes when incubations are conducted in a fluorimeter. Alternatively, products can be detected after amplification using a disposable amplicon containment device that contains an embedded lateral flow strip. Copyright © 2013 John Wiley & Sons, Inc.
Ion-pair chromatography of nucleic acid derivatives

International Nuclear Information System (INIS)

Perrone, P.A.; Brown, P.R.

1985-01-01

Little work has been done on the ion-pair chromatography of nucleic acid constituents, although there is a great potential for the use of this technique in the field. Since the classic work in 1949, nucleotides, as well as nucleosides and bases, have been separated by ion-exchange chromatography. However, ion exchange is a difficult mode and most researchers prefer the use of reversed-phase whenever possible. Although reversed-phase is now the method of choice, ionic compounds like nucleotides and some of the more polar bases are not adequately retained by many systems of this type. In addition, it is difficult to analyze simultaneously members of all three classes of nucleic acid compounds (bases, nucleosides, and nucleotides) using a reversed-phase system, even with gradient elution. Ion pairing can be a useful technique because, theoretically, the separation of nonionic bases and nucleosides along with the ionic nucleotides can be achieved. Additionally, each group of compounds may be separated isocratically. In this chapter, they will discuss ion-pair chromatography as applied to nucleic acid constituents. The current theories, advantages and disadvantages, a limited number of applications, and potential for future work are presented
Study of ion exchange behaviour of some elements at phosphonic-acid cationite in diluted solutions of nitric acid

International Nuclear Information System (INIS)

Razbash, A.A.; Sevast'yanov, Yu.G.

1985-01-01

Ce(3, 4), Eu(3), Gd(3), Sm(3), Sc(3) distribution coefficients are determined in the macroporic phosphonic acid cationite KRF-20T-60 in nitric acid solutions in 0.1-2.0 M concentration interval using statistical method. A simple method of cerium-139 radionuclide extraction from the industrial lanthanum target is developed. The product yield made up more than 99%, specific activity - 2.37x10 9 Bq/mg, radiochemical purity - no less than 99.9%

Methods of introducing nucleic acids into cellular DNA

Energy Technology Data Exchange (ETDEWEB)

Lajoie, Marc J.; Gregg, Christopher J.; Mosberg, Joshua A.; Church, George M.

2017-06-27

A method of introducing a nucleic acid sequence into a cell is provided where the cell has impaired or inhibited or disrupted DnaG primase activity or impaired or inhibited or disrupted DnaB helicase activity, or larger or increased gaps or distance between Okazaki fragments or lowered or reduced frequency of Okazaki fragment initiation, or the cell has increased single stranded DNA (ssDNA) on the lagging strand of the replication fork including transforming the cell through recombination with a nucleic acid oligomer.
Digital Microfluidics for Nucleic Acid Amplification

Directory of Open Access Journals (Sweden)

Beatriz Coelho

2017-06-01

Full Text Available Digital Microfluidics (DMF has emerged as a disruptive methodology for the control and manipulation of low volume droplets. In DMF, each droplet acts as a single reactor, which allows for extensive multiparallelization of biological and chemical reactions at a much smaller scale. DMF devices open entirely new and promising pathways for multiplex analysis and reaction occurring in a miniaturized format, thus allowing for healthcare decentralization from major laboratories to point-of-care with accurate, robust and inexpensive molecular diagnostics. Here, we shall focus on DMF platforms specifically designed for nucleic acid amplification, which is key for molecular diagnostics of several diseases and conditions, from pathogen identification to cancer mutations detection. Particular attention will be given to the device architecture, materials and nucleic acid amplification applications in validated settings.
Nucleic Acid Extraction from Synthetic Mars Analog Soils for in situ Life Detection.

Science.gov (United States)

Mojarro, Angel; Ruvkun, Gary; Zuber, Maria T; Carr, Christopher E

2017-08-01

Biological informational polymers such as nucleic acids have the potential to provide unambiguous evidence of life beyond Earth. To this end, we are developing an automated in situ life-detection instrument that integrates nucleic acid extraction and nanopore sequencing: the Search for Extra-Terrestrial Genomes (SETG) instrument. Our goal is to isolate and determine the sequence of nucleic acids from extant or preserved life on Mars, if, for example, there is common ancestry to life on Mars and Earth. As is true of metagenomic analysis of terrestrial environmental samples, the SETG instrument must isolate nucleic acids from crude samples and then determine the DNA sequence of the unknown nucleic acids. Our initial DNA extraction experiments resulted in low to undetectable amounts of DNA due to soil chemistry-dependent soil-DNA interactions, namely adsorption to mineral surfaces, binding to divalent/trivalent cations, destruction by iron redox cycling, and acidic conditions. Subsequently, we developed soil-specific extraction protocols that increase DNA yields through a combination of desalting, utilization of competitive binders, and promotion of anaerobic conditions. Our results suggest that a combination of desalting and utilizing competitive binders may establish a "universal" nucleic acid extraction protocol suitable for analyzing samples from diverse soils on Mars. Key Words: Life-detection instruments-Nucleic acids-Mars-Panspermia. Astrobiology 17, 747-760.
Nucleic acid purification from plants, animals and microbes in under 30 seconds.

Directory of Open Access Journals (Sweden)

Yiping Zou

2017-11-01

Full Text Available Nucleic acid amplification is a powerful molecular biology tool, although its use outside the modern laboratory environment is limited due to the relatively cumbersome methods required to extract nucleic acids from biological samples. To address this issue, we investigated a variety of materials for their suitability for nucleic acid capture and purification. We report here that untreated cellulose-based paper can rapidly capture nucleic acids within seconds and retain them during a single washing step, while contaminants present in complex biological samples are quickly removed. Building on this knowledge, we have successfully created an equipment-free nucleic acid extraction dipstick methodology that can obtain amplification-ready DNA and RNA from plants, animals, and microbes from difficult biological samples such as blood and leaves from adult trees in less than 30 seconds. The simplicity and speed of this method as well as the low cost and availability of suitable materials (e.g., common paper towelling, means that nucleic acid extraction is now more accessible and affordable for researchers and the broader community. Furthermore, when combined with recent advancements in isothermal amplification and naked eye DNA visualization techniques, the dipstick extraction technology makes performing molecular diagnostic assays achievable in limited resource settings including university and high school classrooms, field-based environments, and developing countries.
[Blood Test Patterns for Blood Donors after Nucleic Acid Detection in the Blood Center].

Science.gov (United States)

Men, Shou-Shan; Lv, Lian-Zhi; Chen, Yuan-Feng; Han, Chun-Hua; Liu, Hong-Yu; Yan, Yan

2017-12-01

To investigate the blood test patterns for blood donors after nucleic acid detection in blood center. The collected blood samples after voluntary blood donors first were detected by conventional ELISA, then 31981 negative samples were detected via HBV/HCV/HIV combined nucleic acid test of 6 mixed samples(22716 cases) or single samples(9265 cases) by means of Roche cobas s201 instrument. The combined detection method as follows: the blood samples were assayed by conventional nucleic acid test of 6 mixed samples, at same time, 6 mixed samples were treated with polyethylene glycol precipitation method to concentrate the virus, then the nucleic acid test of blood samples was performed; the single detection method as follows: firstly the conventional nucleic acid test of single sample was performed, then the positive reactive samples after re-examination were 6-fold diluted to simulate the nucleic acid test of 6-mixed samples. The positive rate of positive samples detected by combined nucleic acid test, positive samples detected by nucleic acid test of mixed virus concentration and positive samples detected by single nucleic acid test was statistically analyzed. In addition, for HBV + persons the serological test yet should be performed. In 22 716 samples detected by nucleic acid test of 6 mixed samples (MP-6-NAT) , 9 cases were HBV + (0.40‰, 9/22716); at same time, the detection of same samples by nucleic acid test of mixed sample virus concentration showed 29 cases of HBV + (1.28‰, 29/22716). In 9265 samples detected by single nucleic acid test(ID-NAT) 12 cases showed HBV + (1.30‰, 12/9265), meanwhile the detection of these 12 samples with HBV + by 6-fold dilution for virus concentration found only 4 samples with HBV + . In serological qualified samples, ID-NAT unqualified rate was 1.28‰, which was higher than that of MP-6-NAT(0.4‰) (χ 2 =8.11, P0.05). In 41 samples with HBsAg - HBV DNA + detected by ELISA, 36 samples were confirmed to be occult HBV
Programming the Assembly of Unnatural Materials with Nucleic Acids

Science.gov (United States)

Mirkin, Chad

Nature directs the assembly of enormously complex and highly functional materials through an encoded class of biomolecules, nucleic acids. The establishment of a similarly programmable code for the construction of synthetic, unnatural materials would allow researchers to impart functionality by precisely positioning all material components. Although it is exceedingly difficult to control the complex interactions between atomic and molecular species in such a manner, interactions between nanoscale components can be directed through the ligands attached to their surface. Our group has shown that nucleic acids can be used as highly programmable surface ligands to control the spacing and symmetry of nanoparticle building blocks in structurally sophisticated and functional materials. These nucleic acids function as programmable ``bonds'' between nanoparticle ``atoms,'' analogous to a nanoscale genetic code for assembling materials. The sequence and length tunability of nucleic acid bonds has allowed us to define a powerful set of design rules for the construction of nanoparticle superlattices with more than 30 unique lattice symmetries, tunable defect structures and interparticle spacings, and several well-defined crystal habits. Further, the nature of the nucleic acid bond enables an additional level of structural control: temporal regulation of dynamic material response to external biomolecular and chemical stimuli. This control allows for the reversible transformation between thermodynamic states with different crystal symmetries, particle stoichiometries, thermal stabilities, and interparticle spacings on demand. Notably, our unique genetic approach affords functional nanoparticle architectures that, among many other applications, can be used to systematically explore and manipulate optoelectronic material properties, such as tunable interparticle plasmonic interactions, microstructure-directed energy emission, and coupled plasmonic and photonic modes.
A DNA origami nanorobot controlled by nucleic acid hybridization.

Science.gov (United States)

Torelli, Emanuela; Marini, Monica; Palmano, Sabrina; Piantanida, Luca; Polano, Cesare; Scarpellini, Alice; Lazzarino, Marco; Firrao, Giuseppe

2014-07-23

A prototype for a DNA origami nanorobot is designed, produced, and tested. The cylindrical nanorobot (diameter of 14 nm and length of 48 nm) with a switchable flap, is able to respond to an external stimulus and reacts by a physical switch from a disarmed to an armed configuration able to deliver a cellular compatible message. In the tested design the robot weapon is a nucleic acid fully contained in the inner of the tube and linked to a single point of the internal face of the flap. Upon actuation the nanorobot moves the flap extracting the nucleic acid that assembles into a hemin/G-quadruplex horseradish peroxidase mimicking DNAzyme catalyzing a colorimetric reaction or chemiluminescence generation. The actuation switch is triggered by an external nucleic acid (target) that interacts with a complementary nucleic acid that is beard externally by the nanorobot (probe). Hybridization of probe and target produces a localized structural change that results in flap opening. The flap movement is studied on a two-dimensional prototype origami using Förster resonance energy transfer and is shown to be triggered by a variety of targets, including natural RNAs. The nanorobot has potential for in vivo biosensing and intelligent delivery of biological activators. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.
A DNA origami nanorobot controlled by nucleic acid hybridization

KAUST Repository

Torelli, Emanuela

2014-03-20

A prototype for a DNA origami nanorobot is designed, produced, and tested. The cylindrical nanorobot (diameter of 14 nm and length of 48 nm) with a switchable flap, is able to respond to an external stimulus and reacts by a physical switch from a disarmed to an armed configuration able to deliver a cellular compatible message. In the tested design the robot weapon is a nucleic acid fully contained in the inner of the tube and linked to a single point of the internal face of the flap. Upon actuation the nanorobot moves the flap extracting the nucleic acid that assembles into a hemin/G-quadruplex horseradish peroxidase mimicking DNAzyme catalyzing a colorimetric reaction or chemiluminescence generation. The actuation switch is triggered by an external nucleic acid (target) that interacts with a complementary nucleic acid that is beard externally by the nanorobot (probe). Hybridization of probe and target produces a localized structural change that results in flap opening. The flap movement is studied on a two-dimensional prototype origami using Förster resonance energy transfer and is shown to be triggered by a variety of targets, including natural RNAs. The nanorobot has potential for in vivo biosensing and intelligent delivery of biological activators. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.
Optimization of Peptide Nucleic Acid Antisense Oligonucleotides for Local and Systemic Dystrophin Splice Correction in the mdx Mouse

Science.gov (United States)

Yin, HaiFang; Betts, Corinne; Saleh, Amer F; Ivanova, Gabriela D; Lee, Hyunil; Seow, Yiqi; Kim, Dalsoo; Gait, Michael J; Wood, Matthew JA

2010-01-01

Antisense oligonucleotides (AOs) have the capacity to alter the processing of pre-mRNA transcripts in order to correct the function of aberrant disease-related genes. Duchenne muscular dystrophy (DMD) is a fatal X-linked muscle degenerative disease that arises from mutations in the DMD gene leading to an absence of dystrophin protein. AOs have been shown to restore the expression of functional dystrophin via splice correction by intramuscular and systemic delivery in animal models of DMD and in DMD patients via intramuscular administration. Major challenges in developing this splice correction therapy are to optimize AO chemistry and to develop more effective systemic AO delivery. Peptide nucleic acid (PNA) AOs are an alternative AO chemistry with favorable in vivo biochemical properties and splice correcting abilities. Here, we show long-term splice correction of the DMD gene in mdx mice following intramuscular PNA delivery and effective splice correction in aged mdx mice. Further, we report detailed optimization of systemic PNA delivery dose regimens and PNA AO lengths to yield splice correction, with 25-mer PNA AOs providing the greatest splice correcting efficacy, restoring dystrophin protein in multiple peripheral muscle groups. PNA AOs therefore provide an attractive candidate AO chemistry for DMD exon skipping therapy. PMID:20068555
Developing nucleic acid-based electrical detection systems

Directory of Open Access Journals (Sweden)

Gabig-Ciminska Magdalena

2006-03-01

Full Text Available Abstract Development of nucleic acid-based detection systems is the main focus of many research groups and high technology companies. The enormous work done in this field is particularly due to the broad versatility and variety of these sensing devices. From optical to electrical systems, from label-dependent to label-free approaches, from single to multi-analyte and array formats, this wide range of possibilities makes the research field very diversified and competitive. New challenges and requirements for an ideal detector suitable for nucleic acid analysis include high sensitivity and high specificity protocol that can be completed in a relatively short time offering at the same time low detection limit. Moreover, systems that can be miniaturized and automated present a significant advantage over conventional technology, especially if detection is needed in the field. Electrical system technology for nucleic acid-based detection is an enabling mode for making miniaturized to micro- and nanometer scale bio-monitoring devices via the fusion of modern micro- and nanofabrication technology and molecular biotechnology. The electrical biosensors that rely on the conversion of the Watson-Crick base-pair recognition event into a useful electrical signal are advancing rapidly, and recently are receiving much attention as a valuable tool for microbial pathogen detection. Pathogens may pose a serious threat to humans, animal and plants, thus their detection and analysis is a significant element of public health. Although different conventional methods for detection of pathogenic microorganisms and their toxins exist and are currently being applied, improvements of molecular-based detection methodologies have changed these traditional detection techniques and introduced a new era of rapid, miniaturized and automated electrical chip detection technologies into pathogen identification sector. In this review some developments and current directions in
Boronic acid-based autoligation of nucleic acids

DEFF Research Database (Denmark)

Barbeyron, R.; Vasseur, J.-J.; Smietana, M.

2013-01-01

Abstract: The development of synthetic systems displaying dynamic and adaptive characteristics is a formidable challenge with wide applications from biotechnology to therapeutics. Recently, we described a dynamic and programmable nucleic acid-based system relying on the formation of reversible bo....... Evidence suggests that geometric and steric factors are key features for controlling the equilibria. Graphical Abstract: [Figure not available: see fulltext.]...
Scaffolding along Nucleic Acid Duplexes Using 2'-Amino-Locked Nucleic Acids

DEFF Research Database (Denmark)

Astakhova, I Kira; Wengel, Jesper

2014-01-01

-LNA nucleotides. By application of different chemical reactions, modification of 2'-amino-LNA scaffolds can be efficiently performed in high yields and with various tags, postsynthetically or during the automated oligonucleotide synthesis. The choice of a synthetic method for scaffolding along 2'-amino-LNA mainly....../DNA probes bind nucleic acid targets with advantages of high affinity and specificity. Thus, molecular motion of nanodevices and programmable self-assembly of chemically modified LNA/DNA nanomaterials can be followed by bright fluorescence signaling from the functionalized LNA units. Another appealing aspect...
Nanoparticulate systems for nucleic acid delivery

NARCIS (Netherlands)

Varkouhi, A.K.

2011-01-01

Development of carrier systems with controllable physicochemical and delivery properties has opened up the possibility of nanomedicines containing nucleic acids. In the last decades, much effort has been dedicated to two exciting approaches in biomedicine, namely gene and RNA interference
A locked nucleic Acid-based nanocrawler

DEFF Research Database (Denmark)

Astakhova, I Kira; Pasternak, Karol; Campbell, Meghan A

2013-01-01

Herein we introduce a novel fluorescent LNA/DNA machine, a nanocrawler, which reversibly moves along a directionally polar complementary road controlled by affinity-enhancing locked nucleic acid (LNA) monomers and additional regulatory strands. Polyaromatic hydrocarbon (PAH) dyes attached to 2...
Effect of time and deposition method on quality of phosphonic acid modifier self-assembled monolayers on indium zinc oxide

Energy Technology Data Exchange (ETDEWEB)

Sang, Lingzi [Department of Chemistry and Biochemistry, University of Arizona, Tucson, AZ 85721 (United States); Knesting, Kristina M. [Department of Chemistry, University of Washington, Seattle, WA 98195-1700 (United States); Bulusu, Anuradha [School of Mechanical Engineering, Georgia Institute of Technology, Atlanta, GA 30332 (United States); Sigdel, Ajaya K. [National Renewable Energy Laboratory, Golden, CO 80401 (United States); Giordano, Anthony J.; Marder, Seth R. [School of Chemistry and Biochemistry and Center for Organic Photonics and Electronics, Georgia Institute of Technology, Atlanta, GA 30332-0400 (United States); Berry, Joseph J. [National Renewable Energy Laboratory, Golden, CO 80401 (United States); Graham, Samuel [School of Mechanical Engineering, Georgia Institute of Technology, Atlanta, GA 30332 (United States); Ginger, David S. [Department of Chemistry, University of Washington, Seattle, WA 98195-1700 (United States); Pemberton, Jeanne E., E-mail: pembertn@email.arizona.edu [Department of Chemistry and Biochemistry, University of Arizona, Tucson, AZ 85721 (United States)

2016-12-15

Highlights: • Deposition of phosphonic acid monolayers on oxides from ethanol solutions occurs by rapid adsorption within 10 s with slower equilibration complete in 48 h. • The slower equilibration step involves molecular reorientation and vacancy filling on the oxide surface. • Soak-free deposition by spray coating and microcontact printing do not provide reproducible, fully-covered, uniform monolayers without substrate etching. • Adjustments to exposure time, substrate temperature, and solution/substrate contact efficiency are necessary to optimize soak-free methods. - Abstract: Phosphonic acid (PA) self-assembled monolayers (SAMs) are utilized at critical interfaces between transparent conductive oxides (TCO) and organic active layers in organic photovoltaic devices (OPVs). The effects of PA deposition method and time on the formation of close-packed, high-quality monolayers is investigated here for SAMs fabricated by solution deposition, micro-contact printing, and spray coating. The solution deposition isotherm for pentafluorinated benzylphosphonic acid (F{sub 5}BnPA) on indium-doped zinc oxide (IZO) is studied using polarization modulation-infrared reflection-absorption spectroscopy (PM-IRRAS) at room temperature as a model PA/IZO system. Fast surface adsorption occurs within the first min; however, well-oriented high-quality SAMs are reached only after ∼48 h, presumably through a continual process of molecular adsorption/desorption and monolayer filling accompanied by molecular reorientation. Two other rapid, soak-free deposition techniques, micro-contact printing and spray coating, are also explored. SAM quality is compared for deposition of phenyl phosphonic acid (PPA), F{sub 13}-octylphosphonic acid (F{sub 13}OPA), and pentafluorinated benzyl phosphonic acid (F{sub 5}BnPA) by solution deposition, micro-contact printing and spray coating using PM-IRRAS. In contrast to micro-contact printing and spray coating techniques, 48–168 h solution
Gene Therapy for Advanced Melanoma: Selective Targeting and Therapeutic Nucleic Acids

Directory of Open Access Journals (Sweden)

Joana R. Viola

2013-01-01

Full Text Available Despite recent advances, the treatment of malignant melanoma still results in the relapse of the disease, and second line treatment mostly fails due to the occurrence of resistance. A wide range of mutations are known to prevent effective treatment with chemotherapeutic drugs. Hence, approaches with biopharmaceuticals including proteins, like antibodies or cytokines, are applied. As an alternative, regimens with therapeutically active nucleic acids offer the possibility for highly selective cancer treatment whilst avoiding unwanted and toxic side effects. This paper gives a brief introduction into the mechanism of this devastating disease, discusses the shortcoming of current therapy approaches, and pinpoints anchor points which could be harnessed for therapeutic intervention with nucleic acids. We bring the delivery of nucleic acid nanopharmaceutics into perspective as a novel antimelanoma therapeutic approach and discuss the possibilities for melanoma specific targeting. The latest reports on preclinical and already clinical application of nucleic acids in melanoma are discussed.
Nucleic Acid Extraction from Synthetic Mars Analog Soils for in situ Life Detection

Science.gov (United States)

Mojarro, Angel; Ruvkun, Gary; Zuber, Maria T.; Carr, Christopher E.

2017-08-01

Biological informational polymers such as nucleic acids have the potential to provide unambiguous evidence of life beyond Earth. To this end, we are developing an automated in situ life-detection instrument that integrates nucleic acid extraction and nanopore sequencing: the Search for Extra-Terrestrial Genomes (SETG) instrument. Our goal is to isolate and determine the sequence of nucleic acids from extant or preserved life on Mars, if, for example, there is common ancestry to life on Mars and Earth. As is true of metagenomic analysis of terrestrial environmental samples, the SETG instrument must isolate nucleic acids from crude samples and then determine the DNA sequence of the unknown nucleic acids. Our initial DNA extraction experiments resulted in low to undetectable amounts of DNA due to soil chemistry-dependent soil-DNA interactions, namely adsorption to mineral surfaces, binding to divalent/trivalent cations, destruction by iron redox cycling, and acidic conditions. Subsequently, we developed soil-specific extraction protocols that increase DNA yields through a combination of desalting, utilization of competitive binders, and promotion of anaerobic conditions. Our results suggest that a combination of desalting and utilizing competitive binders may establish a "universal" nucleic acid extraction protocol suitable for analyzing samples from diverse soils on Mars.
Localization of Bovine Papillomavirus Nucleic Acid in Equine Sarcoids.

Science.gov (United States)

Gaynor, A M; Zhu, K W; Dela Cruz, F N; Affolter, V K; Pesavento, P A

2016-05-01

Bovine papillomaviruses (BPV1/BPV2) have long been associated with equine sarcoids; deciphering their contribution has been difficult due to their ubiquitous presence on skin and in the environment, as well as the lack of decent techniques to interrogate their role in pathogenesis. We have developed and characterized an in situ hybridization (ISH) assay that uses a pool of probes complementary to portions of the E5, E6, and E7 genes. This assay is highly sensitive for direct visualization of viral transcript and nucleic acid in routinely processed histopathologic samples. We demonstrate here the visualization of BPV nucleic acid in 18 of 18 equine sarcoids, whereas no detectable viral DNA was present in 15 of 15 nonsarcoid controls by this technique. In nearly 90% (16/18) of the sarcoids, 50% or more of the fibroblastic cell nuclei distributed throughout the neoplasm had detectable hybridization. In the remaining 2 cases, fewer than half of the fibroblastic cells contained detectable hybridization, but viral nucleic acid was also detected in epithelial cells of the sebaceous glands, hair follicles and epidermis. A sensitive ISH assay is an indispensable addition to the molecular methods used to detect viral nucleic acid in tissue. We have used this technique to determine the specific cellular localization and distribution of BPV in a subset of equine sarcoids. © The Author(s) 2015.
Assessment for Melting Temperature Measurement of Nucleic Acid by HRM.

Science.gov (United States)

Wang, Jing; Pan, Xiaoming; Liang, Xingguo

2016-01-01

High resolution melting (HRM), with a high sensitivity to distinguish the nucleic acid species with small variations, has been widely applied in the mutation scanning, methylation analysis, and genotyping. For the aim of extending HRM for the evaluation of thermal stability of nucleic acid secondary structures on sequence dependence, we investigated effects of the dye of EvaGreen, metal ions, and impurities (such as dNTPs) on melting temperature ( T m ) measurement by HRM. The accuracy of HRM was assessed as compared with UV melting method, and little difference between the two methods was found when the DNA T m was higher than 40°C. Both insufficiency and excessiveness of EvaGreen were found to give rise to a little bit higher T m , showing that the proportion of dye should be considered for precise T m measurement of nucleic acids. Finally, HRM method was also successfully used to measure T m s of DNA triplex, hairpin, and RNA duplex. In conclusion, HRM can be applied in the evaluation of thermal stability of nucleic acid (DNA or RNA) or secondary structural elements (even when dNTPs are present).
Delivery Systems for In Vivo use of Nucleic Acid Drugs

Directory of Open Access Journals (Sweden)

Resende R.R

2007-01-01

Full Text Available The notorious biotechnological advance of the last few decades has allowed the development of experimental methods for understanding molecular mechanisms of genes and new therapeutic approaches. Gene therapy is maturing into a viable, practical method with the potential to cure a variety of human illnesses. Some nucleic-acid-based drugs are now available for controlling the progression of genetic diseases by inhibiting gene expression or the activity of their gene products. New therapeutic strategies employ a wide range of molecular tools such as bacterial plasmids containing transgenic inserts, RNA interference aptamers. A nucleic-acid based constitution confers a lower immunogenic potential and as result of the high stringency selection of large molecular variety, these drugs have high affi nity and selectivity for their targets. However, nucleic acids have poor biostability thus requiring chemical modifications and delivery systems to maintain their activity and ease their cellular internalization. This review discusses some of the mechanisms of action and the application of therapies based on nucleic acids such as aptamers and RNA interference as well as platforms for cellular uptake and intracellular delivery of therapeutic oligonucleotides and their trade-offs.

Effect of Time and Deposition Method on Quality of Phosphonic Acid Modifier Self-Assembled Monolayers on Indium Zinc Oxide

Energy Technology Data Exchange (ETDEWEB)

Sang, Lingzi; Knesting, Kristina M.; Bulusu, Anuradha; Sigdel, Ajaya K.; Giordano, Anthony J.; Marder, Seth R.; Berry, Joseph J.; Graham, Samuel; Ginger, David S.; Pemberton, Jeanne E.

2016-12-15

Phosphonic acid (PA) self-assembled monolayers (SAMs) are utilized at critical interfaces between transparent conductive oxides (TCO) and organic active layers in organic photovoltaic devices (OPVs). The effects of PA deposition method and time on the formation of close-packed, high-quality monolayers is investigated here for SAMs fabricated by solution deposition, micro-contact printing, and spray coating. The solution deposition isotherm for pentafluorinated benzylphosphonic acid (F5BnPA) on indium-doped zinc oxide (IZO) is studied using polarization modulation-infrared reflection-absorption spectroscopy (PM-IRRAS) at room temperature as a model PA/IZO system. Fast surface adsorption occurs within the first min; however, well-oriented high-quality SAMs are reached only after -48 h, presumably through a continual process of molecular adsorption/desorption and monolayer filling accompanied by molecular reorientation. Two other rapid, soak-free deposition techniques, micro-contact printing and spray coating, are also explored. SAM quality is compared for deposition of phenyl phosphonic acid (PPA), F13-octylphosphonic acid (F13OPA), and pentafluorinated benzyl phosphonic acid (F5BnPA) by solution deposition, micro-contact printing and spray coating using PM-IRRAS. In contrast to micro-contact printing and spray coating techniques, 48-168 h solution deposition at both room temperature and 70 degrees C result in contamination- and surface etch-free close-packed monolayers with good reproducibility. SAMs fabricated by micro-contact printing and spray coating are much less well ordered.
A non-covalent peptide-based carrier for in vivo delivery of DNA mimics

OpenAIRE

Morris, May C.; Gros, Edwige; Aldrian-Herrada, Gudrun; Choob, Michael; Archdeacon, John; Heitz, Frederic; Divita, Gilles

2007-01-01

The dramatic acceleration in identification of new nucleic-acid-based therapeutic molecules has provided new perspectives in pharmaceutical research. However, their development is limited by their poor cellular uptake and inefficient trafficking. Here we describe a short amphipathic peptide, Pep-3, that combines a tryptophan/phenylalanine domain with a lysine/arginine-rich hydrophilic motif. Pep-3 forms stable nano-size complexes with peptide-nucleic acid analogues and promotes their efficien...
Nucleic acid compositions and the encoding proteins

Science.gov (United States)

Preston, III, James F.; Chow, Virginia; Nong, Guang; Rice, John D.; St. John, Franz J.

2014-09-02

The subject invention provides at least one nucleic acid sequence encoding an aldouronate-utilization regulon isolated from Paenibacillus sp. strain JDR-2, a bacterium which efficiently utilizes xylan and metabolizes aldouronates (methylglucuronoxylosaccharides). The subject invention also provides a means for providing a coordinately regulated process in which xylan depolymerization and product assimilation are coupled in Paenibacillus sp. strain JDR-2 to provide a favorable system for the conversion of lignocellulosic biomass to biobased products. Additionally, the nucleic acid sequences encoding the aldouronate-utilization regulon can be used to transform other bacteria to form organisms capable of producing a desired product (e.g., ethanol, 1-butanol, acetoin, 2,3-butanediol, 1,3-propanediol, succinate, lactate, acetate, malate or alanine) from lignocellulosic biomass.
Recent advances in delivery of drug-nucleic acid combinations for cancer treatment.

Science.gov (United States)

Li, Jing; Wang, Yan; Zhu, Yu; Oupický, David

2013-12-10

Cancer treatment that uses a combination of approaches with the ability to affect multiple disease pathways has been proven highly effective in the treatment of many cancers. Combination therapy can include multiple chemotherapeutics or combinations of chemotherapeutics with other treatment modalities like surgery or radiation. However, despite the widespread clinical use of combination therapies, relatively little attention has been given to the potential of modern nanocarrier delivery methods, like liposomes, micelles, and nanoparticles, to enhance the efficacy of combination treatments. This lack of knowledge is particularly notable in the limited success of vectors for the delivery of combinations of nucleic acids with traditional small molecule drugs. The delivery of drug-nucleic acid combinations is particularly challenging due to differences in the physicochemical properties of the two types of agents. This review discusses recent advances in the development of delivery methods using combinations of small molecule drugs and nucleic acid therapeutics to treat cancer. This review primarily focuses on the rationale used for selecting appropriate drug-nucleic acid combinations as well as progress in the development of nanocarriers suitable for simultaneous delivery of drug-nucleic acid combinations. Copyright © 2013 Elsevier B.V. All rights reserved.
Identification of nucleic acid binding sites on translin-associated factor X (TRAX protein.

Directory of Open Access Journals (Sweden)

Gagan Deep Gupta

Full Text Available Translin and TRAX proteins play roles in very important cellular processes such as DNA recombination, spatial and temporal expression of mRNA, and in siRNA processing. Translin forms a homomeric nucleic acid binding complex and binds to ssDNA and RNA. However, a mutant translin construct that forms homomeric complex lacking nucleic acid binding activity is able to form fully active heteromeric translin-TRAX complex when co-expressed with TRAX. A substantial progress has been made in identifying translin sites that mediate its binding activity, while TRAX was thought not to bind DNA or RNA on its own. We here for the first time demonstrate nucleic acid binding to TRAX by crosslinking radiolabeled ssDNA to heteromeric translin-TRAX complex using UV-laser. The TRAX and translin, photochemically crosslinked with ssDNA, were individually detected on SDS-PAGE. We mutated two motifs in TRAX and translin, designated B2 and B3, to help define the nucleic acid binding sites in the TRAX sequence. The most pronounced effect was observed in the mutants of B3 motif that impaired nucleic acid binding activity of the heteromeric complexes. We suggest that both translin and TRAX are binding competent and contribute to the nucleic acid binding activity.
Identification of Nucleic Acid Binding Sites on Translin-Associated Factor X (TRAX) Protein

Science.gov (United States)

Gupta, Gagan Deep; Kumar, Vinay

2012-01-01

Translin and TRAX proteins play roles in very important cellular processes such as DNA recombination, spatial and temporal expression of mRNA, and in siRNA processing. Translin forms a homomeric nucleic acid binding complex and binds to ssDNA and RNA. However, a mutant translin construct that forms homomeric complex lacking nucleic acid binding activity is able to form fully active heteromeric translin-TRAX complex when co-expressed with TRAX. A substantial progress has been made in identifying translin sites that mediate its binding activity, while TRAX was thought not to bind DNA or RNA on its own. We here for the first time demonstrate nucleic acid binding to TRAX by crosslinking radiolabeled ssDNA to heteromeric translin-TRAX complex using UV-laser. The TRAX and translin, photochemically crosslinked with ssDNA, were individually detected on SDS-PAGE. We mutated two motifs in TRAX and translin, designated B2 and B3, to help define the nucleic acid binding sites in the TRAX sequence. The most pronounced effect was observed in the mutants of B3 motif that impaired nucleic acid binding activity of the heteromeric complexes. We suggest that both translin and TRAX are binding competent and contribute to the nucleic acid binding activity. PMID:22427937
Novel quinolinone-phosphonic acid AMPA antagonists devoid of nephrotoxicity.

Science.gov (United States)

Cordi, Alex A; Desos, Patrice; Ruano, Elisabeth; Al-Badri, Hashim; Fugier, Claude; Chapman, Astrid G; Meldrum, Brian S; Thomas, Jean-Yves; Roger, Anita; Lestage, Pierre

2002-10-01

We reported previously the synthesis and structure-activity relationships (SAR) in a series of 2-(1H)-oxoquinolines bearing different acidic functions in the 3-position. Exploiting these SAR, we were able to identify 6,7-dichloro-2-(1H)-oxoquinoline-3-phosphonic acid compound 3 (S 17625) as a potent, in vivo active AMPA antagonist. Unfortunately, during the course of the development, nephrotoxicity was manifest at therapeutically effective doses. Considering that some similitude exists between S 17625 and probenecid, a compound known to protect against the nephrotoxicity and/or slow the clearance of different drugs, we decided to synthesise some new analogues of S 17625 incorporating some of the salient features of probenecid. Replacement of the chlorine in position 6 by a sulfonylamine led to very potent AMPA antagonists endowed with good in vivo activity and lacking nephrotoxicity potential. Amongst the compounds evaluated, derivatives 7a and 7s appear to be the most promising and are currently evaluated in therapeutically relevant stroke models.
Novel fluorescent nanoparticles for ultrasensitive identification of nucleic acids by optical methods

DEFF Research Database (Denmark)

Mulberg, Mads Westergaard; Taskova, Maria; Thomsen, Rasmus P.

2017-01-01

For decades, the detection of nucleic acids and their interactions at low abundances has been a challenging task. Present nucleic acid diagnostics are primarily based on enzymatic reactions including sequencing, polymerase-chain reaction and microarrays. However, the use of enzymatic amplificatio...
Toward Peptide Nucleic Acid (PNA) Directed Peptide Translation Using Ester Based Aminoacyl Transfer

DEFF Research Database (Denmark)

Singhal, Abhishek; Bagnacani, Valentina; Corradini, Roberto

2014-01-01

Peptide synthesis is a fundamental feature of life. However, it still remains unclear how the contemporary translation apparatus evolved from primitive prebiotic systems and at which stage of the evolution peptide synthesis emerged. Using simple molecular architectures, in which aminoacyl transfe...
Peptide Nucleic Acid Knockdown and Intra-host Cell Complementation of Ehrlichia Type IV Secretion System Effector

Directory of Open Access Journals (Sweden)

Pratibha Sharma

2017-06-01

Full Text Available Survival of Ehrlichia chaffeensis depends on obligatory intracellular infection. One of the barriers to E. chaffeensis research progress has been the inability, using conventional techniques, to generate knock-out mutants for genes essential for intracellular infection. This study examined the use of Peptide Nucleic Acids (PNAs technology to interrupt type IV secretion system (T4SS effector protein expression in E. chaffeensis followed by intracellular complementation of the effector to determine its requirement for infection. Successful E. chaffeensis infection depends on the E. chaffeensis-specific T4SS protein effector, ehrlichial translocated factor-1 (Etf-1, which induces Rab5-regulated autophagy to provide host cytosolic nutrients required for E. chaffeensis proliferation. Etf-1 is also imported by host cell mitochondria where it inhibits host cell apoptosis to prolong its infection. We designed a PNA specific to Etf-1 and showed that the PNA bound to the target region of single-stranded Etf-1 RNA using a competitive binding assay. Electroporation of E. chaffeensis with this PNA significantly reduced Etf-1 mRNA and protein, and the bacteria's ability to induce host cell autophagy and infect host cells. Etf-1 PNA-mediated inhibition of ehrlichial Etf-1 expression and E. chaffeensis infection could be intracellularly trans-complemented by ectopic expression of Etf-1-GFP in host cells. These data affirmed the critical role of bacterial T4SS effector in host cell autophagy and E. chaffeensis infection, and demonstrated the use of PNA to analyze the gene functions of obligate intracellular bacteria.
Nucleic acid protocols: Extraction and optimization

Directory of Open Access Journals (Sweden)

Saeed El-Ashram

2016-12-01

Full Text Available Yield and quality are fundamental features for any researchers during nucleic acid extraction. Here, we describe a simplified, semi-unified, effective, and toxic material free protocol for extracting DNA and RNA from different prokaryotic and eukaryotic sources exploiting the physical and chemical properties of nucleic acids. Furthermore, this protocol showed that DNA and RNA are under triple protection (i.e. EDTA, SDS and NaCl during lysis step, and this environment is improper for RNase to have DNA liberated of RNA and even for DNase to degrade the DNA. Therefore, the complete removal of RNA under RNase influence is achieved when RNase is added after DNA extraction, which gives optimal quality with any protocols. Similarly, DNA contamination in an isolated RNA is degraded by DNase to obtain high-quality RNA. Our protocol is the protocol of choice in terms of simplicity, recovery time, environmental safety, amount, purity, PCR and RT-PCR applicability.
Structure, stability and behaviour of nucleic acids in ionic liquids

Science.gov (United States)

Tateishi-Karimata, Hisae; Sugimoto, Naoki

2014-01-01

Nucleic acids have become a powerful tool in nanotechnology because of their conformational polymorphism. However, lack of a medium in which nucleic acid structures exhibit long-term stability has been a bottleneck. Ionic liquids (ILs) are potential solvents in the nanotechnology field. Hydrated ILs, such as choline dihydrogen phosphate (choline dhp) and deep eutectic solvent (DES) prepared from choline chloride and urea, are ‘green’ solvents that ensure long-term stability of biomolecules. An understanding of the behaviour of nucleic acids in hydrated ILs is necessary for developing DNA materials. We here review current knowledge about the structures and stabilities of nucleic acids in choline dhp and DES. Interestingly, in choline dhp, A–T base pairs are more stable than G–C base pairs, the reverse of the situation in buffered NaCl solution. Moreover, DNA triplex formation is markedly stabilized in hydrated ILs compared with aqueous solution. In choline dhp, the stability of Hoogsteen base pairs is comparable to that of Watson–Crick base pairs. Moreover, the parallel form of the G-quadruplex is stabilized in DES compared with aqueous solution. The behaviours of various DNA molecules in ILs detailed here should be useful for designing oligonucleotides for the development of nanomaterials and nanodevices. PMID:25013178
Highly simplified lateral flow-based nucleic acid sample preparation and passive fluid flow control

Science.gov (United States)

Cary, Robert E.

2015-12-08

Highly simplified lateral flow chromatographic nucleic acid sample preparation methods, devices, and integrated systems are provided for the efficient concentration of trace samples and the removal of nucleic acid amplification inhibitors. Methods for capturing and reducing inhibitors of nucleic acid amplification reactions, such as humic acid, using polyvinylpyrrolidone treated elements of the lateral flow device are also provided. Further provided are passive fluid control methods and systems for use in lateral flow assays.
Highly simplified lateral flow-based nucleic acid sample preparation and passive fluid flow control

Energy Technology Data Exchange (ETDEWEB)

Cary, Robert B.

2018-04-17

Highly simplified lateral flow chromatographic nucleic acid sample preparation methods, devices, and integrated systems are provided for the efficient concentration of trace samples and the removal of nucleic acid amplification inhibitors. Methods for capturing and reducing inhibitors of nucleic acid amplification reactions, such as humic acid, using polyvinylpyrrolidone treated elements of the lateral flow device are also provided. Further provided are passive fluid control methods and systems for use in lateral flow assays.
Nucleic Acid Templated Reactions for Chemical Biology.

Science.gov (United States)

Di Pisa, Margherita; Seitz, Oliver

2017-06-21

Nucleic acid directed bioorthogonal reactions offer the fascinating opportunity to unveil and redirect a plethora of intracellular mechanisms. Nano- to picomolar amounts of specific RNA molecules serve as templates and catalyze the selective formation of molecules that 1) exert biological effects, or 2) provide measurable signals for RNA detection. Turnover of reactants on the template is a valuable asset when concentrations of RNA templates are low. The idea is to use RNA-templated reactions to fully control the biodistribution of drugs and to push the detection limits of DNA or RNA analytes to extraordinary sensitivities. Herein we review recent and instructive examples of conditional synthesis or release of compounds for in cellulo protein interference and intracellular nucleic acid imaging. © 2017 The Authors. Published by Wiley-VCH Verlag GmbH & Co. KGaA.
Design of Tail-Clamp Peptide Nucleic Acid Tethered with Azobenzene Linker for Sequence-Specific Detection of Homopurine DNA

Directory of Open Access Journals (Sweden)

Shinjiro Sawada

2017-10-01

Full Text Available DNA carries genetic information in its sequence of bases. Synthetic oligonucleotides that can sequence-specifically recognize a target gene sequence are a useful tool for regulating gene expression or detecting target genes. Among the many synthetic oligonucleotides, tail-clamp peptide nucleic acid (TC-PNA offers advantages since it has two homopyrimidine PNA strands connected via a flexible ethylene glycol-type linker that can recognize complementary homopurine sequences via Watson-Crick and Hoogsteen base pairings and form thermally-stable PNA/PNA/DNA triplex structures. Here, we synthesized a series of TC-PNAs that can possess different lengths of azobenzene-containing linkers and studied their binding behaviours to homopurine single-stranded DNA. Introduction of azobenzene at the N-terminus amine of PNA increased the thermal stability of PNA-DNA duplexes. Further extension of the homopyrimidine PNA strand at the N-terminus of PNA-AZO further increased the binding stability of the PNA/DNA/PNA triplex to the target homopurine sequence; however, it induced TC-PNA/DNA/TC-PNA complex formation. Among these TC-PNAs, 9W5H-C4-AZO consisting of nine Watson-Crick bases and five Hoogsteen bases tethered with a beta-alanine conjugated azobenzene linker gave a stable 1:1 TC-PNA/ssDNA complex and exhibited good mismatch recognition. Our design for TC-PNA-AZO can be utilized for detecting homopurine sequences in various genes.
The relationship between odd- and branched-chain fatty acids and microbial nucleic acid bases in rumen

Directory of Open Access Journals (Sweden)

Keyuan Liu

2017-11-01

Full Text Available Objective This study aims to identify the relationship between odd- and branched-chain fatty acids (OBCFAs and microbial nucleic acid bases in the rumen, and to establish a model to accurately predict microbial protein flow by using OBCFA. Methods To develop the regression equations, data on the rumen contents of individual cows were obtained from 2 feeding experiments. In the first experiment, 3 rumen-fistulated dry dairy cows arranged in a 3×3 Latin square were fed diets of differing forage to concentration ratios (F:C. The second experiment consisted of 9 lactating Holstein dairy cows of similar body weights at the same stage of pregnancy. For each lactation stage, 3 cows with similar milk production were selected. The rumen contents were sampled at 4 time points of every two hours after morning feeding 6 h, and then to analyse the concentrations of OBCFA and microbial nucleic acid bases in the rumen samples. Results The ruminal bacteria nucleic acid bases were significantly influenced by feeding diets of differing forge to concentration ratios and lactation stages of dairy cows (p<0.05. The concentrations of OBCFAs, especially odd-chain fatty acids and C15:0 isomers, strongly correlated with the microbial nucleic acid bases in the rumen (p<0.05. The equations of ruminal microbial nucleic acid bases established by ruminal OBCFAs contents showed a good predictive capacity, as indicated by reasonably low standard errors and high R-squared values. Conclusion This finding suggests that the rumen OBCFA composition could be used as an internal marker of rumen microbial matter.
Logic gates and antisense DNA devices operating on a translator nucleic Acid scaffold.

Science.gov (United States)

Shlyahovsky, Bella; Li, Yang; Lioubashevski, Oleg; Elbaz, Johann; Willner, Itamar

2009-07-28

A series of logic gates, "AND", "OR", and "XOR", are designed using a DNA scaffold that includes four "footholds" on which the logic operations are activated. Two of the footholds represent input-recognition strands, and these are blocked by complementary nucleic acids, whereas the other two footholds are blocked by nucleic acids that include the horseradish peroxidase (HRP)-mimicking DNAzyme sequence. The logic gates are activated by either nucleic acid inputs that hybridize to the respective "footholds", or by low-molecular-weight inputs (adenosine monophosphate or cocaine) that yield the respective aptamer-substrate complexes. This results in the respective translocation of the blocking nucleic acids to the footholds carrying the HRP-mimicking DNAzyme sequence, and the concomitant release of the respective DNAzyme. The released product-strands then self-assemble into the hemin/G-quadruplex-HRP-mimicking DNAzyme that biocatalyzes the formation of a colored product and provides an output signal for the different logic gates. The principle of the logic operation is, then, implemented as a possible paradigm for future nanomedicine. The nucleic acid inputs that bind to the blocked footholds result in the translocation of the blocking nucleic acids to the respective footholds carrying the antithrombin aptamer. The released aptamer inhibits, then, the hydrolytic activity of thrombin. The system demonstrates the regulation of a biocatalytic reaction by a translator system activated on a DNA scaffold.
Application of locked nucleic acid-based probes in fluorescence in situ hybridization

DEFF Research Database (Denmark)

Fontenete, Sílvia; Carvalho, Daniel R; Guimarães, Nuno

2016-01-01

of nucleic acid mimics used as mixmers in LNA-based probes strongly influence the efficiency of detection. LNA probes with 10 to 15 mers showed the highest efficiency. Additionally, the combination of 2′-OMe RNA with LNA allowed an increase on the fluorescence intensities of the probes. Overall......Fluorescence in situ hybridization (FISH) employing nucleic acid mimics as probes is becoming an emerging molecular tool in the microbiology area for the detection and visualization of microorganisms. However, the impact that locked nucleic acid (LNA) and 2′-O-methyl (2′-OMe) RNA modifications have...
Optimizing scoring function of protein-nucleic acid interactions with both affinity and specificity.

Directory of Open Access Journals (Sweden)

Zhiqiang Yan

Full Text Available Protein-nucleic acid (protein-DNA and protein-RNA recognition is fundamental to the regulation of gene expression. Determination of the structures of the protein-nucleic acid recognition and insight into their interactions at molecular level are vital to understanding the regulation function. Recently, quantitative computational approach has been becoming an alternative of experimental technique for predicting the structures and interactions of biomolecular recognition. However, the progress of protein-nucleic acid structure prediction, especially protein-RNA, is far behind that of the protein-ligand and protein-protein structure predictions due to the lack of reliable and accurate scoring function for quantifying the protein-nucleic acid interactions. In this work, we developed an accurate scoring function (named as SPA-PN, SPecificity and Affinity of the Protein-Nucleic acid interactions for protein-nucleic acid interactions by incorporating both the specificity and affinity into the optimization strategy. Specificity and affinity are two requirements of highly efficient and specific biomolecular recognition. Previous quantitative descriptions of the biomolecular interactions considered the affinity, but often ignored the specificity owing to the challenge of specificity quantification. We applied our concept of intrinsic specificity to connect the conventional specificity, which circumvents the challenge of specificity quantification. In addition to the affinity optimization, we incorporated the quantified intrinsic specificity into the optimization strategy of SPA-PN. The testing results and comparisons with other scoring functions validated that SPA-PN performs well on both the prediction of binding affinity and identification of native conformation. In terms of its performance, SPA-PN can be widely used to predict the protein-nucleic acid structures and quantify their interactions.

Cleavage and protection of locked nucleic acid-modified DNA by restriction endonucleases

DEFF Research Database (Denmark)

Crouzier, Lucile; Dubois, Camille; Wengel, Jesper

2012-01-01

Locked nucleic acid (LNA) is one of the most prominent nucleic acid analogues reported so far. We herein for the first time report cleavage by restriction endonuclease of LNA-modified DNA oligonucleotides. The experiments revealed that RsaI is an efficient enzyme capable of recognizing and cleaving...
Bis-Pyrene-Modified Unlocked Nucleic Acids: Synthesis, Hybridization Studies, and Fluorescent Properties

Czech Academy of Sciences Publication Activity Database

Perlíková, Pavla; Ejlersen, M.; Langkjaer, N.; Wengel, J.

2014-01-01

Roč. 9, č. 9 (2014), s. 2120-2127 ISSN 1860-7179 Grant - others:European Research Council(XE) FP7-268776 Institutional support: RVO:61388963 Keywords : fluorescence * nucleic acid hybridization * oligonucleotides * pyrenes * unlocked nucleic acids Subject RIV: CC - Organic Chemistry Impact factor: 2.968, year: 2014
Recent progress in nucleic acids isotachophoresis

Czech Academy of Sciences Publication Activity Database

Datinská, Vladimíra; Voráčová, Ivona; Schlecht, U.; Berka, J.; Foret, František

2018-01-01

Roč. 41, č. 1 (2018), s. 236-247 ISSN 1615-9306 R&D Projects: GA ČR(CZ) GBP206/12/G014 Institutional support: RVO:68081715 Keywords : isotachophoresis * nucleic acids * sample preparation Subject RIV: CB - Analytical Chemistry, Separation OBOR OECD: Analytical chemistry Impact factor: 2.557, year: 2016
Recent progress in nucleic acids isotachophoresis

Czech Academy of Sciences Publication Activity Database

Datinská, Vladimíra; Voráčová, Ivona; Schlecht, U.; Berka, J.; Foret, František

2018-01-01

Roč. 41, č. 1 (2018), s. 236-247 ISSN 1615-9306 R&D Projects: GA ČR(CZ) GBP206/12/G014 Institutional support: RVO:68081715 Keywords : isotachophoresis * nucleic acid s * sample preparation Subject RIV: CB - Analytical Chemistry, Separation OBOR OECD: Analytical chemistry Impact factor: 2.557, year: 2016
Non-viral Nucleic Acid Delivery Strategies to the Central Nervous System

Directory of Open Access Journals (Sweden)

James-Kevin Tan

2016-11-01

Full Text Available With an increased prevalence and understanding of central nervous system injuries and neurological disorders, nucleic acid therapies are gaining promise as a way to regenerate lost neurons or halt disease progression. While more viral vectors have been used clinically as tools for gene delivery, non-viral vectors are gaining interest due to lower safety concerns and the ability to deliver all types of nucleic acids. Nevertheless, there are still a number of barriers to nucleic acid delivery. In this focused review, we explore the in vivo challenges hindering non-viral nucleic acid delivery to the central nervous system and the strategies and vehicles used to overcome them. Advantages and disadvantages of different routes of administration including: systemic injection, cerebrospinal fluid injection, intraparenchymal injection, and peripheral administration are discussed. Non-viral vehicles and treatment strategies that have overcome delivery barriers and demonstrated in vivo gene transfer to the central nervous system are presented. These approaches can be used as guidelines in developing synthetic gene delivery vectors for central nervous system applications and will ultimately bring non-viral vectors closer to clinical application.
Single-Stranded Nucleic Acids Bind to the Tetramer Interface of SAMHD1 and Prevent Formation of the Catalytic Homotetramer.

Science.gov (United States)

Seamon, Kyle J; Bumpus, Namandjé N; Stivers, James T

2016-11-08

Sterile alpha motif and HD domain protein 1 (SAMHD1) is a unique enzyme that plays important roles in nucleic acid metabolism, viral restriction, and the pathogenesis of autoimmune diseases and cancer. Although much attention has been focused on its dNTP triphosphohydrolase activity in viral restriction and disease, SAMHD1 also binds to single-stranded RNA and DNA. Here we utilize a UV cross-linking method using 5-bromodeoxyuridine-substituted oligonucleotides coupled with high-resolution mass spectrometry to identify the binding site for single-stranded nucleic acids (ssNAs) on SAMHD1. Mapping cross-linked amino acids on the surface of existing crystal structures demonstrated that the ssNA binding site lies largely along the dimer-dimer interface, sterically blocking the formation of the homotetramer required for dNTPase activity. Surprisingly, the disordered C-terminus of SAMHD1 (residues 583-626) was also implicated in ssNA binding. An interaction between this region and ssNA was confirmed in binding studies using the purified SAMHD1 583-626 peptide. Despite a recent report that SAMHD1 possesses polyribonucleotide phosphorylase activity, we did not detect any such activity in the presence of inorganic phosphate, indicating that nucleic acid binding is unrelated to this proposed activity. These data suggest an antagonistic regulatory mechanism in which the mutually exclusive oligomeric state requirements for ssNA binding and dNTP hydrolase activity modulate these two functions of SAMHD1 within the cell.
Comparative evaluation of three commercial systems for nucleic acid extraction from urine specimens.

Science.gov (United States)

Tang, Yi-Wei; Sefers, Susan E; Li, Haijing; Kohn, Debra J; Procop, Gary W

2005-09-01

A nucleic acid extraction system that can handle small numbers of specimens with a short test turnaround time and short hands-on time is desirable for emergent testing. We performed a comparative validation on three systems: the MagNA Pure compact system (Compact), the NucliSens miniMAG extraction instrument (miniMAG), and the BioRobot EZ1 system (EZ1). A total of 75 urine specimens submitted for polyomavirus BK virus detection were used. The human beta-actin gene was detected on 75 (100%), 75 (100%), and 72 (96%) nucleic acid extracts prepared by the miniMAG, EZ1, and Compact, respectively. The miniMAG produced the highest quantity of nucleic acids and the best precision among the three systems. The agreement rate was 100% for BKV detection on nucleic acid extracts prepared by the three extraction systems. When a full panel of specimens was run, the hands-on time and test turnaround time were 105.7 and 121.1 min for miniMAG, 6.1 and 22.6 min for EZ1, and 7.4 and 33.7 min for Compact, respectively. The EZ1 and Compact systems processed automatic nucleic acid extraction properly, providing a good solution to the need for sporadic but emergent specimen detection. The miniMAG yielded the highest quantity of nucleic acids, suggesting that this system would be the best for specimens containing a low number of microorganisms of interest.
Probing the mechanical properties, conformational changes, and interactions of nucleic acids with magnetic tweezers.

Science.gov (United States)

Kriegel, Franziska; Ermann, Niklas; Lipfert, Jan

2017-01-01

Nucleic acids are central to the storage and transmission of genetic information. Mechanical properties, along with their sequence, both enable and fundamentally constrain the biological functions of DNA and RNA. For small deformations from the equilibrium conformations, nucleic acids are well described by an isotropic elastic rod model. However, external forces and torsional strains can induce conformational changes, giving rise to a complex force-torque phase diagram. This review focuses on magnetic tweezers as a powerful tool to precisely determine both the elastic parameters and conformational transitions of nucleic acids under external forces and torques at the single-molecule level. We review several variations of magnetic tweezers, in particular conventional magnetic tweezers, freely orbiting magnetic tweezers and magnetic torque tweezers, and discuss their characteristic capabilities. We then describe the elastic rod model for DNA and RNA and discuss conformational changes induced by mechanical stress. The focus lies on the responses to torque and twist, which are crucial in the mechanics and interactions of nucleic acids and can directly be measured using magnetic tweezers. We conclude by highlighting several recent studies of nucleic acid-protein and nucleic acid-small-molecule interactions as further applications of magnetic tweezers and give an outlook of some exciting developments to come. Copyright © 2016. Published by Elsevier Inc.
Characterization of peptide-oligonucleotide heteroconjugates by mass spectrometry.

OpenAIRE

Jensen, O N; Kulkarni, S; Aldrich, J V; Barofsky, D F

1996-01-01

Two peptide-oligothymidylic acids, prepared by joining an 11 residue synthetic peptide containing one internal carboxyl group (Asp side chain) to amino-linker-5'pdT6 and amino-linker-5'pdT10 oligonucleotides, were analyzed by matrix-assisted laser desorption/ionization (MALDI) on a linear time-of-flight mass spectrometer and by electrospray ionization (ESI) on a triple-quadrupole system. These synthetic compounds model peptide-nucleic acid heteroconjugates encountered in antisense research an...
Nucleic acid polymeric properties and electrostatics: Directly comparing theory and simulation with experiment.

Science.gov (United States)

Sim, Adelene Y L

2016-06-01

Nucleic acids are biopolymers that carry genetic information and are also involved in various gene regulation functions such as gene silencing and protein translation. Because of their negatively charged backbones, nucleic acids are polyelectrolytes. To adequately understand nucleic acid folding and function, we need to properly describe its i) polymer/polyelectrolyte properties and ii) associating ion atmosphere. While various theories and simulation models have been developed to describe nucleic acids and the ions around them, many of these theories/simulations have not been well evaluated due to complexities in comparison with experiment. In this review, I discuss some recent experiments that have been strategically designed for straightforward comparison with theories and simulation models. Such data serve as excellent benchmarks to identify limitations in prevailing theories and simulation parameters. Copyright © 2015 Elsevier B.V. All rights reserved.
Comparison of Zirconium Phosphonate-Modified Surfaces for Immobilizing Phosphopeptides and Phosphate-Tagged Proteins.

Science.gov (United States)

Forato, Florian; Liu, Hao; Benoit, Roland; Fayon, Franck; Charlier, Cathy; Fateh, Amina; Defontaine, Alain; Tellier, Charles; Talham, Daniel R; Queffélec, Clémence; Bujoli, Bruno

2016-06-07

Different routes for preparing zirconium phosphonate-modified surfaces for immobilizing biomolecular probes are compared. Two chemical-modification approaches were explored to form self-assembled monolayers on commercially available primary amine-functionalized slides, and the resulting surfaces were compared to well-characterized zirconium phosphonate monolayer-modified supports prepared using Langmuir-Blodgett methods. When using POCl3 as the amine phosphorylating agent followed by treatment with zirconyl chloride, the result was not a zirconium-phosphonate monolayer, as commonly assumed in the literature, but rather the process gives adsorbed zirconium oxide/hydroxide species and to a lower extent adsorbed zirconium phosphate and/or phosphonate. Reactions giving rise to these products were modeled in homogeneous-phase studies. Nevertheless, each of the three modified surfaces effectively immobilized phosphopeptides and phosphopeptide tags fused to an affinity protein. Unexpectedly, the zirconium oxide/hydroxide modified surface, formed by treating the amine-coated slides with POCl3/Zr(4+), afforded better immobilization of the peptides and proteins and efficient capture of their targets.
Development of a peptide nucleic acid polymerase chain reaction clamping assay for semiquantitative evaluation of genetically modified organism content in food.

Science.gov (United States)

Peano, C; Lesignoli, F; Gulli, M; Corradini, R; Samson, M C; Marchelli, R; Marmiroli, N

2005-09-15

In the present study a peptide nucleic acid (PNA)-mediated polymerase chain reaction (PCR) clamping method was developed and applied to the detection of genetically modified organisms (GMO), to test PCR products for band identity and to obtain a semiquantitative evaluation of GMO content. The minimal concentration of PNA necessary to block the PCR was determined by comparing PCRs containing a constant amount of DNA in the presence of increasing concentration of target-specific PNA. The lowest PNA concentration at which specific inhibition took place, by the inhibition of primer extension and/or steric hindrance, was the most efficient condition. Optimization of PCR clamping by PNA was observed by testing five different PNAs with a minimum of 13 bp to a maximum of 15 bp, designed on the target sequence of Roundup Ready soybean. The results obtained on the DNA extracted from Roundup Ready soybean standard flour were verified also on DNA extracted from standard flours of maize GA21, Bt176, Bt11, and MON810. A correlation between the PNA concentration necessary for inducing PCR clamping and the percentage of the GMO target sequence in the sample was found.
Covering all the bases : Coarse-grained model design and application for nucleic acids

NARCIS (Netherlands)

Uusitalo, Jaakko Juhani

2016-01-01

Nucleic acids play a crucial role in the storage, transportation and expression of our genetic information. They have also become an interesting tool for many applications in nanotechnology. Studying biomolecular systems containing nucleic acids using experimental and imaging techniques has its
Locked nucleic acid (LNA): High affinity targeting of RNA for diagnostics and therapeutics

DEFF Research Database (Denmark)

Kauppinen, S.; Vester, Birte; Wengel, Jesper

2005-01-01

Locked nucleic acid (LNA) is a nucleic acid analogue containing one or more LNA nucleotide monomers with a bicyclic furanose unit locked in an RNA mimicking sugar conformation. This conformational restriction results in unprecedented hybridization affinity towards complementary single stranded RN...
[Comparison of two nucleic acid extraction methods for norovirus in oysters].

Science.gov (United States)

Yuan, Qiao; Li, Hui; Deng, Xiaoling; Mo, Yanling; Fang, Ling; Ke, Changwen

2013-04-01

To explore a convenient and effective method for norovirus nucleic acid extraction from oysters suitable for long-term viral surveillance. Two methods, namely method A (glycine washing and polyethylene glycol precipitation of the virus followed by silica gel centrifugal column) and method B (protease K digestion followed by application of paramagnetic silicon) were compared for their performance in norovirus nucleic acid extraction from oysters. Real-time RT-PCR was used to detect norovirus in naturally infected oysters and in oysters with induced infection. The two methods yielded comparable positive detection rates for the samples, but the recovery rate of the virus was higher with method B than with method A. Method B is a more convenient and rapid method for norovirus nucleic acid extraction from oysters and suitable for long-term surveillance of norovirus.
Conformational analysis of Infectious bursal disease virus (IBDV derived cell penetrating peptide (CPP analogs

Directory of Open Access Journals (Sweden)

Vinay G. Joshi

2013-12-01

Full Text Available Aim: This study was designed to develop peptide analogs of Infectious Bursal Disease (IBD virus VP5 protein segment having cell penetrating ability to improve their interaction with cargo molecule (Nucleic acid without affecting the backbone conformation. Materials and Methods: IBDV VP5 protein segment designated as RATH peptide were synthesized using solid phase peptide synthesis and their solution conformation was elucidated using CD spectroscopy in polar (water and apolar (TFE solvents. Cell penetrating ability of RATH-CONH2 was observed using FITC labeled peptide internalization in to HeLa cells under fluorescent microscopy. The efficacy of RATH analog interactions with nucleic acids was evaluated using FITC labeled oligonucleotides by fluorescence spectroscopy and plasmid constructs in gel retardation assay. Results: CD spectra of RATH analogs in water and apolar trifluroethanol (TFE helped to compare their secondary structures which were almost similar with dominant beta conformations suggesting successful induction of positive charge in the analogs without affecting back bone conformation of CPP designed. Cell penetrating ability of RATH CONH2 in HeLa cell was more than 90%. The fluorescence spectroscopy and plasmid constructs in gel retardation assay demonstrated successful interaction of amide analogs with nucleic acid. Conclusion: Intentional changes made in IBDV derived peptide RATH COOH to RATH CONH2 did not showed major changes in backbone conformation and such modifications may help to improve the cationic charge in most CPPs to interact with nucleic acid. [Vet World 2013; 6(6.000: 307-312
Rapid hybridization of nucleic acids using isotachophoresis

Science.gov (United States)

Bercovici, Moran; Han, Crystal M.; Liao, Joseph C.; Santiago, Juan G.

2012-01-01

We use isotachophoresis (ITP) to control and increase the rate of nucleic acid hybridization reactions in free solution. We present a new physical model, validation experiments, and demonstrations of this assay. We studied the coupled physicochemical processes of preconcentration, mixing, and chemical reaction kinetics under ITP. Our experimentally validated model enables a closed form solution for ITP-aided reaction kinetics, and reveals a new characteristic time scale which correctly predicts order 10,000-fold speed-up of chemical reaction rate for order 100 pM reactants, and greater enhancement at lower concentrations. At 500 pM concentration, we measured a reaction time which is 14,000-fold lower than that predicted for standard second-order hybridization. The model and method are generally applicable to acceleration of reactions involving nucleic acids, and may be applicable to a wide range of reactions involving ionic reactants. PMID:22733732
The association between low-grade inflammation, iron status and nucleic acid oxidation in the elderly

DEFF Research Database (Denmark)

Broedbaek, Kasper; Siersma, Volkert Dirk; Andersen, Jon T

2011-01-01

This study applied a case-control approach to investigate the association between low-grade inflammation, defined by high values within the normal range of C-reactive protein (CRP) and interleukin-6 (IL-6), and urinary markers of nucleic acid oxidation. No differences in excretion of urinary...... markers of nucleic acid oxidation between cases and controls were found and multivariable linear regression analysis showed no association between urinary markers of nucleic acid oxidation and inflammatory markers. Post-hoc multivariable linear regression analysis showed significant associations between...... suggest that low-grade inflammation only has a negligible impact on whole body nucleic acid oxidation, whereas iron status seems to be of great importance....
Accuracy assessment of the linear Poisson-Boltzmann equation and reparametrization of the OBC generalized Born model for nucleic acids and nucleic acid-protein complexes.

Science.gov (United States)

Fogolari, Federico; Corazza, Alessandra; Esposito, Gennaro

2015-04-05

The generalized Born model in the Onufriev, Bashford, and Case (Onufriev et al., Proteins: Struct Funct Genet 2004, 55, 383) implementation has emerged as one of the best compromises between accuracy and speed of computation. For simulations of nucleic acids, however, a number of issues should be addressed: (1) the generalized Born model is based on a linear model and the linearization of the reference Poisson-Boltmann equation may be questioned for highly charged systems as nucleic acids; (2) although much attention has been given to potentials, solvation forces could be much less sensitive to linearization than the potentials; and (3) the accuracy of the Onufriev-Bashford-Case (OBC) model for nucleic acids depends on fine tuning of parameters. Here, we show that the linearization of the Poisson Boltzmann equation has mild effects on computed forces, and that with optimal choice of the OBC model parameters, solvation forces, essential for molecular dynamics simulations, agree well with those computed using the reference Poisson-Boltzmann model. © 2015 Wiley Periodicals, Inc.
Possibilities for prognostication of radiation injury in rats by leucocyte nucleic acid levels

International Nuclear Information System (INIS)

Minkova, M.; Pantev, T.

1988-01-01

The possibilities to prognosticate acute radiation injury by the changes in the amount of nucleic acids in the leucocytes was studied. Experiments were carried out on male Wistar albino rats, gamma-irradiated with nonlethal and sublethal doses of 0.5, 2 and 4 Gy and lethal dose of 8 Gy (LD 90/30 ). The nucleic acid content and the total leucocyte count were determined at definite intervals on days 1-30. The changes in the nucleic acids in nonlethally and sublethally irradiated animals had phase nature, with a clear-cut abortive increase in their amount on days 7-10. In lethally irradiated animals the phase character of the changes was lost and the abortive peak disappeared. By reducing the effectiveness of the lethal radiation dose survival of the population increased from 10-75% through physical and from 10-70% - through chemical protection. The nucleic acid dynamics showed features typical for an injury with possible survival - appearance of abortive peak and resumption of their normal values. It is assumed that determination of leucocyte nucleic acid content may be used for early prognostication of radiation injury, as it allows keen differentiation of the lethal from nonlethal outcome of radiation sickness. The absence of abortive peak (over 50%) by day 14 post-irradiation is a poor prognostic sign

Biomimetic High Density Lipoprotein Nanoparticles For Nucleic Acid Delivery

Science.gov (United States)

McMahon, Kaylin M.; Mutharasan, R. Kannan; Tripathy, Sushant; Veliceasa, Dorina; Bobeica, Mariana; Shumaker, Dale K.; Luthi, Andrea J.; Helfand, Brian T.; Ardehali, Hossein; Mirkin, Chad A.; Volpert, Olga; Thaxton, C. Shad

2014-01-01

We report a gold nanoparticle-templated high density lipoprotein (HDL AuNP) platform for gene therapy which combines lipid-based nucleic acid transfection strategies with HDL biomimicry. For proof-of-concept, HDL AuNPs are shown to adsorb antisense cholesterylated DNA. The conjugates are internalized by human cells, can be tracked within cells using transmission electron microscopy (TEM), and regulate target gene expression. Overall, the ability to directly image the AuNP core within cells, the chemical tailorability of the HDL AuNP platform, and the potential for cell-specific targeting afforded by HDL biomimicry make this platform appealing for nucleic acid delivery. PMID:21319839
Nucleic acid aptamers: an emerging frontier in cancer therapy.

Science.gov (United States)

Zhu, Guizhi; Ye, Mao; Donovan, Michael J; Song, Erqun; Zhao, Zilong; Tan, Weihong

2012-11-04

The last two decades have witnessed the development and application of nucleic acid aptamers in a variety of fields, including target analysis, disease therapy, and molecular and cellular engineering. The efficient and widely applicable aptamer selection, reproducible chemical synthesis and modification, generally impressive target binding selectivity and affinity, relatively rapid tissue penetration, low immunogenicity, and rapid systemic clearance make aptamers ideal recognition elements for use as therapeutics or for in vivo delivery of therapeutics. In this feature article, we discuss the development and biomedical application of nucleic acid aptamers, with emphasis on cancer cell aptamer isolation, targeted cancer therapy, oncology biomarker identification and drug discovery.
Predicting nucleic acid binding interfaces from structural models of proteins.

Science.gov (United States)

Dror, Iris; Shazman, Shula; Mukherjee, Srayanta; Zhang, Yang; Glaser, Fabian; Mandel-Gutfreund, Yael

2012-02-01

The function of DNA- and RNA-binding proteins can be inferred from the characterization and accurate prediction of their binding interfaces. However, the main pitfall of various structure-based methods for predicting nucleic acid binding function is that they are all limited to a relatively small number of proteins for which high-resolution three-dimensional structures are available. In this study, we developed a pipeline for extracting functional electrostatic patches from surfaces of protein structural models, obtained using the I-TASSER protein structure predictor. The largest positive patches are extracted from the protein surface using the patchfinder algorithm. We show that functional electrostatic patches extracted from an ensemble of structural models highly overlap the patches extracted from high-resolution structures. Furthermore, by testing our pipeline on a set of 55 known nucleic acid binding proteins for which I-TASSER produces high-quality models, we show that the method accurately identifies the nucleic acids binding interface on structural models of proteins. Employing a combined patch approach we show that patches extracted from an ensemble of models better predicts the real nucleic acid binding interfaces compared with patches extracted from independent models. Overall, these results suggest that combining information from a collection of low-resolution structural models could be a valuable approach for functional annotation. We suggest that our method will be further applicable for predicting other functional surfaces of proteins with unknown structure. Copyright © 2011 Wiley Periodicals, Inc.
Chemical consequences of irradiating nucleic acids

International Nuclear Information System (INIS)

Ward, J.F.

1976-01-01

On the basis of literature data, a discussion is presented of the DNA damage which would be produced in a cellular environment and an attempt is made to place this damage in perspective as a potential hazard in food irradiation. The topics discussed are radiation damage mechanisms, OH reactions with DNA, base products, sugar products, and evaluation of damage from irradiated nucleic acids
Microwave-assisted hydrolysis of phosphonate diesters: an efficient protocol for the preparation of phosphonic acids

Czech Academy of Sciences Publication Activity Database

Jansa, Petr; Baszczyňski, Ondřej; Procházková, Eliška; Dračínský, Martin; Janeba, Zlatko

2012-01-01

Roč. 14, č. 8 (2012), s. 2282-2288 ISSN 1463-9262 R&D Projects: GA MV VG20102015046 Institutional research plan: CEZ:AV0Z40550506 Keywords : acyclic nucleoside phosphonates * DNA virus * retrovirus infections * cidofovir * organic-synthesis Subject RIV: CC - Organic Chemistry Impact factor: 6.828, year: 2012
New nucleic acid testing devices to diagnose infectious diseases in resource-limited settings.

Science.gov (United States)

Maffert, P; Reverchon, S; Nasser, W; Rozand, C; Abaibou, H

2017-10-01

Point-of-care diagnosis based on nucleic acid testing aims to incorporate all the analytical steps, from sample preparation to nucleic acid amplification and detection, in a single device. This device needs to provide a low-cost, robust, sensitive, specific, and easily readable analysis. Microfluidics has great potential for handling small volumes of fluids on a single platform. Microfluidic technology has recently been applied to paper, which is already used in low-cost lateral flow tests. Nucleic acid extraction from a biological specimen usually requires cell filtration and lysis on specific membranes, while affinity matrices, such as chitosan or polydiacetylene, are well suited to concentrating nucleic acids for subsequent amplification. Access to electricity is often difficult in resource-limited areas, so the amplification step needs to be equipment-free. Consequently, the reaction has to be isothermal to alleviate the need for a thermocycler. LAMP, NASBA, HDA, and RPA are examples of the technologies available. Nucleic acid detection techniques are currently based on fluorescence, colorimetry, or chemiluminescence. For point-of-care diagnostics, the results should be readable with the naked eye. Nowadays, interpretation and communication of results to health professionals could rely on a smartphone, used as a telemedicine device. The major challenge of creating an "all-in-one" diagnostic test involves the design of an optimal solution and a sequence for each analytical step, as well as combining the execution of all these steps on a single device. This review provides an overview of available materials and technologies which seem to be adapted to point-of-care nucleic acid-based diagnosis, in low-resource areas.
Non-invasive Oil-Based Method to Increase Topical Delivery of Nucleic Acids to Skin.

Science.gov (United States)

Vij, Manika; Alam, Shamshad; Gupta, Nidhi; Gotherwal, Vishvabandhu; Gautam, Hemlata; Ansari, Kausar M; Santhiya, Deenan; Natarajan, Vivek T; Ganguli, Munia

2017-06-07

Topical delivery of nucleic acids to skin has huge prospects in developing therapeutic interventions for cutaneous disorders. In spite of initial success, clinical translation is vastly impeded by the constraints of bioavailability as well as stability in metabolically active environment of skin. Various physical and chemical methods used to overcome these limitations involve invasive procedures or compounds that compromise skin integrity. Hence, there is an increasing demand for developing safe skin penetration enhancers for efficient nucleic acid delivery to skin. Here, we demonstrate that pretreatment of skin with silicone oil can increase the transfection efficiency of non-covalently associated peptide-plasmid DNA nanocomplexes in skin ex vivo and in vivo. The method does not compromise skin integrity, as indicated by microscopic evaluation of cellular differentiation, tissue architecture, enzyme activity assessment, dye penetration tests using Franz assay, and cytotoxicity and immunogenicity analyses. Stability of nanocomplexes is not hampered on pretreatment, thereby avoiding nuclease-mediated degradation. The mechanistic insights through Fourier transform infrared (FTIR) spectroscopy reveal some alterations in the skin hydration status owing to possible occlusion effects of the enhancer. Overall, we describe a topical, non-invasive, efficient, and safe method that can be used to increase the penetration and delivery of plasmid DNA to skin for possible therapeutic applications. Copyright © 2017 The American Society of Gene and Cell Therapy. Published by Elsevier Inc. All rights reserved.
Carbon composite micro- and nano-tubes-based electrodes for detection of nucleic acids

Directory of Open Access Journals (Sweden)

Huska Dalibor

2011-01-01

Full Text Available Abstract The first aim of this study was to fabricate vertically aligned multiwalled carbon nanotubes (MWCNTs. MWCNTs were successfully prepared by using plasma enhanced chemical vapour deposition. Further, three carbon composite electrodes with different content of carbon particles with various shapes and sizes were prepared and tested on measuring of nucleic acids. The dependences of adenine peak height on the concentration of nucleic acid sample were measured. Carbon composite electrode prepared from a mixture of glassy and spherical carbon powder and MWCNTs had the highest sensitivity to nucleic acids. Other interesting result is the fact that we were able to distinguish signals for all bases using this electrode.
Engineering nucleic acid structures for programmable molecular circuitry and intracellular biocomputation

Science.gov (United States)

Li, Jiang; Green, Alexander A.; Yan, Hao; Fan, Chunhai

2017-11-01

Nucleic acids have attracted widespread attention due to the simplicity with which they can be designed to form discrete structures and programmed to perform specific functions at the nanoscale. The advantages of DNA/RNA nanotechnology offer numerous opportunities for in-cell and in-vivo applications, and the technology holds great promise to advance the growing field of synthetic biology. Many elegant examples have revealed the potential in integrating nucleic acid nanostructures in cells and in vivo where they can perform important physiological functions. In this Review, we summarize the current abilities of DNA/RNA nanotechnology to realize applications in live cells and then discuss the key problems that must be solved to fully exploit the useful properties of nanostructures. Finally, we provide viewpoints on how to integrate the tools provided by DNA/RNA nanotechnology and related new technologies to construct nucleic acid nanostructure-based molecular circuitry for synthetic biology.
Key Aspects of Nucleic Acid Library Design for in Vitro Selection

Science.gov (United States)

Vorobyeva, Maria A.; Davydova, Anna S.; Vorobjev, Pavel E.; Pyshnyi, Dmitrii V.; Venyaminova, Alya G.

2018-01-01

Nucleic acid aptamers capable of selectively recognizing their target molecules have nowadays been established as powerful and tunable tools for biospecific applications, be it therapeutics, drug delivery systems or biosensors. It is now generally acknowledged that in vitro selection enables one to generate aptamers to almost any target of interest. However, the success of selection and the affinity of the resulting aptamers depend to a large extent on the nature and design of an initial random nucleic acid library. In this review, we summarize and discuss the most important features of the design of nucleic acid libraries for in vitro selection such as the nature of the library (DNA, RNA or modified nucleotides), the length of a randomized region and the presence of fixed sequences. We also compare and contrast different randomization strategies and consider computer methods of library design and some other aspects. PMID:29401748
Molecular structure and interactions of nucleic acid components in nanoparticles: ab initio calculations

International Nuclear Information System (INIS)

Rubin, Yu.V.; Belous, L.F.

2012-01-01

Self-associates of nucleic acid components (stacking trimers and tetramers of the base pairs of nucleic acids) and short fragments of nucleic acids are nanoparticles (linear sizes of these particles are more than 10 A). Modern quantum-mechanical methods and softwares allow one to perform ab initio calculations of the systems consisting of 150-200 atoms with enough large basis sets (for example, 6-31G * ). The aim of this work is to reveal the peculiarities of molecular and electronic structures, as well as the energy features of nanoparticles of nucleic acid components. We had carried out ab initio calculations of the molecular structure and interactions in the stacking dimer, trimer, and tetramer of nucleic base pairs and in the stacking (TpG)(ApC) dimer and (TpGpC) (ApCpG) trimer of nucleotides, which are small DNA fragments. The performed calculations of molecular structures of dimers and trimers of nucleotide pairs showed that the interplanar distance in the structures studied is equal to 3.2 A on average, and the helical angle in a trimer is approximately equal to 30 o : The distance between phosphor atoms in neighboring chains is 13.1 A. For dimers and trimers under study, we calculated the horizontal interaction energies. The analysis of interplanar distances and angles between nucleic bases and their pairs in the calculated short oligomers of nucleic acid base pairs (stacking dimer, trimer, and tetramer) has been carried out. Studies of interactions in the calculated short oligomers showed a considerable role of the cross interaction in the stabilization of the structures. The contribution of cross interactions to the horizontal interactions grows with the length of an oligomer. Nanoparticle components get electric charges in nanoparticles. Longwave low-intensity bands can appear in the electron spectra of nanoparticles.
RNA:DNA Ratio and Other Nucleic Acid Derived Indices in Marine Ecology

Directory of Open Access Journals (Sweden)

Luis ChÃƒÂcharo

2008-08-01

Full Text Available Some of most used indicators in marine ecology are nucleic acid-derived indices. They can be divided by target levels in three groups: 1 at the organism level as ecophysiologic indicators, indicators such as RNA:DNA ratios, DNA:dry weight and RNA:protein, 2 at the population level, indicators such as growth rate, starvation incidence or fisheries impact indicators, and 3 at the community level, indicators such as trophic interactions, exergy indices and prey identification. The nucleic acids derived indices, especially RNA:DNA ratio, have been applied with success as indicators of nutritional condition, well been and growth in marine organisms. They are also useful as indicators of natural or anthropogenic impacts in marine population and communities, such as upwelling or dredge fisheries, respectively. They can help in understanding important issues of marine ecology such as trophic interactions in marine environment, fish and invertebrate recruitment failure and biodiversity changes, without laborious work of counting, measuring and identification of small marine organisms. Besides the objective of integrate nucleic acid derived indices across levels of organization, the paper will also include a general characterization of most used nucleic acid derived indices in marine ecology and also advantages and limitations of them. We can conclude that using indicators, such RNA:DNA ratios and other nucleic acids derived indices concomitantly with organism and ecosystems measures of responses to climate change (distribution, abundance, activity, metabolic rate, survival will allow for the development of more rigorous and realistic predictions of the effects of anthropogenic climate change on marine systems.
Pyrazine Nucleic Acids: From Small Molecules to Proto-Informational Polymers

Science.gov (United States)

Wong, S. B.; Gately, M.; Young, E.; Krishnamurthy, R.; Weber, A. L.; Campbell, T.

2017-07-01

Pyrazine nucleosides are derivable from amino acid amides and pentoses under plausibly prebiotic conditions. Pyrazines share features similar to adenine or thymine, and may behave as an informational polymer when polymerized as pyrazine nucleic acid.
Phosphonate biosynthesis and catabolism: a treasure trove of unusual enzymology.

Science.gov (United States)

Peck, Spencer C; van der Donk, Wilfred A

2013-08-01

Natural product biosynthesis has proven a fertile ground for the discovery of novel chemistry. Herein we review the progress made in elucidating the biosynthetic pathways of phosphonate and phosphinate natural products such as the antibacterial compounds dehydrophos and fosfomycin, the herbicidal phosphinothricin-containing peptides, and the antimalarial compound FR-900098. In each case, investigation of the pathway has yielded unusual, and often unprecedented, biochemistry. Likewise, recent investigations have uncovered novel ways to cleave the CP bond to yield phosphate under phosphorus starvation conditions. These include the discovery of novel oxidative cleavage of the CP bond catalyzed by PhnY and PhnZ as well as phosphonohydrolases that liberate phosphate from phosphonoacetate. Perhaps the crown jewel of phosphonate catabolism has been the recent resolution of the longstanding problem of the C-P lyase responsible for reductively cleaving the CP bond of a number of different phosphonates to release phosphate. Taken together, the strides made on both metabolic and catabolic fronts illustrate an array of fascinating biochemistry. Copyright © 2013 Elsevier Ltd. All rights reserved.
Nucleic acid secondary structure prediction and display.

OpenAIRE

Stüber, K

1986-01-01

A set of programs has been developed for the prediction and display of nucleic acid secondary structures. Information from experimental data can be used to restrict or enforce secondary structural elements. The predictions can be displayed either on normal line printers or on graphic devices like plotters or graphic terminals.
Sol–gel synthesis of tantalum oxide and phosphonic acid-modified carbon nanotubes composite coatings on titanium surfaces

International Nuclear Information System (INIS)

Maho, Anthony; Detriche, Simon; Delhalle, Joseph; Mekhalif, Zineb

2013-01-01

Carbon nanotubes used as fillers in composite materials are more and more appreciated for the outstanding range of accessible properties and functionalities they generate in numerous domains of nanotechnologies. In the framework of biological and medical sciences, and particularly for orthopedic applications and devices (prostheses, implants, surgical instruments, …), titanium substrates covered by tantalum oxide/carbon nanotube composite coatings have proved to constitute interesting and successful platforms for the conception of solid and biocompatible biomaterials inducing the osseous regeneration processes (hydroxyapatite growth, osteoblasts attachment). This paper describes an original strategy for the conception of resistant and homogeneous tantalum oxide/carbon nanotubes layers on titanium through the introduction of carbon nanotubes functionalized by phosphonic acid moieties (-P(=O)(OH) 2 ). Strong covalent C-P bonds are specifically inserted on their external sidewalls with a ratio of two phosphonic groups per anchoring point. Experimental results highlight the stronger “tantalum capture agent” effect of phosphonic-modified nanotubes during the sol–gel formation process of the deposits compared to nanotubes bearing oxidized functions (-OH, -C=O, -C(=O)OH). Particular attention is also paid to the relative impact of the rate of functionalization and the dispersion degree of the carbon nanotubes in the coatings, as well as their wrapping level by the tantalum oxide matrix material. The resulting effect on the in vitro growth of hydroxyapatite is also evaluated to confirm the primary osseous bioactivity of those materials. Chemical, structural and morphological features of the different composite deposits described herein are assessed by X-ray photoelectron spectroscopy (XPS), scanning (SEM) and transmission (TEM) electronic microscopies, energy dispersive X-rays analysis (EDX) and peeling tests. Highlights: ► Formation of tantalum/carbon nanotube
Sol–gel synthesis of tantalum oxide and phosphonic acid-modified carbon nanotubes composite coatings on titanium surfaces

Energy Technology Data Exchange (ETDEWEB)

Maho, Anthony [Laboratory of Chemistry and Electrochemistry of Surfaces, University of Namur (FUNDP), Rue de Bruxelles 61, B-5000 Namur (Belgium); Fonds pour la Formation à la Recherche dans l' Industrie et dans l' Agriculture (FRIA), Rue d' Egmont 5, B-1000 Bruxelles (Belgium); Detriche, Simon; Delhalle, Joseph [Laboratory of Chemistry and Electrochemistry of Surfaces, University of Namur (FUNDP), Rue de Bruxelles 61, B-5000 Namur (Belgium); Mekhalif, Zineb, E-mail: zineb.mekhalif@fundp.ac.be [Laboratory of Chemistry and Electrochemistry of Surfaces, University of Namur (FUNDP), Rue de Bruxelles 61, B-5000 Namur (Belgium)

2013-07-01

Carbon nanotubes used as fillers in composite materials are more and more appreciated for the outstanding range of accessible properties and functionalities they generate in numerous domains of nanotechnologies. In the framework of biological and medical sciences, and particularly for orthopedic applications and devices (prostheses, implants, surgical instruments, …), titanium substrates covered by tantalum oxide/carbon nanotube composite coatings have proved to constitute interesting and successful platforms for the conception of solid and biocompatible biomaterials inducing the osseous regeneration processes (hydroxyapatite growth, osteoblasts attachment). This paper describes an original strategy for the conception of resistant and homogeneous tantalum oxide/carbon nanotubes layers on titanium through the introduction of carbon nanotubes functionalized by phosphonic acid moieties (-P(=O)(OH){sub 2}). Strong covalent C-P bonds are specifically inserted on their external sidewalls with a ratio of two phosphonic groups per anchoring point. Experimental results highlight the stronger “tantalum capture agent” effect of phosphonic-modified nanotubes during the sol–gel formation process of the deposits compared to nanotubes bearing oxidized functions (-OH, -C=O, -C(=O)OH). Particular attention is also paid to the relative impact of the rate of functionalization and the dispersion degree of the carbon nanotubes in the coatings, as well as their wrapping level by the tantalum oxide matrix material. The resulting effect on the in vitro growth of hydroxyapatite is also evaluated to confirm the primary osseous bioactivity of those materials. Chemical, structural and morphological features of the different composite deposits described herein are assessed by X-ray photoelectron spectroscopy (XPS), scanning (SEM) and transmission (TEM) electronic microscopies, energy dispersive X-rays analysis (EDX) and peeling tests. Highlights: ► Formation of tantalum
Structural aspects of catalytic mechanisms of endonucleases and their binding to nucleic acids

Energy Technology Data Exchange (ETDEWEB)

Zhukhlistova, N. E.; Balaev, V. V.; Lyashenko, A. V.; Lashkov, A. A., E-mail: alashkov83@gmail.com [Russian Academy of Sciences, Shubnikov Institute of Crystallography (Russian Federation)

2012-05-15

Endonucleases (EC 3.1) are enzymes of the hydrolase class that catalyze the hydrolytic cleavage of deoxyribonucleic and ribonucleic acids at any region of the polynucleotide chain. Endonucleases are widely used both in biotechnological processes and in veterinary medicine as antiviral agents. Medical applications of endonucleases in human cancer therapy hold promise. The results of X-ray diffraction studies of the spatial organization of endonucleases and their complexes and the mechanism of their action are analyzed and generalized. An analysis of the structural studies of this class of enzymes showed that the specific binding of enzymes to nucleic acids is characterized by interactions with nitrogen bases and the nucleotide backbone, whereas the nonspecific binding of enzymes is generally characterized by interactions only with the nucleic-acid backbone. It should be taken into account that the specificity can be modulated by metal ions and certain low-molecular-weight organic compounds. To test the hypotheses about specific and nonspecific nucleic-acid-binding proteins, it is necessary to perform additional studies of atomic-resolution three-dimensional structures of enzyme-nucleic-acid complexes by methods of structural biology.
Nucleic acid-based diagnostics for infectious diseases in public health affairs.

Science.gov (United States)

Yu, Albert Cheung-Hoi; Vatcher, Greg; Yue, Xin; Dong, Yan; Li, Mao Hua; Tam, Patrick H K; Tsang, Parker Y L; Wong, April K Y; Hui, Michael H K; Yang, Bin; Tang, Hao; Lau, Lok-Ting

2012-06-01

Infectious diseases, mostly caused by bacteria and viruses but also a result of fungal and parasitic infection, have been one of the most important public health concerns throughout human history. The first step in combating these pathogens is to get a timely and accurate diagnosis at an affordable cost. Many kinds of diagnostics have been developed, such as pathogen culture, biochemical tests and serological tests, to help detect and fight against the causative agents of diseases. However, these diagnostic tests are generally unsatisfactory because they are not particularly sensitive and specific and are unable to deliver speedy results. Nucleic acid-based diagnostics, detecting pathogens through the identification of their genomic sequences, have shown promise to overcome the above limitations and become more widely adopted in clinical tests. Here we review some of the most popular nucleic acid-based diagnostics and focus on their adaptability and applicability to routine clinical usage. We also compare and contrast the characteristics of different types of nucleic acid-based diagnostics.
Assembly of phosphonic acids on GaN and AlGaN

Energy Technology Data Exchange (ETDEWEB)

Simpkins, B S; Stine, R; Theodore, N D; Pehrsson, P E [Chemistry Division, Naval Research Laboratory, Washington DC (United States); Hong, S [Thomas Jefferson High School, McClean, VA (United States); Maekinen, A J [Optical Sciences Division, Naval Research Laboratory, Washington, DC (United States); Mastro, M A; Eddy, C R Jr [Electronics Science and Technology Division, Naval Research Laboratory, Washington, DC (United States)

2010-01-13

Self-assembled monolayers of octadecylphosphonic acid and 16-phosphonohexadecanoic acid (PHDA) were formed on the semiconductor substrates gallium nitride (GaN) and aluminium gallium nitride (AlGaN). The presence of the molecular layers was verified through x-ray photoelectron spectroscopy and ultraviolet photoelectron spectroscopy. Structural information was acquired with infrared spectroscopy which verified the bonding orientation of the carboxyl-containing PHDA. The impact of the molecular layers on the channel conductivity and the surface electronic structure of an AlGaN/GaN heterostructure was measured. Our results indicate that pinning of the surface Fermi level prohibits modification of the channel conductivity by the layer. However, a surface dipole of {approx}0.8 eV is present and associated with both phosphonic acid layers. These results are of direct relevance to field-effect-based biochemical sensors and metal-semiconductor contact formation for this system and provide a fundamental basis for further applications of GaN and AlGaN technology in the fields of biosensing and microelectronics.

The relationship between odd- and branched-chain fatty acids and microbial nucleic acid bases in rumen.

Science.gov (United States)

Liu, Keyuan; Hao, Xiaoyan; Li, Yang; Luo, Guobin; Zhang, Yonggen; Xin, Hangshu

2017-11-01

This study aims to identify the relationship between odd- and branched-chain fatty acids (OBCFAs) and microbial nucleic acid bases in the rumen, and to establish a model to accurately predict microbial protein flow by using OBCFA. To develop the regression equations, data on the rumen contents of individual cows were obtained from 2 feeding experiments. In the first experiment, 3 rumen-fistulated dry dairy cows arranged in a 3×3 Latin square were fed diets of differing forage to concentration ratios (F:C). The second experiment consisted of 9 lactating Holstein dairy cows of similar body weights at the same stage of pregnancy. For each lactation stage, 3 cows with similar milk production were selected. The rumen contents were sampled at 4 time points of every two hours after morning feeding 6 h, and then to analyse the concentrations of OBCFA and microbial nucleic acid bases in the rumen samples. The ruminal bacteria nucleic acid bases were significantly influenced by feeding diets of differing forge to concentration ratios and lactation stages of dairy cows (pacids and C15:0 isomers, strongly correlated with the microbial nucleic acid bases in the rumen (pacid bases established by ruminal OBCFAs contents showed a good predictive capacity, as indicated by reasonably low standard errors and high R-squared values. This finding suggests that the rumen OBCFA composition could be used as an internal marker of rumen microbial matter.
Novel open-framework architectures in lanthanide phosphonates

Science.gov (United States)

Groves, John A.; Stephens, Nicholas F.; Wright, Paul A.; Lightfoot, Philip

2006-03-01

Two novel three-dimensional lanthanide coordination polymers have been prepared hydrothermally with the phosphonic acid N,N-piperazine bis(methylenephosphonic acid), H 2O 3PCH 2N(C 2H 4) 2NCH 2PO 3H 2 ( LH 4). The structures of Gd 2( LH 2) 3ṡ3H 2O (I) and Nd 2( LH 2) 3ṡ9H 2O (II) have been characterised by single crystal X-ray techniques. One-dimensional 'lanthanide-phosphate' chains are a key feature in both structures, although there are major structural differences between the chains, with (I) displaying octahedral GdO 6 coordination and (II) showing eight-coordinate NdO 8 polyhedra. In each case, three-dimensional connectivity is completed by coordination of the phosphonate group resulting in open framework structures encapsulating loosely bound water molecules. Isostructural Y 3+ and Yb 3+ analogues of (I) have been prepared, suggesting that cation size is a key factor in controlling the differing reaction products. In the case of Y 2( LH 2) 3ṡ5H 2O, isostructural to (I), it is shown that the extra-framework water molecules may be removed reversibly without framework collapse. Structural relationships to other known lanthanide phosphonates are discussed.
Spherical Nucleic Acids as Intracellular Agents for Nucleic Acid Based Therapeutics

Science.gov (United States)

Hao, Liangliang

Recent functional discoveries on the noncoding sequences of human genome and transcriptome could lead to revolutionary treatment modalities because the noncoding RNAs (ncRNAs) can be applied as therapeutic agents to manipulate disease-causing genes. To date few nucleic acid-based therapeutics have been translated into the clinic due to challenges in the delivery of the oligonucleotide agents in an effective, cell specific, and non-toxic fashion. Unmodified oligonucleotide agents are destroyed rapidly in biological fluids by enzymatic degradation and have difficulty crossing the plasma membrane without the aid of transfection reagents, which often cause inflammatory, cytotoxic, or immunogenic side effects. Spherical nucleic acids (SNAs), nanoparticles consisting of densely organized and highly oriented oligonucleotides, pose one possible solution to circumventing these problems in both the antisense and RNA interference (RNAi) pathways. The unique three dimensional architecture of SNAs protects the bioactive oligonucleotides from unspecific degradation during delivery and supports their targeting of class A scavenger receptors and endocytosis via a lipid-raft-dependent, caveolae-mediated pathway. Owing to their unique structure, SNAs are able to cross cell membranes and regulate target genes expression as a single entity, without triggering the cellular innate immune response. Herein, my thesis has focused on understanding the interactions between SNAs and cellular components and developing SNA-based nanostructures to improve therapeutic capabilities. Specifically, I developed a novel SNA-based, nanoscale agent for delivery of therapeutic oligonucleotides to manipulate microRNAs (miRNAs), the endogenous post-transcriptional gene regulators. I investigated the role of SNAs involving miRNAs in anti-cancer or anti-inflammation responses in cells and in in vivo murine disease models via systemic injection. Furthermore, I explored using different strategies to construct
Nanopore biosensors for detection of proteins and nucleic acids

NARCIS (Netherlands)

Maglia, Giovanni; Soskine, Mikhael

2014-01-01

Described herein are nanopore biosensors based on a modified cytolysin protein. The nanopore biosensors accommodate macromoiecules including proteins and nucleic acids, and may additionally comprise ligands with selective binding properties.
Real-time electrochemical monitoring of isothermal helicase-dependent amplification of nucleic acids.

Science.gov (United States)

Kivlehan, Francine; Mavré, François; Talini, Luc; Limoges, Benoît; Marchal, Damien

2011-09-21

We described an electrochemical method to monitor in real-time the isothermal helicase-dependent amplification of nucleic acids. The principle of detection is simple and well-adapted to the development of portable, easy-to-use and inexpensive nucleic acids detection technologies. It consists of monitoring a decrease in the electrochemical current response of a reporter DNA intercalating redox probe during the isothermal DNA amplification. The method offers the possibility to quantitatively analyze target nucleic acids in less than one hour at a single constant temperature, and to perform at the end of the isothermal amplification a DNA melt curve analysis for differentiating between specific and non-specific amplifications. To illustrate the potentialities of this approach for the development of a simple, robust and low-cost instrument with high throughput capability, the method was validated with an electrochemical system capable of monitoring up to 48 real-time isothermal HDA reactions simultaneously in a disposable microplate consisting of 48-electrochemical microwells. Results obtained with this approach are comparable to that obtained with a well-established but more sophisticated and expensive fluorescence-based method. This makes for a promising alternative detection method not only for real-time isothermal helicase-dependent amplification of nucleic acid, but also for other isothermal DNA amplification strategies.
A quantitative measure of chirality inside nucleic acid databank.

Science.gov (United States)

Pietropaolo, Adriana; Parrinello, Michele

2011-08-01

We show the capability of a chirality index (Pietropaolo et al., Proteins 2008;70:667-677) to investigate nucleic acid structures because of its high sensitivity to helical conformations. By analyzing selected structures of DNA and RNA, we have found that sequences rich in cytosine and guanine have a tendency to left-handed chirality, in contrast to regions rich in adenine or thymine which show strong negative, right-handed, chirality values. We also analyze RNA structures, where specific loops and hairpin motifs are characterized by a well-defined chirality value. We find that in nucleosome the chirality is exalted, whereas in ribosome it is reduced. Our results illustrate the sensitivity of this descriptor for nucleic acid conformations. Copyright © 2011 Wiley-Liss, Inc.
Labelling of nucleic acid components with tritium by hydrogenolysis of corresponding precursors

International Nuclear Information System (INIS)

Myasoedov, V.F.; Kvyatkovskij, Yu.G.; Sidorov, G.V.

1981-01-01

To desalt the luates of liquid column chromatography containing components of the nucleic acids different types of activated carbons are used: AG-5, Ou-4, KAD, BAU (SU), Norit (GB) and Carboafin (CS). The Carborafin (CS) carbon proved to be the most efficient for the purpose. Dependences of the adsorption degree on pH, the time of the phases contact, temperature, concentration of the salt background (ammonium formite, lithium chloride) as well as adsorption isotherm are determined for the activated carbon. Desorption conditions of the nucleic acids components from the carbon are studied. It is shown that quantitative desorption is achieved when 1n solution of ammonia is used in 50% ethanole for 50-60 min. Data on practical application of the method to desalt the eluates containing tritiated nucleic acid components with a high activity are presented [ru
Tetrofuranose nucleoside phosphonic acids: Synthesis and properties

Czech Academy of Sciences Publication Activity Database

Poláková, Ivana; Buděšínský, Miloš; Točík, Zdeněk; Rosenberg, Ivan

2011-01-01

Roč. 76, č. 5 (2011), s. 503-536 ISSN 0010-0765 R&D Projects: GA AV ČR KAN200520801; GA ČR GA203/09/0820; GA MŠk(CZ) LC06061; GA MŠk(CZ) LC06077 Institutional research plan: CEZ:AV0Z40550506 Keywords : tetrofuranosyl phosphonate * nucleotide analogues * phosphonomethoxy nucleosides * sugar hydroxyphosphonates Subject RIV: CC - Organic Chemistry Impact factor: 1.283, year: 2011
Dioxaphosphorinane-constrained nucleic Acid dinucleotides as tools for structural tuning of nucleic acids.

Science.gov (United States)

Catana, Dan-Andrei; Renard, Brice-Loïc; Maturano, Marie; Payrastre, Corinne; Tarrat, Nathalie; Escudier, Jean-Marc

2012-01-01

We describe a rational approach devoted to modulate the sugar-phosphate backbone geometry of nucleic acids. Constraints were generated by connecting one oxygen of the phosphate group to a carbon of the sugar moiety. The so-called dioxaphosphorinane rings were introduced at key positions along the sugar-phosphate backbone allowing the control of the six-torsion angles α to ζ defining the polymer structure. The syntheses of all the members of the D-CNA family are described, and we emphasize the effect on secondary structure stabilization of a couple of diastereoisomers of α,β-D-CNA exhibiting wether B-type canonical values or not.
Dioxaphosphorinane-Constrained Nucleic Acid Dinucleotides as Tools for Structural Tuning of Nucleic Acids

Directory of Open Access Journals (Sweden)

Dan-Andrei Catana

2012-01-01

Full Text Available We describe a rational approach devoted to modulate the sugar-phosphate backbone geometry of nucleic acids. Constraints were generated by connecting one oxygen of the phosphate group to a carbon of the sugar moiety. The so-called dioxaphosphorinane rings were introduced at key positions along the sugar-phosphate backbone allowing the control of the six-torsion angles α to ζ defining the polymer structure. The syntheses of all the members of the D-CNA family are described, and we emphasize the effect on secondary structure stabilization of a couple of diastereoisomers of α,β-D-CNA exhibiting wether B-type canonical values or not.
System for portable nucleic acid testing in low resource settings

Science.gov (United States)

Lu, Hsiang-Wei; Roskos, Kristina; Hickerson, Anna I.; Carey, Thomas; Niemz, Angelika

2013-03-01

Our overall goal is to enable timely diagnosis of infectious diseases through nucleic acid testing at the point-of-care and in low resource settings, via a compact system that integrates nucleic acid sample preparation, isothermal DNA amplification, and nucleic acid lateral flow (NALF) detection. We herein present an interim milestone, the design of the amplification and detection subsystem, and the characterization of thermal and fluidic control and assay execution within this system. Using an earlier prototype of the amplification and detection unit, comprised of a disposable cartridge containing flexible pouches, passive valves, and electrolysis-driven pumps, in conjunction with a small heater, we have demonstrated successful execution of an established and clinically validated isothermal loop-mediated amplification (LAMP) reaction targeting Mycobacterium tuberculosis (M.tb) DNA, coupled to NALF detection. The refined design presented herein incorporates miniaturized and integrated electrolytic pumps, novel passive valves, overall design changes to facilitate integration with an upstream sample preparation unit, and a refined instrument design that automates pumping, heating, and timing. Nucleic acid amplification occurs in a two-layer pouch that facilitates fluid handling and appropriate thermal control. The disposable cartridge is manufactured using low-cost and scalable techniques and forms a closed system to prevent workplace contamination by amplicons. In a parallel effort, we are developing a sample preparation unit based on similar design principles, which performs mechanical lysis of mycobacteria and DNA extraction from liquefied and disinfected sputum. Our next step is to combine sample preparation, amplification, and detection in a final integrated cartridge and device, to enable fully automated sample-in to answer-out diagnosis of active tuberculosis in primary care facilities of low-resource and high-burden countries.
Nucleic acid detection based on the use of microbeads: a review

International Nuclear Information System (INIS)

Rödiger, Stefan; Liebsch, Claudia; Schmidt, Carsten; Schierack, Peter; Lehmann, Werner; Resch-Genger, Ute; Schedler, Uwe

2014-01-01

Microbead-based technologies represent elegant and versatile approaches for highly parallelized quantitative multiparameter assays. They also form the basis of various techniques for detection and quantification of nucleic acids and proteins. Nucleic acid-based methods include hybridization assays, solid-phase PCR, sequencing, and trapping assays. Microbead assays have been improved in the past decades and are now important tools in routine and point-of-care diagnostics as well as in life science. Its advances include low costs, low workload, high speed and high-throughput automation. The potential of microbead-based assays therefore is apparent, and commercial applications can be found in the detection and discrimination of single nucleotide polymorphism, of pathogens, and in trapping assays. This review provides an overview on microbead-based platforms for biosensing with a main focus on nucleic acid detection (including amplification strategies and on selected probe systems using fluorescent labeling). Specific sections cover chemical properties of microbeads, the coupling of targets onto solid surfaces, microbead probe systems (mainly oligonucleotide probes), microbead detection schemes (with subsections on suspension arrays, microfluidic devices, and immobilized microbeads), quantification of nucleic acids, PCR in solution and the detection of amplicons, and methods for solid-phase amplification. We discuss selected trends such as microbead-coupled amplification, heterogeneous and homogenous DNA hybridization assays, real-time assays, melting curve analysis, and digital microbead assays. We finally discuss the relevance and trends of the methods in terms of high-level multiplexed analysis and their potential in diagnosis and personalized medicine. (author)
Recent advances in nanopore-based nucleic acid analysis and sequencing

International Nuclear Information System (INIS)

Shi, Jidong; Fang, Ying; Hou, Junfeng

2016-01-01

Nanopore-based sequencing platforms are transforming the field of genomic science. This review (containing 116 references) highlights some recent progress on nanopore-based nucleic acid analysis and sequencing. These studies are classified into three categories, biological, solid-state, and hybrid nanopores, according to their nanoporous materials. We begin with a brief description of the translocation-based detection mechanism of nanopores. Next, specific examples are given in nanopore-based nucleic acid analysis and sequencing, with an emphasis on identifying strategies that can improve the resolution of nanopores. This review concludes with a discussion of future research directions that will advance the practical applications of nanopore technology. (author)
The role of immunostimulatory nucleic acids in septic shock

Science.gov (United States)

Bleiblo, Farag; Michael, Paul; Brabant, Danielle; Ramana, Chilakamarti V; Tai, TC; Saleh, Mazen; Parrillo, Joseph E; Kumar, Anand; Kumar, Aseem

2012-01-01

Sepsis and its associated syndromes represent the systemic host response to severe infection and is manifested by varying degrees of hypotension, coagulopathy, and multiorgan dysfunction. Despite great efforts being made to understand this condition and designing therapies to treat sepsis, mortality rates are still high in septic patients. Characterization of the complex molecular signaling networks between the various components of host-pathogen interactions, highlights the difficulty in identifying a single driving force responsible for sepsis. Although triggering the inflammatory response is generally considered as protective against pathogenic threats, the interplay between the signaling pathways that are induced or suppressed during sepsis may harm the host. Numerous surveillance mechanisms have evolved to discriminate self from foreign agents and accordingly provoke an effective cellular response to target the pathogens. Nucleic acids are not only an essential genetic component, but sensing their molecular signature is also an important quality control mechanism which has evolved to maintain the integrity of the human genome. Evidence that has accumulated recently indicated that distinct pattern recognition receptors sense nucleic acids released from infectious organisms or from damaged host cells, resulting in the modulation of intracellular signalling cascades. Immunoreceptor-mediated detection of these nucleic acids induces antigen-specific immunity, secretion of proinflammatory cytokines and reactive oxygen/nitrogen species and thus are implicated in a range of diseases including septic shock. PMID:22328944
Syntheses and crystal structures of a series of new divalent metal phosphonates with imino-bis(methylphosphonic acid)

International Nuclear Information System (INIS)

Yang Bingping; Prosvirin, Andrey V.; Zhao, Han-Hua; Mao, Jiang-Gao

2006-01-01

Hydrothermal reactions of divalent transition metal salts with imino-bis(methylphosphonic acid), NH(CH 2 PO 3 H 2 ) 2 (H 4 L) afforded three new metal phosphonates, namely, Cu[NH(CH 2 PO 3 H) 2 ] 1, {Co[NH 2 (CH 2 PO 3 H)(CH 2 PO 3 )](H 2 O) 2 }.H 2 O 2 and Mn[NH 2 (CH 2 PO 3 H)(CH 2 PO 3 )](H 2 O) 3. When HO 2 C(CH 2 ) 3 N(CH 2 PO 3 H 2 ) 2 was used as the phosphonate ligand and 4,4'-bipy as the second metal linker, {Cu 4 [NH(CH 2 PO 3 ) 2 ] 2 (4,4'-bipy)(H 2 O) 4 }.9H 2 O 4 with a pillared layered architecture was obtained. The NH(CH 2 PO 3 ) 2 anion resulted from the cleavage of the HO 2 C(CH 2 ) 3 -group during the reaction. Although compounds 1-3 have a same M/L ratio (1:1), they exhibit totally different structures.Compound 1 has a linear chain structure, in which each pair of square-pyramidal coordinated copper(II) ions are bridged by two phosphonate oxygen atoms to form a Cu 2 O 2 dimeric unit, and such dimeric units are further interconnected via phosphonate groups to form a [010] chain. Compound 2 has a layered architecture built from CoO 6 octahedra bridged by phosphonate ligands. In compound 3, the interconnection of the manganese(II) ions by bridging imino-diphosphonate ligands leads to a 3D network. Compound 4 has a pillar-layered structure, the layers composed of Cu(II) ions bridged by aminodiphosphonate ligands are interconnected by 4,4'-bipy ligands to form channels along c-axis. Several factors that affect the structures of the metal phosphonates formed have also been discussed. Compounds 2 and 3 show predominant antiferromagnetic interactions between magnetic centers. -- Graphical abstract: Four new metal phosphonates, namely, Cu[NH(CH 2 PO 3 H) 2 ] 1, {Co[NH 2 (CH 2 PO 3 H)(CH 2 PO 3 )](H 2 O) 2 }.H 2 O 2, Mn[NH 2 (CH 2 PO 3 H)(CH 2 PO 3 )](H 2 O) 3 and {Cu 4 [NH(CH 2 PO 3 ) 2 ] 2 (4,4'-bipy)(H 2 O) 4 }.9H 2 O 4 have been synthesized and structurally characterized. Compound 1 has a linear chain structure, and compound 2 is layered. Compound 3 is
The isolation of nucleic acids from fixed, paraffin-embedded tissues-which methods are useful when?

DEFF Research Database (Denmark)

Gilbert, M Thomas P; Haselkorn, Tamara; Bunce, Michael

2007-01-01

. Cross-linking not only complicates isolation of nucleic acid but also introduces polymerase "blocks" during PCR. A wide variety of methods exists for the recovery of DNA and RNA from archival tissues, and although a number of previous studies have qualitatively compared the relative merits....... These include methods of pre-treating the samples prior to extraction, extraction and nucleic acid purification methods themselves, and a post-extraction enzymatic repair technique. We find that although many of the published methods have distinct positive effects on some characteristics of the nucleic acids...
Corrosion protective performance of amino trimethylene phosphonic acid-metal complex layers fabricated on the cold-rolled steel substrate via one-step assembly

Science.gov (United States)

Yan, Ru; He, Wei; Zhai, Tianhua; Ma, Houyi

2018-06-01

Seeing that amino trimethylene phosphonic acid (ATMP) possesses very strong complexation ability to metal ions and the phosphonic acid group has good affinity for the oxidized iron surface, herein a simple and rapid film-forming method (one-step assembly method) was developed to construct the ATMP-Zn complex conversion layers (ATMP-Zn layers for short) on the cold-rolled steel (CRS) substrate. Zinc ions were found to participate in the formation process of ATMP-based composite film, which made the Zn-containing ATMP film significantly different in appearance, thickness, microstructure and film-forming mechanisms from the Zn-free ATMP film. There was mainly iron (ш) phosphonate in the Zn-free ATMP film, whereas there were Zn2+-ATMP complex and a certain amount of ZnO in the ATMP-Zn composite film. In addition, electrochemical test results clearly indicate that corrosion resistance of ATMP-Zn composite film was greatly enhanced due to the presence of Zn component. Moreover, the corrosion resistance performance could be controlled by adjusting film-forming time, pH and ATMP concentration in the film-forming solutions. The present study provides a new method for the design and fabrication of high-quality environmentally-friendly conversion layers.
Styrene-divinylbenzene copolymer grafted with phosphonic acid dialkylesters

Directory of Open Access Journals (Sweden)

SMARANDA ILIESCU

2004-12-01

Full Text Available The functionalization of a crosslinked chloromethylated polystyrene 8% divinylbenzene copolymer with phosphonic ester groups is detailed. The reacton conditions were studied in order to determine the optimal conditions for obtaining only diesters. A statistical method for the calculation of the fraction of repetive units for the inited and final resin is proposed.
Lateral flow devices for nucleic acid analysis exploiting quantum dots as reporters

Energy Technology Data Exchange (ETDEWEB)

Sapountzi, Eleni A.; Tragoulias, Sotirios S.; Kalogianni, Despina P. [Department of Chemistry, University of Patras, GR-26504 Patras (Greece); Ioannou, Penelope C. [Department of Chemistry, University of Athens, GR-15771 Athens (Greece); Christopoulos, Theodore K., E-mail: tchrist@upatras.gr [Department of Chemistry, University of Patras, GR-26504 Patras (Greece); Institute of Chemical Engineering and High Temperature Processes, Foundation of Research and Technology Hellas, GR-26504 Patras (Greece)

2015-03-15

Highlights: • Dipstick tests for DNA hybridization assays and genotyping of single-nucleotide polymorphisms. • Use of quantum dots as reporters. • Visual detection without the need for expensive instrumentation. • Simplicity and low-cost of the assays. - Abstract: There is a growing interest in the development of biosensors in the form of simple lateral flow devices that enable visual detection of nucleic acid sequences while eliminating several steps required for pipetting, incubation and washing out the excess of reactants. In this work, we present the first dipstick-type nucleic acid biosensors based on quantum dots (QDs) as reporters. The biosensors enable sequence confirmation of the target DNA by hybridization and simple visual detection of the emitted fluorescence under a UV lamp. The ‘diagnostic’ membrane of the biosensor contains a test zone (TZ) and a control zone (CZ). The CZ always fluoresces in order to confirm the proper function of the biosensor. Fluorescence is emitted from the TZ, only when the specific nucleic acid sequence is present. We have developed two general types of QD-based nucleic acid biosensors, namely, Type I and Type II, in which the TZ consists of either immobilized streptavidin (Type I) or immobilized oligodeoxynucleotides (Type II). The control zone consists of immobilized biotinylated albumin. No purification steps are required prior to the application of the DNA sample on the strip. The QD-based nucleic acid biosensors performed accurately and reproducibly when applied to (a) the visual detection of PCR amplification products and (b) visual genotyping of single nucleotide polymorphisms (SNPs) in human genomic DNA from clinical samples. As low as 1.5 fmol of double-stranded DNA were clearly detected by naked eye and the dynamic range extended to 200 fmol. The %CV were estimated to be 4.3–8.2.
Conformationally constrained nucleoside phosphonic acids - potent inhibitors of human mitochondrial and cytosolic 5'(3')-nucleotidases

Czech Academy of Sciences Publication Activity Database

Šimák, Ondřej; Pachl, Petr; Fábry, Milan; Buděšínský, Miloš; Jandušík, T.; Hnízda, Aleš; Skleničková, Radka; Petrová, Magdalena; Veverka, Václav; Řezáčová, Pavlína; Brynda, Jiří; Rosenberg, Ivan

2014-01-01

Roč. 12, č. 40 (2014), s. 7971-7982 ISSN 1477-0520 R&D Projects: GA ČR GA203/09/0820; GA ČR GA13-24880S; GA ČR GA13-26526S; GA MŠk(CZ) LK11205; GA AV ČR KAN200520801 Institutional support: RVO:61388963 ; RVO:68378050 Keywords : 5'(3')-nucleotidase * structure * inhibition * cdN * mdN * nucleoside * SAR * phosphonic acid Subject RIV: CC - Organic Chemistry Impact factor: 3.562, year: 2014

The fragile X mental retardation protein has nucleic acid chaperone properties.

Science.gov (United States)

Gabus, Caroline; Mazroui, Rachid; Tremblay, Sandra; Khandjian, Edouard W; Darlix, Jean-Luc

2004-01-01

The fragile X syndrome is the most common cause of inherited mental retardation resulting from the absence of the fragile X mental retardation protein (FMRP). FMRP contains two K-homology (KH) domains and one RGG box that are landmarks characteristic of RNA-binding proteins. In agreement with this, FMRP associates with messenger ribonucleoparticles (mRNPs) within actively translating ribosomes, and is thought to regulate translation of target mRNAs, including its own transcript. To investigate whether FMRP might chaperone nucleic acid folding and hybridization, we analysed the annealing and strand exchange activities of DNA oligonucleotides and the enhancement of ribozyme-directed RNA substrate cleavage by FMRP and deleted variants relative to canonical nucleic acid chaperones, such as the cellular YB-1/p50 protein and the retroviral nucleocapsid protein HIV-1 NCp7. FMRP was found to possess all the properties of a potent nucleic acid chaperone, requiring the KH motifs and RGG box for optimal activity. These findings suggest that FMRP may regulate translation by acting on RNA-RNA interactions and thus on the structural status of mRNAs.
Velocity and processivity of helicase unwinding of double-stranded nucleic acids

International Nuclear Information System (INIS)

Betterton, M D; Juelicher, F

2005-01-01

Helicases are molecular motors which unwind double-stranded nucleic acids (dsNA) in cells. Many helicases move with directional bias on single-stranded (ss) nucleic acids, and couple their directional translocation to strand separation. A model of the coupling between translocation and unwinding uses an interaction potential to represent passive and active helicase mechanisms. A passive helicase must wait for thermal fluctuations to open dsNA base pairs before it can advance and inhibit NA closing. An active helicase directly destabilizes dsNA base pairs, accelerating the opening rate. Here we extend this model to include helicase unbinding from the nucleic-acid strand. The helicase processivity depends on the form of the interaction potential. A passive helicase has a mean attachment time which does not change between ss translocation and ds unwinding, while an active helicase in general shows a decrease in attachment time during unwinding relative to ss translocation. In addition, we describe how helicase unwinding velocity and processivity vary if the base-pair binding free energy is changed
Operating Cooperatively (OC sensor for highly specific recognition of nucleic acids.

Directory of Open Access Journals (Sweden)

Evan M Cornett

Full Text Available Molecular Beacon (MB probes have been extensively used for nucleic acid analysis because of their ability to produce fluorescent signal in solution instantly after hybridization. The indirect binding of MB probe to a target analyte offers several advantages, including: improved genotyping accuracy and the possibility to analyse folded nucleic acids. Here we report on a new design for MB-based sensor, called 'Operating Cooperatively' (OC, which takes advantage of indirect binding of MB probe to a target analyte. The sensor consists of two unmodified DNA strands, which hybridize to a universal MB probe and a nucleic acid analyte to form a fluorescent complex. OC sensors were designed to analyze two human SNPs and E. coli 16S rRNA. High specificity of the approach was demonstrated by the detection of true analyte in over 100 times excess amount of single base substituted analytes. Taking into account the flexibility in the design and the simplicity in optimization, we conclude that OC sensors may become versatile and efficient tools for instant DNA and RNA analysis in homogeneous solution.
Determination of perfluoroalkyl carboxylic, sulfonic, and phosphonic acids in food.

Science.gov (United States)

Ullah, Shahid; Alsberg, Tomas; Vestergren, Robin; Berger, Urs

2012-11-01

A sensitive and accurate method was developed and validated for simultaneous analysis of perfluoroalkyl carboxylic acids, sulfonic acids, and phosphonic acids (PFPAs) at low picograms per gram concentrations in a variety of food matrices. The method employed extraction with acetonitrile/water and cleanup on a mixed-mode co-polymeric sorbent (C8 + quaternary amine) using solid-phase extraction. High-performance liquid chromatographic separation was achieved on a C18 column using a mobile phase gradient containing 5 mM 1-methyl piperidine for optimal chromatographic resolution of PFPAs. A quadrupole time-of-flight high-resolution mass spectrometer operating in negative ion mode was used as detector. Method detection limits were in the range of 0.002 to 0.02 ng g(-1) for all analytes. Sample preparation (extraction and cleanup) recoveries at a spiking level of 0.1 ng g(-1) to a baby food composite were in the range of 59 to 98 %. A strong matrix effect was observed in the analysis of PFPAs in food extracts, which was tentatively assigned to sorption of PFPAs to the injection vial in the solvent-based calibration standard. The method was successfully applied to a range of different food matrices including duplicate diet samples, vegetables, meat, and fish samples.
Chimeric peptide beacons: a direct polypeptide analog of DNA molecular beacons†

OpenAIRE

Oh, Kenneth J.; Cash, Kevin J.; Lubin, Arica A.; Plaxco, Kevin W.

2007-01-01

We have developed a new biosensor architecture, which is comprised of a polypeptide–peptide nucleic acid tri-block copolymer and which we have termed chimeric peptide beacons (CPB), that generates an optical output via a mechanism analogous to that employed in DNA-based molecular beacons.
Circulating nucleic acids as a new diagnostic tool

Czech Academy of Sciences Publication Activity Database

Urbanová, Markéta; Plzák, J.; Strnad, Hynek; Betka, J.

2010-01-01

Roč. 15, č. 2 (2010), s. 242-259 ISSN 1425-8153 R&D Projects: GA MŠk 2B06106 Institutional research plan: CEZ:AV0Z50520514 Keywords : circulating nucleic acids * diagnostics * cancer Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 1.455, year: 2010
PYRENE INTERCALATING NUCLEIC ACIDS WITH A CARBON LINKER

DEFF Research Database (Denmark)

Østergaard, Michael E.; Wamberg, Michael Chr.; Pedersen, Erik Bjerregaard

2011-01-01

geminally attached. Fluorescence studies of this intercalating nucleic acid with the pyrene moieties inserted as a bulge showed formation of an excimer band. When a mismatch was introduced at the site of the intercalator, an excimer band was formed for the destabilized duplexes whereas an exciplex band...
Optical nano-biosensing interface via nucleic acid amplification strategy: construction and application.

Science.gov (United States)

Zhou, Hong; Liu, Jing; Xu, Jing-Juan; Zhang, Shu-Sheng; Chen, Hong-Yuan

2018-03-21

Modern optical detection technology plays a critical role in current clinical detection due to its high sensitivity and accuracy. However, higher requirements such as extremely high detection sensitivity have been put forward due to the clinical needs for the early finding and diagnosing of malignant tumors which are significant for tumor therapy. The technology of isothermal amplification with nucleic acids opens up avenues for meeting this requirement. Recent reports have shown that a nucleic acid amplification-assisted modern optical sensing interface has achieved satisfactory sensitivity and accuracy, high speed and specificity. Compared with isothermal amplification technology designed to work completely in a solution system, solid biosensing interfaces demonstrated better performances in stability and sensitivity due to their ease of separation from the reaction mixture and the better signal transduction on these optical nano-biosensing interfaces. Also the flexibility and designability during the construction of these nano-biosensing interfaces provided a promising research topic for the ultrasensitive detection of cancer diseases. In this review, we describe the construction of the burgeoning number of optical nano-biosensing interfaces assisted by a nucleic acid amplification strategy, and provide insightful views on: (1) approaches to the smart fabrication of an optical nano-biosensing interface, (2) biosensing mechanisms via the nucleic acid amplification method, (3) the newest strategies and future perspectives.
Phosphonate-hydroxyapatite hybrid compounds prepared by hydrothermal method

Energy Technology Data Exchange (ETDEWEB)

Agougui, H.; Aissa, A. [Laboratoire de Physico-Chimie des Materiaux, Faculte des sciences de Monastir, 5019 Monastir (Tunisia); Maggi, S. [CNR-IC Istituto di Cristallografia, via Amendola 122/O, 70126 Bari (Italy); Debbabi, M., E-mail: m.debbabi@yahoo.fr [Laboratoire de Physico-Chimie des Materiaux, Faculte des sciences de Monastir, 5019 Monastir (Tunisia)

2010-12-15

Calcium hydroxyapatite (CaHAp) was prepared in the presence of two alkylphosphonates, the tert-butyl phosphonic acid TBPOH and the 2-carboxyletylphosphonic acid 2-CEPA, by hydrothermal method at 120 deg. C for 15 h. The modification of hydroxyapatite by grafting organic moieties is confirmed by IR and NMR MAS ({sup 1}H and {sup 31}P) spectroscopy and chemical analysis. X-ray powder diffraction patterns show that the incorporation of organic moieties induces a significant loss of the material crystallinity and a clear increase of the unit cell lattice parameter a as function of 2-CEPA grafting rate. The specific surface area (SSA) increases with increasing phosphonate amount especially for 2-CEPA. All techniques show the lower reactivity of TBPOH due to the steric effects of tert-butyl, whereas the 2-CEPA with a linear chain and double acidic functions is more reactive and can replace the OH{sup -} groups of the apatitic structure.
Phosphonate-hydroxyapatite hybrid compounds prepared by hydrothermal method

International Nuclear Information System (INIS)

Agougui, H.; Aissa, A.; Maggi, S.; Debbabi, M.

2010-01-01

Calcium hydroxyapatite (CaHAp) was prepared in the presence of two alkylphosphonates, the tert-butyl phosphonic acid TBPOH and the 2-carboxyletylphosphonic acid 2-CEPA, by hydrothermal method at 120 deg. C for 15 h. The modification of hydroxyapatite by grafting organic moieties is confirmed by IR and NMR MAS ( 1 H and 31 P) spectroscopy and chemical analysis. X-ray powder diffraction patterns show that the incorporation of organic moieties induces a significant loss of the material crystallinity and a clear increase of the unit cell lattice parameter a as function of 2-CEPA grafting rate. The specific surface area (SSA) increases with increasing phosphonate amount especially for 2-CEPA. All techniques show the lower reactivity of TBPOH due to the steric effects of tert-butyl, whereas the 2-CEPA with a linear chain and double acidic functions is more reactive and can replace the OH - groups of the apatitic structure.
Devices, systems, and methods for detecting nucleic acids using sedimentation

Energy Technology Data Exchange (ETDEWEB)

Koh, Chung-Yan; Schaff, Ulrich Y.; Sommer, Gregory J.

2017-10-24

Embodiments of the present invention are directed toward devices, systems, and method for conducting nucleic acid purification and quantification using sedimentation. In one example, a method includes generating complexes which bind to a plurality of beads in a fluid sample, individual ones of the complexes comprising a nucleic acid molecule such as DNA or RNA and a labeling agent. The plurality of beads including the complexes may be transported through a density media, wherein the density media has a density lower than a density of the beads and higher than a density of the fluid sample, and wherein the transporting occurs, at least in part, by sedimentation. Signal may be detected from the labeling agents of the complexes.
Preparation of polyvinyl alcohol graphene oxide phosphonate film and research of thermal stability and mechanical properties.

Science.gov (United States)

Li, Jihui; Song, Yunna; Ma, Zheng; Li, Ning; Niu, Shuai; Li, Yongshen

2018-05-01

In this article, flake graphite, nitric acid, peroxyacetic acid and phosphoric acid are used to prepare graphene oxide phosphonic and phosphinic acids (GOPAs), and GOPAs and polyvinyl alcohol (PVA) are used to synthesize polyvinyl alcohol graphene oxide phosphonate and phosphinate (PVAGOPs) in the case of faint acidity and ultrasound irradiation, and PVAGOPs are used to fabricate PVAGOPs film, and the structure and morphology of GOPAs, PVAGOPs and PVAGOPs film are characterized, and the thermal stability and mechanical properties of PVAGOPs film are investigated. Based on these, it has been proved that GOPAs consist of graphene oxide phosphonic acid and graphene oxide phosphinic acid, and there are CP covalent bonds between them, and PVAGOPs are composed of GOPAs and PVA, and there are six-member lactone rings between GOPAs and PVA, and the thermal stability and mechanical properties of PVAGOPs film are improved effectively. Copyright © 2018 Elsevier B.V. All rights reserved.
Circulating nucleic acids and evolution.

Science.gov (United States)

Anker, Philippe; Stroun, Maurice

2012-06-01

J.B. Lamarck in 1809 was the first to present a theory of evolution. He proposed it was due to the adaptation of species to environmental changes, this adaptation being acquired by the offspring. In 1868, Darwin suggested that cells excrete gemmules, which circulate through the body and reach the gonads where they are transmitted to the next generation. His main argument came from graft hybrids. In the fifties and sixties, Russian geneticists, rejecting neo-Darwinism, said that acquired characteristics were the basis of evolution. The main experiments on which they based their theory were the transmission of hereditary characteristics by a special technique of grafting between two varieties of plants. We repeated this kind of experiment and also succeeded in obtaining hereditary modifications of the pupil plants that acquired some characteristics of the mentor variety. Rather than adopting the views of the Russian scientists, we suggested that DNA was circulating between the mentor and pupil plants. Hirata's group have shown recently, by using molecular techniques such as cloning, RFLP PCR and sequencing some genes of their graft hybrids of pepper plants, that transfer of informative molecules from the mentor to the pupil plant does exist. Nucleic acids are actively released by cells; they circulate in the body. They can transform oncogenically or trigger antibody response but the only genetic transformation showing that DNA can go from the soma to the germen comes from graft hybrids. This suggests that circulating nucleic acids, in this case DNA, like Darwin's gemmules, play a role in the mechanism of evolution.
NaVirCept - Nucleic Acid-Based Anti-Viral Project

International Nuclear Information System (INIS)

Stephen, E. R.; Wong, J.; Van Loon, D.

2007-01-01

Vaccines are generally considered to be the most effective countermeasures to bacterial and viral diseases, however, licensed vaccines against many disease agents are either not available or their efficacies have not been demonstrated. Vaccines are generally agent specific in terms of treatment spectrum and are subject to defeat through natural mutation or through directed efforts. With respect to viral therapeutics, one of the major limitations associated with antiviral drugs is acquired drug resistance caused by antigenic shift or drift. A number of next-generation prophylactic and/or therapeutic measures are on the horizon. Of these, nucleic acid-based drugs are showing great antiviral potential. These drugs elicit long-lasting, broad spectrum protective immune responses, especially to respiratory viral pathogens. The Nucleic Acid-Based Antiviral (NaVirCept) project provides the opportunity to demonstrate the effectiveness of novel medical countermeasures against military-significant endemic and other viral threat agents. This project expands existing DRDC drug delivery capability development, in the form of proprietary liposome intellectual property, by coupling it with leading-edge nucleic acid-based technology to deliver effective medical countermeasures that will protect deployed personnel and the warfighter against a spectrum of viral disease agents. The technology pathway will offer a means to combat emerging viral diseases or modified threat agents such as the bird flu or reconstructed Spanish flu without going down the laborious, time-consuming and expensive paths to develop countermeasures for each new and/or emerging viral disease organism.(author)
Assays for urinary biomarkers of oxidatively damaged nucleic acids

DEFF Research Database (Denmark)

Weimann, Allan; Broedbaek, Kasper; Henriksen, Trine

2012-01-01

Abstract The analysis of oxidized nucleic acid metabolites can be performed by a variety of methodologies: liquid chromatography coupled with electrochemical or mass-spectrometry detection, gas chromatography coupled with mass spectrometry, capillary electrophoresis and ELISA (Enzyme-linked immun...
Pharmacokinetic Profiling of Conjugated Therapeutic Oligonucleotides: A High-Throughput Method Based Upon Serial Blood Microsampling Coupled to Peptide Nucleic Acid Hybridization Assay.

Science.gov (United States)

Godinho, Bruno M D C; Gilbert, James W; Haraszti, Reka A; Coles, Andrew H; Biscans, Annabelle; Roux, Loic; Nikan, Mehran; Echeverria, Dimas; Hassler, Matthew; Khvorova, Anastasia

2017-12-01

Therapeutic oligonucleotides, such as small interfering RNAs (siRNAs), hold great promise for the treatment of incurable genetically defined disorders by targeting cognate toxic gene products for degradation. To achieve meaningful tissue distribution and efficacy in vivo, siRNAs must be conjugated or formulated. Clear understanding of the pharmacokinetic (PK)/pharmacodynamic behavior of these compounds is necessary to optimize and characterize the performance of therapeutic oligonucleotides in vivo. In this study, we describe a simple and reproducible methodology for the evaluation of in vivo blood/plasma PK profiles and tissue distribution of oligonucleotides. The method is based on serial blood microsampling from the saphenous vein, coupled to peptide nucleic acid hybridization assay for quantification of guide strands. Performed with minimal number of animals, this method allowed unequivocal detection and sensitive quantification without the need for amplification, or further modification of the oligonucleotides. Using this methodology, we compared plasma clearances and tissue distribution profiles of two different hydrophobically modified siRNAs (hsiRNAs). Notably, cholesterol-hsiRNA presented slow plasma clearances and mainly accumulated in the liver, whereas, phosphocholine-docosahexaenoic acid-hsiRNA was rapidly cleared from the plasma and preferably accumulated in the kidney. These data suggest that the PK/biodistribution profiles of modified hsiRNAs are determined by the chemical nature of the conjugate. Importantly, the method described in this study constitutes a simple platform to conduct pilot assessments of the basic clearance and tissue distribution profiles, which can be broadly applied for evaluation of new chemical variants of siRNAs and micro-RNAs.
Label-free direct surface-enhanced Raman scattering (SERS) of nucleic acids (Conference Presentation)

Science.gov (United States)

Guerrini, Luca; Morla-Folch, Judit; Gisbert-Quilis, Patricia; Xie, Hainan; Alvarez-Puebla, Ramon

2016-03-01

Recently, plasmonic-based biosensing has experienced an unprecedented level of attention, with a particular focus on the nucleic acid detection, offering efficient solutions to engineer simple, fast, highly sensitive sensing platforms while overcoming important limitations of PCR and microarray techniques. In the broad field of plasmonics, surface-enhanced Raman scattering (SERS) spectroscopy has arisen as a powerful analytical tool for detection and structural characterization of biomolecules. Today applications of SERS to nucleic acid analysis largely rely on indirect strategies, which have been demonstrated very effective for pure sensing purposes but completely dismiss the exquisite structural information provided by the direct acquisition of the biomolecular vibrational fingerprint. Contrarily, direct label-free SERS of nucleic acid shows an outstanding potential in terms of chemical-specific information which, however, remained largely unexpressed mainly because of the inherent poor spectral reproducibility and/or limited sensitivity. To address these limitations, we developed a fast and affordable high-throughput screening direct SERS method for gaining detailed genomic information on nucleic acids (DNA and RNA) and for the characterization and quantitative recognition of DNA interactions with exogenous agents. The simple strategy relies on the electrostatic adhesion of DNA/RNA onto positively-charged silver colloids that promotes the nanoparticle aggregation into stable clusters yielding intense and reproducible SERS spectra at picogram level (i.e. the analysis can be performed without the necessity of amplification steps thus providing realistic direct information of the nucleic acid in its native state). We anticipate this method to gain a vast impact and set of applications in different fields, including medical diagnostics, genomic screening, drug discovery, forensic science and even molecular electronics.
Content and synthesis of nucleic acids in the cartilage in chondromalacia patellae.

Science.gov (United States)

Lund, F; Telhag, H

1978-12-01

The content and the synthesis of nucleic acids in chondromalacian, osteoarthritis and normal cartilage was compared. The chondromalacian cartilage differed from osteoarthritis in that the content of nucleic acids was less. Also, the cell density was less in chondromalacian than in normal cartilage as opposed to previous findings in osteoarthritis. The synthesis of DNA was greater in chondromalacian than in normal cartilage but less than in osteoarthritis. With regard to the RNA synthesis, however, the chondromalacian cartilage showed a higher rate than both normal and osteoarthritic cartilage.
The inhibition of crystal growth of mirabilite in aqueous solutions in the presence of phosphonates

Science.gov (United States)

Vavouraki, A. I.; Koutsoukos, P. G.

2016-02-01

The formation of sodium sulfate decahydrate (Mirabilite) has been known to cause serious damages to structural materials both of modern and of historical buildings. Methods which can retard or completely suppress the development of mirabilte crystals are urgently needed especially as remedies or preventive measures for the preservation of the built cultural heritage. In the present work we present results on the effect of the presence of phosphonate compounds on the kinetics of crystal growth from aqueous supersaturated solutions at 18 °C using the seeded growth technique. The phosphonate compounds tested differed with respect to the number of ionizable phosphonate groups and with respect to the number of amino groups in the respective molecules. The crystal growth process was monitored by the temperature changes during the exothermic crystallization of mirabilite in the stirred supersaturated solutions. The crystal growth of mirabilite in the presence of: (1-hydroxyethylidene)-1, 1-diphosphonic acid (HEDP), amino tri (methylene phosphonic acid) (ATMP), hexamethylenediaminetetra (methylene)phosphonic acid (HTDMP), and diethylene triamine penta(methylene phosphonic acid)(DETPMP) over a range of concentrations between 0.1-5% w/w resulted in significant decrease of the rates of mirabilite crystal growth. All phosphonic compounds tested reduced the crystallization rates up to 60% in comparison with additive-free solutions. The presence of the test compounds did not cause changes of the mechanism of crystal growth which was surface diffusion controlled, as shown by the second order dependence of the rates of mirabilite crystal growth on the relative supersaturation. The excellent fit of the measured rates to a kinetic Langmuir-type model suggested that the activity of the tested inhibitors could be attributed to the adsorption and subsequent reduction of the active crystal growth sites of the seed crystals. In all cases, the inhibitory activity was reduced with
Fluorescence In Situ Hybridization with Peptide Nucleic Acid Probes for Rapid Identification of Candida albicans Directly from Blood Culture Bottles

Science.gov (United States)

Rigby, Susan; Procop, Gary W.; Haase, Gerhard; Wilson, Deborah; Hall, Geraldine; Kurtzman, Cletus; Oliveira, Kenneth; Von Oy, Sabina; Hyldig-Nielsen, Jens J.; Coull, James; Stender, Henrik

2002-01-01

A new fluorescence in situ hybridization (FISH) method that uses peptide nucleic acid (PNA) probes for identification of Candida albicans directly from positive-blood-culture bottles in which yeast was observed by Gram staining (herein referred to as yeast-positive blood culture bottles) is described. The test (the C. albicans PNA FISH method) is based on a fluorescein-labeled PNA probe that targets C. albicans 26S rRNA. The PNA probe is added to smears made directly from the contents of the blood culture bottle and hybridized for 90 min at 55°C. Unhybridized PNA probe is removed by washing of the mixture (30 min), and the smears are examined by fluorescence microscopy. The specificity of the method was confirmed with 23 reference strains representing phylogenetically related yeast species and 148 clinical isolates covering the clinically most significant yeast species, including C. albicans (n = 72), C. dubliniensis (n = 58), C. glabrata (n = 5), C. krusei (n = 2), C. parapsilosis (n = 4), and C. tropicalis (n = 3). The performance of the C. albicans PNA FISH method as a diagnostic test was evaluated with 33 routine and 25 simulated yeast-positive blood culture bottles and showed 100% sensitivity and 100% specificity. It is concluded that this 2.5-h method for the definitive identification of C. albicans directly from yeast-positive blood culture bottles provides important information for optimal antifungal therapy and patient management. PMID:12037084

Peptide nucleic acid probe-based fluorescence melting curve analysis for rapid screening of common JAK2, MPL, and CALR mutations.

Science.gov (United States)

Park, Joonhong; Song, Minsik; Jang, Woori; Chae, Hyojin; Lee, Gun Dong; Kim, KyungTak; Park, Heekyung; Kim, Myungshin; Kim, Yonggoo

2017-02-01

We developed and evaluated the feasibility of peptide nucleic acid (PNA)-based fluorescence melting curve analysis (FMCA) to detect common mutations in myeloproliferative neoplasms (MPNs). We have set up two separate reactions of PNA-based FMCA: JAK2 V617F &CALR p.Leu367fs*46 (set A) and MPL W515L/K &CALR p.Lys385fs*47 (set B). Clinical usefulness was validated with allele-specific real-time PCR, fragment analysis, Sanger sequencing in 57 BCR-ABL1-negative MPNs. The limit of detection (LOD) of PNA-based FMCA was approximately 10% for each mutation and interference reactions using mixtures of different mutations were not observed. Non-specific amplification was not observed in normal control. PNA-based FMCA was able to detect all JAK2 V617F (n=20), CALR p.Leu367fs*46 (n=10) and p.Lys385fs*47 (n=8). Three of six MPL mutations were detected except three samples with low mutant concentration in out of LOD. JAK2 exon 12 mutations (n=7) were negative without influencing V617F results. Among six variant CALR exon 9 mutations, two were detected by this method owing to invading of probe binding site. PNA-based FMCA for detecting common JAK2, MPL, and CALR mutations is a rapid, simple, and sensitive technique in BCR-ABL1-negative MPNs with >10% mutant allele at the time of initial diagnosis. Copyright © 2016 Elsevier B.V. All rights reserved.
A novel automated device for rapid nucleic acid extraction utilizing a zigzag motion of magnetic silica beads

International Nuclear Information System (INIS)

Yamaguchi, Akemi; Matsuda, Kazuyuki; Uehara, Masayuki; Honda, Takayuki; Saito, Yasunori

2016-01-01

We report a novel automated device for nucleic acid extraction, which consists of a mechanical control system and a disposable cassette. The cassette is composed of a bottle, a capillary tube, and a chamber. After sample injection in the bottle, the sample is lysed, and nucleic acids are adsorbed on the surface of magnetic silica beads. These magnetic beads are transported and are vibrated through the washing reagents in the capillary tube under the control of the mechanical control system, and thus, the nucleic acid is purified without centrifugation. The purified nucleic acid is automatically extracted in 3 min for the polymerase chain reaction (PCR). The nucleic acid extraction is dependent on the transport speed and the vibration frequency of the magnetic beads, and optimizing these two parameters provided better PCR efficiency than the conventional manual procedure. There was no difference between the detection limits of our novel device and that of the conventional manual procedure. We have already developed the droplet-PCR machine, which can amplify and detect specific nucleic acids rapidly and automatically. Connecting the droplet-PCR machine to our novel automated extraction device enables PCR analysis within 15 min, and this system can be made available as a point-of-care testing in clinics as well as general hospitals. - Highlights: • Automatic nucleic acid extraction is performed in 3 min. • Zigzag motion of magnetic silica beads yields rapid and efficient extraction. • The present our device provides better performance than the conventional procedure.
Multiplexed Detection of Attomoles of Nucleic Acids Using Fluorescent Nanoparticle Counting Platform.

Science.gov (United States)

Pei, Xiaojing; Yin, Haoyan; Lai, Tiancheng; Zhang, Junlong; Liu, Feng; Xu, Xiao; Li, Na

2018-01-16

The sensitive multiplexed detection of nucleic acids in a single sample by a simple manner is of pivotal importance for the diagnosis and therapy of human diseases. Herein, we constructed an automatic fluorescent nanoparticle (FNP) counting platform with a common fluorescence microscopic imaging setup for nonamplification multiplexed detection of attomoles of nucleic acids. Taking the advantages of the highly bright, multicolor emitting FNPs and magnetic separation, the platform enables sensitive multiplexed detection without the need for extra fluorescent labels. Quantification for multiplex DNAs, multiplex microRNAs (miRNA), as well as a DNA and miRNA mixture was achieved with a similar dynamic range, a limit of detection down to 5 amol (5 μL detection volume), and a 81-115% spike recovery from different biological sample matrices. In particular, the sensitivity for multiplex miRNA is by far among the highest without using amplification or the lock nucleic acid hybridization enhancement strategy. Results regarding miRNA-141 from four different cell lines were agreeable with those of the quantitative reverse transcription polymerase chain reaction. Simultaneous detection of miRNA-141 and miRNA-21 in four different cell lines yielded consistent results with publications, indicating the potential for monitoring multiplex miRNA expression associated with the collaborative regulation of important cellular events. This work expands the rule set of multiplex nucleic acid detection strategies and shows promising potential application in clinical diagnosis.
Surface plasmon resonance sensing of nucleic acids: A review

Czech Academy of Sciences Publication Activity Database

Šípová, Hana; Homola, Jiří

-, č. 773 (2013), s. 9-23 ISSN 0003-2670 R&D Projects: GA MŠk(CZ) LH11102 Institutional support: RVO:67985882 Keywords : Surface plasmon resonance * Nucleic acid * Biosensor Subject RIV: JB - Sensors, Measurment, Regulation Impact factor: 4.517, year: 2013
Bio-Orthogonal Mediated Nucleic Acid Transfection of Cells via Cell Surface Engineering.

Science.gov (United States)

O'Brien, Paul J; Elahipanah, Sina; Rogozhnikov, Dmitry; Yousaf, Muhammad N

2017-05-24

The efficient delivery of foreign nucleic acids (transfection) into cells is a critical tool for fundamental biomedical research and a pillar of several biotechnology industries. There are currently three main strategies for transfection including reagent, instrument, and viral based methods. Each technology has significantly advanced cell transfection; however, reagent based methods have captured the majority of the transfection market due to their relatively low cost and ease of use. This general method relies on the efficient packaging of a reagent with nucleic acids to form a stable complex that is subsequently associated and delivered to cells via nonspecific electrostatic targeting. Reagent transfection methods generally use various polyamine cationic type molecules to condense with negatively charged nucleic acids into a highly positively charged complex, which is subsequently delivered to negatively charged cells in culture for association, internalization, release, and expression. Although this appears to be a straightforward procedure, there are several major issues including toxicity, low efficiency, sorting of viable transfected from nontransfected cells, and limited scope of transfectable cell types. Herein, we report a new strategy (SnapFect) for nucleic acid transfection to cells that does not rely on electrostatic interactions but instead uses an integrated approach combining bio-orthogonal liposome fusion, click chemistry, and cell surface engineering. We show that a target cell population is rapidly and efficiently engineered to present a bio-orthogonal functional group on its cell surface through nanoparticle liposome delivery and fusion. A complementary bio-orthogonal nucleic acid complex is then formed and delivered to which chemoselective click chemistry induced transfection occurs to the primed cell. This new strategy requires minimal time, steps, and reagents and leads to superior transfection results for a broad range of cell types
Origin of nucleic acids

International Nuclear Information System (INIS)

Prieur, B.E.

1995-01-01

The appearance of nucleic acids is the first event after the birth of membranes which made it possible to assure the perenniality of information. The complexity of these molecules has led some scientists to propose that they were not prebiotic but rather derived a more simple and achiral primitive ancestor. This hypothesis suggests that ribose possesses properties that allowed the formation of certain polysaccharides which evolved to RNA. The first step of the hypothesis is the selection and concentration of ribofuranose. This sugar has chelating properties and its alpha-ribofuranose is favoured in the chelating position. The density of the sugar with a heavy cation is greater than water and thus the complex can escape the UV radiation at the surface of the ocean. The particularity of ribose is to be able to form a homochiral regular array of these basic chelating structures with pyrophosphite. These arrays evolve towards the formation of polysaccharides (poly ribose phosphate) which have a very organized structure. These polysaccharides in turn evolve to RNA by binding of adenine and deoxyguanine which are HCN derivatives that can react with the polysaccharides. The primitive RNA is methylated and oxidized to form prebiotic RNA with adenosine, cytidine, 7methyl-guanosine and ribothymidine as nucleic bases. The pathway of biosynthesis of DNA form RNA will be studied. I suggest that the appearance of DNA results form the interaction between prebiotic double stranded RNA and proteins. DNA could be a product of RNA degradation by proteins. The catabolism of RNA to DNA requires a source of free radicals, protons and hydrides. RNA cannot produce free radicals, which are provided by the phenol group of the amino acid tyrosien. Protons are provided by the medium and hydrides are provided by 7-methyl-guanosine which can fix hydrides coming from hydrogen gas and donate them for the transformation of a riboside to a deoxyriboside. This pathway suggests that DNA appeared at
Molecular modeling of nucleic Acid structure: electrostatics and solvation.

Science.gov (United States)

Bergonzo, Christina; Galindo-Murillo, Rodrigo; Cheatham, Thomas E

2014-12-19

This unit presents an overview of computer simulation techniques as applied to nucleic acid systems, ranging from simple in vacuo molecular modeling techniques to more complete all-atom molecular dynamics treatments that include an explicit representation of the environment. The third in a series of four units, this unit focuses on critical issues in solvation and the treatment of electrostatics. UNITS 7.5 & 7.8 introduced the modeling of nucleic acid structure at the molecular level. This included a discussion of how to generate an initial model, how to evaluate the utility or reliability of a given model, and ultimately how to manipulate this model to better understand its structure, dynamics, and interactions. Subject to an appropriate representation of the energy, such as a specifically parameterized empirical force field, the techniques of minimization and Monte Carlo simulation, as well as molecular dynamics (MD) methods, were introduced as a way of sampling conformational space for a better understanding of the relevance of a given model. This discussion highlighted the major limitations with modeling in general. When sampling conformational space effectively, difficult issues are encountered, such as multiple minima or conformational sampling problems, and accurately representing the underlying energy of interaction. In order to provide a realistic model of the underlying energetics for nucleic acids in their native environments, it is crucial to include some representation of solvation (by water) and also to properly treat the electrostatic interactions. These subjects are discussed in detail in this unit. Copyright © 2014 John Wiley & Sons, Inc.
PrPC has nucleic acid chaperoning properties similar to the nucleocapsid protein of HIV-1.

Science.gov (United States)

Derrington, Edmund; Gabus, Caroline; Leblanc, Pascal; Chnaidermann, Jonas; Grave, Linda; Dormont, Dominique; Swietnicki, Wieslaw; Morillas, Manuel; Marck, Daniel; Nandi, Pradip; Darlix, Jean-Luc

2002-01-01

The function of the cellular prion protein (PrPC) remains obscure. Studies suggest that PrPC functions in several processes including signal transduction and Cu2+ metabolism. PrPC has also been established to bind nucleic acids. Therefore we investigated the properties of PrPC as a putative nucleic acid chaperone. Surprisingly, PrPC possesses all the nucleic acid chaperoning properties previously specific to retroviral nucleocapsid proteins. PrPC appears to be a molecular mimic of NCP7, the nucleocapsid protein of HIV-1. Thus PrPC, like NCP7, chaperones the annealing of tRNA(Lys) to the HIV-1 primer binding site, the initial step of retrovirus replication. PrPC also chaperones the two DNA strand transfers required for production of a complete proviral DNA with LTRs. Concerning the functions of NCP7 during budding, PrPC also mimices NCP7 by dimerizing the HIV-1 genomic RNA. These data are unprecedented because, although many cellular proteins have been identified as nucleic acid chaperones, none have the properties of retroviral nucleocapsid proteins.
Vibrational spectroscopy and principal component analysis for conformational study of virus nucleic acids

Science.gov (United States)

Dovbeshko, G. I.; Repnytska, O. P.; Pererva, T.; Miruta, A.; Kosenkov, D.

2004-07-01

Conformation analysis of mutated DNA-bacteriophages (PLys-23, P23-2, P47- the numbers have been assigned by T. Pererva) induced by MS2 virus incorporated in Ecoli AB 259 Hfr 3000 has been done. Surface enhanced infrared absorption (SEIRA) spectroscopy and principal component analysis has been applied for solving this problem. The nucleic acids isolated from the mutated phages had a form of double stranded DNA with different modifications. The nucleic acid from phage P47 was undergone the structural rearrangement in the most degree. The shape and position ofthe fine structure of the Phosphate asymmetrical band at 1071cm-1 as well as the stretching OH vibration at 3370-3390 cm-1 has indicated to the appearance ofadditional OH-groups. The Z-form feature has been found in the base vibration region (1694 cm-1) and the sugar region (932 cm-1). A supposition about modification of structure of DNA by Z-fragments for P47 phage has been proposed. The P23-2 and PLys-23 phages have showed the numerous minor structural changes also. On the basis of SEIRA spectra we have determined the characteristic parameters of the marker bands of nucleic acid used for construction of principal components. Contribution of different spectral parameters of nucleic acids to principal components has been estimated.
Adsorption of nucleic acid bases and amino acids on single-walled carbon and boron nitride nanotubes: a first-principles study.

Science.gov (United States)

Zheng, Jiaxin; Song, Wei; Wang, Lu; Lu, Jing; Luo, Guangfu; Zhou, Jing; Qin, Rui; Li, Hong; Gao, Zhengxiang; Lai, Lin; Li, Guangping; Mei, Wai Ning

2009-11-01

We study the adsorptions of nucleic acid bases adenine (A), cytosine (C), guanine (G), thymine (T), and uracil (U) and four amino acids phenylalanine, tyrosine, tryptophan, alanine on the single-walled carbon nanotubes (SWCNTs) and boron nitride nanotubes (SWBNNTs) by using density functional theory. We find that the aromatic content plays a critical role in the adsorption. The adsorptions of nucleic acid bases and amino acids on the (7, 7) SWBNNT are stronger than those on the (7, 7) SWCNT. Oxidative treatment of SWCNTs favors the adsorption of biomolecules on nanotubes.
Ultramild protein-mediated click chemistry creates efficient oligonucleotide probes for targeting and detecting nucleic acids

DEFF Research Database (Denmark)

Nåbo, Lina J.; Madsen, Charlotte S.; Jensen, Knud J.

2015-01-01

Functionalized synthetic oligonucleotides are finding growing applications in research, clinical studies, and therapy. However, it is not easy to prepare them in a biocompatible and highly efficient manner. We report a new strategy to synthesize oligonucleotides with promising nucleic acid...... targeting and detection properties. We focus in particular on the pH sensitivity of these new probes and their high target specificity. For the first time, human copper(I)-binding chaperon Cox17 was applied to effectively catalyze click labeling of oligonucleotides. This was performed under ultramild...... conditions with fluorophore, peptide, and carbohydrate azide derivatives. In thermal denaturation studies, the modified probes showed specific binding to complementary DNA and RNA targets. Finally, we demonstrated the pH sensitivity of the new rhodamine-based fluorescent probes in vitro and rationalize our...
Injury-induced inhibition of small intestinal protein and nucleic acid synthesis

International Nuclear Information System (INIS)

Carter, E.A.; Hatz, R.A.; Yarmush, M.L.; Tompkins, R.G.

1990-01-01

Small intestinal mucosal weight and nutrient absorption are significantly diminished early after cutaneous thermal injuries. Because these intestinal properties are highly dependent on rates of nucleic acid and protein synthesis, in vivo incorporation of thymidine, uridine, and leucine into small intestinal deoxyribonucleic acid, ribonucleic acid, and proteins were measured. Deoxyribonucleic acid synthesis was markedly decreased with the lowest thymidine incorporation in the jejunum (p less than 0.01); these findings were confirmed by autoradiographic identification of radiolabeled nuclei in the intestinal crypts. Protein synthesis was decreased by 6 h postinjury (p less than 0.01) but had returned to normal by 48 h. Consistent with a decreased rate of protein synthesis, ribonucleic acid synthesis was also decreased 18 h postinjury (p less than 0.01). These decreased deoxyribonucleic acid, ribonucleic acid, and protein synthesis rates are not likely a result of ischemia because in other studies of this injury model, intestinal blood flow was not significantly changed by the burn injury. Potentially, factors initiating the acute inflammatory reaction may directly inhibit nucleic acid and protein synthesis and lead to alterations in nutrient absorption and intestinal barrier function after injury
Metal phosphonate hybrid mesostructures: environmentally friendly multifunctional materials for clean energy and other applications.

Science.gov (United States)

Ma, Tian-Yi; Yuan, Zhong-Yong

2011-10-17

The synthesis of porous hybrid materials has been extended to mesoporous non-silica-based organic-inorganic hybrid materials, in which mesoporous metal phosphonates represent an important family. By using organically bridged polyphosphonic acids as coupling molecules, the homogeneous incorporation of a considerable number of organic functional groups into the metal phosphonate hybrid framework has been realized. Small amounts of organic additives and the pH value of the reaction solution have a large impact on the morphology and textural properties of the resultant hybrid mesoporous metal phosphonate solids. Cationic and nonionic surfactants can be used as templates for the synthesis of ordered mesoporous metal phosphonates. The materials are used as efficient adsorbents for heavy metal ions, CO₂, and aldehydes, as well as in the separation of polycyclic aromatic hydrocarbons. They are also useful photocatalysts under UV and simulated solar light irradiation for organic dye degradation. Further functionalization of the synthesized mesoporous hybrids makes them oxidation and acid catalysts, both with impressive performances in the fields of sustainable energy and environment. Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.
Phosphonates as alternative to tributyl phosphate for the separation of actinides from fission products

International Nuclear Information System (INIS)

Vyas, Chirag K.; Joshirao, Pranav M.; Manchanda, Vijay K.; Rao, C.V.S. Brahmmananda; Jayalakshmi, S.

2015-01-01

The present work investigates the role of increase in the basicity of organophosphorus extractant (dialkylalkyl phosphonates) on the uptake of actinides and fission products vis-a-vis tributyl phosphate (TBP), currently employed as a universal extractant. Two dialkylalkyl phosphonates viz. dibutylpropyl phosphonate (DBPrP) and dibutylpentyl phosphonate (DBPeP) were synthesized, characterized and evaluated for their solvent extraction behavior towards U(VI), Th(IV), Eu(III) and Tc(VII) in nitric acid medium ranging from 0.01-6 M. It was observed that increasing the basicity of the phosphoryl oxygen enhanced the uptake of the actinides and the distribution coefficient values were significantly larger as compared to TBP. The limiting organic concentration (LOC) value was estimated for Th(IV) for these extractants and compared with the TBP system. The separation factors of actinides with phosphonates over Tc(VII) are distinctly better than that with TBP.
High-throughput strategy for molecular identification of Vel- blood donors employing nucleic acids extracted from plasma pools used for viral nucleic acid test screening.

Science.gov (United States)

Dezan, Marcia R; Dinardo, Carla L; Bosi, Silvia R A; Vega, Sileni; Salles, Nanci A; Mendrone-Júnior, Alfredo; Levi, José E

2016-06-01

Serologic methods to determine the Vel- phenotype require the use of rare human antisera and do not allow for many samples to be tested simultaneously, which limits their application as a tool to search for rare donors. This study developed a low-cost molecular screening strategy using real-time polymerase chain reaction (PCR) and DNA, extracted from plasma pools for viral nucleic acid test (NAT) screening, to identify Vel- and Vel+(W) donors. A total of 4680 blood donors from the Brazilian southeast region were genotyped through real-time PCR targeting the 17-nucleotide (c.64_80del) deletion in the SMIM1 gene, which determines the Vel- phenotype, by using remaining nucleic acid from plasma pools of six donors, routinely discarded after the release of viral NAT results. Twenty pools tested reactive and individual testing of samples from reactive pools identified 19 heterozygous donors with the SMIM1*64_80del deletion (0.40%) and one homozygous donor (0.02%). Fourteen of the 19 donors were confirmed as Vel- or Vel+(W) using anti-Vel human antiserum. The DNA pool genotyping strategy using real-time PCR designed to detect the deletion in the SMIM1 gene proved effective and accurate in identifying donors with the Vel- and Vel+(W) phenotypes. The fact that remaining nucleic acid from routine viral NAT screening was used makes this technique economically attractive and definitely superior to the serologic techniques available to search for this rare phenotype. © 2016 AABB.
Microfluidic "Pouch" Chips for Immunoassays and Nucleic Acid Amplification Tests.

Science.gov (United States)

Mauk, Michael G; Liu, Changchun; Qiu, Xianbo; Chen, Dafeng; Song, Jinzhao; Bau, Haim H

2017-01-01

Microfluidic cassettes ("chips") for processing and analysis of clinical specimens and other sample types facilitate point-of-care (POC) immunoassays and nucleic acid based amplification tests. These single-use test chips can be self-contained and made amenable to autonomous operation-reducing or eliminating supporting instrumentation-by incorporating laminated, pliable "pouch" and membrane structures for fluid storage, pumping, mixing, and flow control. Materials and methods for integrating flexible pouch compartments and diaphragm valves into hard plastic (e.g., acrylic and polycarbonate) microfluidic "chips" for reagent storage, fluid actuation, and flow control are described. We review several versions of these pouch chips for immunoassay and nucleic acid amplification tests, and describe related fabrication techniques. These protocols thus offer a "toolbox" of methods for storage, pumping, and flow control functions in microfluidic devices.
Obtention of Samarium and Gadolinium concentrates by solvent extraction using mono-2-ethylhexyl ester of 2-ethylhexyl phosphonic acid

International Nuclear Information System (INIS)

Miranda Junior, Pedro

1996-01-01

The rare earth chlorides solution employed in this study, which is constituted by medium and heavy fractions, is derived from monazite processing accomplished by NUCLEMON-Mineroquimica (SP). This solution shows an acidity about 1.18 M and 189 g/L of rare earth oxides, containing as main constituents: Sm(34.55%), Gd(23.85%), Dy (6.82%), and Y (24.45%). It was used, as organic phase, 2-ethylhexyl phosphonic acid, mono-2-ethylhexylester diluted to 1 M in isododecane. (author)
Nucleic Acid Amplification as used in the Diagnosis and ...

African Journals Online (AJOL)

Nucleic Acid Amplification as used in the Diagnosis and Management of Viral Diseases: A Review. ... Bayero Journal of Pure and Applied Sciences ... are highly unculturable, fastidious or hazardous to the laboratory personnel and diagnosis depends on serological methods or culture in an expensive bio-safety level.
Study of nucleic acids variations in children in nearest areas to the Chernobyl accident

International Nuclear Information System (INIS)

Morera, L.; Navarro, I.; Proenza, E.; Estrada, L.

1996-01-01

The technique used to determine the nucleic acids concentration of leucocytes in peripheral blood is simple, quick and reproducible. Mathematical patterns for small doses have been achieved by means of this technique, which is also useful as biochemical indicator for evaluating exposure to ionizing radiations. The following information is the result of a research in 445 children from 43 locations that suffered, in different degrees, the effect of this accident. Children were grouped taking into account different degrees of superficial pollution for Cs-137 of original places. Groups were built as follows: G-1, 165 children from 20 superficially polluted places between 0-37 KBq/m 2 ; G-2, 135 children from 15 locations with a level of superficial pollution higher than 37 KBq/m 2 and lower than 185 KBq/m 2 ; G-3, 85 children from 7 locations with a pollution degree higher than 296 KBq/m 2 and G-4, 60 children from a place with a non-determined superficial pollution. Values within the interval 1,29 - 4,89 mg/100 ml of blood samples taken from healthy children in group G-1 were considered as normal. The increase in dose led neither to a decrease in nucleic acids medium value, nor to a meaningful growth in the number of cases with nucleic acids figures under rank considered normal. On the other hand, thyroid hyperplasia did not lead either to an increase in medium values of nucleic acids or to an increase in the number of cases with nucleic acids figures over intervals considered as normal. (authors). 10 refs., 2 tabs
Nucleic acids and smart materials: advanced building blocks for logic systems.

Science.gov (United States)

Pu, Fang; Ren, Jinsong; Qu, Xiaogang

2014-09-03

Logic gates can convert input signals into a defined output signal, which is the fundamental basis of computing. Inspired by molecular switching from one state to another under an external stimulus, molecular logic gates are explored extensively and recognized as an alternative to traditional silicon-based computing. Among various building blocks of molecular logic gates, nucleic acid attracts special attention owing to its specific recognition abilities and structural features. Functional materials with unique physical and chemical properties offer significant advantages and are used in many fields. The integration of nucleic acids and functional materials is expected to bring about several new phenomena. In this Progress Report, recent progress in the construction of logic gates by combining the properties of a range of smart materials with nucleic acids is introduced. According to the structural characteristics and composition, functional materials are categorized into three classes: polymers, noble-metal nanomaterials, and inorganic nanomaterials. Furthermore, the unsolved problems and future challenges in the construction of logic gates are discussed. It is hoped that broader interests in introducing new smart materials into the field are inspired and tangible applications for these constructs are found. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Synergistic Combination of Unquenching and Plasmonic Fluorescence Enhancement in Fluorogenic Nucleic Acid Hybridization Probes.

Science.gov (United States)

Vietz, Carolin; Lalkens, Birka; Acuna, Guillermo P; Tinnefeld, Philip

2017-10-11

Fluorogenic nucleic acid hybridization probes are widely used for detecting and quantifying nucleic acids. The achieved sensitivity strongly depends on the contrast between a quenched closed form and an unquenched opened form with liberated fluorescence. So far, this contrast was improved by improving the quenching efficiency of the closed form. In this study, we modularly combine these probes with optical antennas used for plasmonic fluorescence enhancement and study the effect of the nanophotonic structure on the fluorescence of the quenched and the opened form. As quenched fluorescent dyes are usually enhanced more by fluorescence enhancement, a detrimental reduction of the contrast between closed and opened form was anticipated. In contrast, we could achieve a surprising increase of the contrast with full additivity of quenching of the dark form and fluorescence enhancement of the bright form. Using single-molecule experiments, we demonstrate that the additivity of the two mechanisms depends on the perfect quenching in the quenched form, and we delineate the rules for new nucleic acid probes for enhanced contrast and absolute brightness. Fluorogenic hybridization probes optimized not only for quenching but also for the brightness of the open form might find application in nucleic acid assays with PCR avoiding detection schemes.
Crystallization and X-ray diffraction analysis of an ‘all-locked’ nucleic acid duplex derived from a tRNASer microhelix

International Nuclear Information System (INIS)

Behling, Katja; Eichert, André; Fürste, Jens P.; Betzel, Christian; Erdmann, Volker A.; Förster, Charlotte

2009-01-01

A completely ‘all-locked’ nucleic acid duplex was designed from an E. coli tRNA Ser microhelix. The helix consists exclusively of LNA building blocks and was crystallized. The crystals diffracted to 1.9 Å resolution. Modified nucleic acids are of great interest with respect to their nuclease resistance and enhanced thermostability. In therapeutical and diagnostic applications, such molecules can substitute for labile natural nucleic acids that are targeted against particular diseases or applied in gene therapy. The so-called ‘locked nucleic acids’ contain modified sugar moieties such as 2′-O,4′-C-methylene-bridged β-d-ribofuranose and are known to be very stable nucleic acid derivatives. The structure of locked nucleic acids in single or multiple LNA-substituted natural nucleic acids and in LNA–DNA or LNA–RNA heteroduplexes has been well investigated, but the X-ray structure of an ‘all-locked’ nucleic acid double helix has not been described to date. Here, the crystallization and X-ray diffraction data analysis of an ‘all-locked’ nucleic acid helix, which was designed as an LNA originating from a tRNA Ser microhelix RNA structure, is presented. The crystals belonged to space group C2, with unit-cell parameters a = 77.91, b = 40.74, c = 30.06 Å, β = 91.02°. A high-resolution and a low-resolution data set were recorded, with the high-resolution data showing diffraction to 1.9 Å resolution. The crystals contained two double helices per asymmetric unit, with a Matthews coefficient of 2.48 Å 3 Da −1 and a solvent content of 66.49% for the merged data
Solid-state NMR studies of nucleic acid components

Czech Academy of Sciences Publication Activity Database

Dračínský, Martin; Hodgkinson, P.

2015-01-01

Roč. 5, č. 16 (2015), s. 12300-12310 ISSN 2046-2069 R&D Projects: GA ČR GA13-24880S Institutional support: RVO:61388963 Keywords : NMR spectroscopy * nucleic acid s * solid-state NMR Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 3.289, year: 2015 http://pubs.rsc.org/en/content/articlepdf/2015/ra/c4ra14404j
Watson-Crick hydrogen bonding of unlocked nucleic acids

DEFF Research Database (Denmark)

Langkjær, Niels; Wengel, Jesper; Pasternak, Anna

2015-01-01

We herein describe the synthesis of two new unlocked nucleic acid building blocks containing hypoxanthine and 2,6-diaminopurine as nucleobase moieties and their incorporation into oligonucleotides. The modified oligonucleotides were used to examine the thermodynamic properties of UNA against unmo...... unmodified oligonucleotides and the resulting thermodynamic data support that the hydrogen bonding face of UNA is Watson-Crick like....
Adsorbed States of phosphonate derivatives of N-heterocyclic aromatic compounds, imidazole, thiazole, and pyridine on colloidal silver: comparison with a silver electrode.

Science.gov (United States)

Podstawka, Edyta; Olszewski, Tomasz K; Boduszek, Bogdan; Proniewicz, Leonard M

2009-09-03

Here, we report a systematic surface-enhanced Raman spectroscopy (SERS) study of the structures of phosphonate derivatives of the N-heterocyclic aromatic compounds imidazole (ImMeP ([hydroxy(1H-imidazol-5-yl)methyl]phosphonic acid) and (ImMe)(2)P (bis[hydroxy-(1H-imidazol-4-yl)-methyl]phosphinic acid)), thiazole (BAThMeP (butylaminothiazol-2-yl-methyl)phosphonic acid) and BzAThMeP (benzylaminothiazol-2-yl-methyl)phosphonic acid)), and pyridine ((PyMe)(2)P (bis[(hydroxypyridin-3-yl-methyl)]phosphinic acid)) adsorbed on nanometer-sized colloidal particles. We compared these structures to those on a roughened silver electrode surface to determine the relationship between the adsorption strength and the geometry. For example, we showed that all of these biomolecules interact with the colloidal surface through aromatic rings. However, for BzAThMeP, a preferential interaction between the benzene ring and the colloidal silver surface is observed more so than that between the thiazole ring and this substrate. The PC(OH)C fragment does not take part in the adsorption process, and the phosphonate moiety of ImMeP and (ImMe)(2)P, being removed from the surface, only assists in this process.
Immune activation by nucleic acids: A role in pregnancy complications.

Science.gov (United States)

Konečná, B; Lauková, L; Vlková, B

2018-04-01

Cell-free self-DNA or RNA may induce an immune response by activating specific sensing receptors. During pregnancy, placental nucleic acids present in the maternal circulation further activate these receptors due to the presence of unmethylated CpG islands. A higher concentration of cell-free foetal DNA is associated with pregnancy complications and a higher risk for foetal rejection. Cell-free foetal DNA originates from placental trophoblasts. It appears in different forms: free, bound to histones in nucleosomes, in neutrophil extracellular traps (NETs) and in extracellular vesicles (EVs). In several pregnancy complications, cell-free foetal DNA triggers the production of proinflammatory cytokines, and this production results in a cellular and humoral immune response. This review discusses preeclampsia, systemic lupus erythematosus, foetal growth restriction, gestational diabetes, rheumatoid arthritis and obesity in pregnancy from an immunological point of view and closely examines the different pathways that result in maternal inflammation. Understanding the role of cell-free nucleic acids, as well as the biogenesis of NETs and EVs, will help us to specify their functions or targets, which seem to be important in pregnancy complications. It is still not clear whether higher concentrations of cell-free nucleic acids in the maternal circulation are the cause or consequence of various complications. Therefore, further clinical studies and, even more importantly, animal experiments that focus on the involved immunological pathways are needed. © 2018 The Foundation for the Scandinavian Journal of Immunology.
Effects of radiation, acid, and base on the extractant dihexyl-[(diethylcarbamoyl)methyl] phosphonate

International Nuclear Information System (INIS)

Bahner, C.T.; Shoun, R.R.; McDowell, W.J.

1981-11-01

The effects of exposure to gamma radiation ( 60 Co) and of contact with acidic and basic aqueous solutions on dihexyl[(diethylcarbamoyl)methyl]phosphonate (DHDECMP) were studied. Gamma radiation decomposes DHDECMP into a variety of products. The most troublesome of those are the acidic compounds that cause problems in stripping the actinides and lanthanides from the extractant at low acid concentrations. The rate of degradation of DHDECMP by radiation is about the same or only slightly higher than that of tri-n-butyl phosphate (TBP). It is relatively easy to remove the radiation-produced impurities by equilibration (scrubbing) with sodium carbonate or sodium hydroxide or by column chromatographic methods. The hydrolysis of DHDECMP in contact with aqueous solutions containing less than 3 M HNO 3 is not more severe than that of TBP under the same conditions but is significant above that acid concentration. Hydrolysis of DHDECMP in contact with aqueous sodium hydroxide solution does occur, but it should not pose an important problem with the short contact times such as those anticipated for the removal of the radiation-induced degradation products by caustic scrubbing. Results of various chromatographic tests to characterize the degradation products of DHDECMP are also given
Nucleic acid stains as indicators of Giardia muris viability following cyst inactivation.

Science.gov (United States)

Taghi-Kilani, R; Gyürék, L L; Millard, P J; Finch, G R; Belosevic, M

1996-06-01

A reliable viability assay for Giardia is required for the development of disinfection process design criteria and pathogen monitoring by water treatment utilities. Surveys of single-staining nucleic acid dyes (stain dead parasites only), and double-staining vital dye kits from Molecular Probes (stain live and dead parasites) were conducted to assess the viability of untreated, heat-killed, and chemically inactivated Giardia muris cysts. Nucleic acid staining results were compared to those of in vitro excystation and animal infectivity. Nucleic acid stain, designated as SYTO-9, was considered the best among the single-staining dyes for its ability to stain dead cysts brightly and its relatively slow decay rate of visible light emission following DNA binding. SYTO-9 staining was correlated to animal infectivity. A Live/Dead BacLight was found to be the better of 2 double-staining viability kits tested. Logarithmic survival ratios based on SYTO-9 and Live/Dead BacLight were compared to excystation and infectivity results for G. muris cysts exposed to ozone or free chlorine. The results indicate that SYTO-9 and Live/Dead BacLight staining is stable following treatment of cysts with chemical disinfectants.
A Tat-conjugated Peptide Nucleic Acid Tat-PNA-DR Inhibits Hepatitis B Virus Replication In Vitro and In Vivo by Targeting LTR Direct Repeats of HBV RNA

Science.gov (United States)

Zeng, Zhengyang; Han, Shisong; Hong, Wei; Lang, Yange; Li, Fangfang; Liu, Yongxiang; Li, Zeyong; Wu, Yingliang; Li, Wenxin; Zhang, Xianzheng; Cao, Zhijian

2016-01-01

Hepatitis B virus (HBV) infection is a major cause of chronic active hepatitis, cirrhosis, and primary hepatocellular carcinoma, all of which are severe threats to human health. However, current clinical therapies for HBV are limited by potential side effects, toxicity, and drug-resistance. In this study, a cell-penetrating peptide-conjugated peptide nucleic acid (PNA), Tat-PNA-DR, was designed to target the direct repeat (DR) sequences of HBV. Tat-PNA-DR effectively inhibited HBV replication in HepG2.2.15 cells. Its anti-HBV effect relied on the binding of Tat-PNA-DR to the DR, whereby it suppressed the translation of hepatitis B e antigen (HBeAg), HBsAg, HBV core, hepatitis B virus x protein, and HBV reverse transcriptase (RT) and the reverse transcription of the HBV genome. Furthermore, Tat-PNA-DR administered by intravenous injection efficiently cleared HBeAg and HBsAg in an acute hepatitis B mouse model. Importantly, it induced an 80% decline in HBV DNA in mouse serum, which was similar to the effect of the widely used clinical drug Lamivudine (3TC). Additionally, a long-term hydrodynamics HBV mouse model also demonstrated Tat-PNA-DR's antiviral effect. Interestingly, Tat-PNA-DR displayed low cytotoxicity, low mouse acute toxicity, low immunogenicity, and high serum stability. These data indicate that Tat-PNA-DR is a unique PNA and a promising drug candidate against HBV. PMID:26978579
A Tat-conjugated Peptide Nucleic Acid Tat-PNA-DR Inhibits Hepatitis B Virus Replication In Vitro and In Vivo by Targeting LTR Direct Repeats of HBV RNA

Directory of Open Access Journals (Sweden)

Zhengyang Zeng

2016-01-01

Full Text Available Hepatitis B virus (HBV infection is a major cause of chronic active hepatitis, cirrhosis, and primary hepatocellular carcinoma, all of which are severe threats to human health. However, current clinical therapies for HBV are limited by potential side effects, toxicity, and drug-resistance. In this study, a cell-penetrating peptide-conjugated peptide nucleic acid (PNA, Tat-PNA-DR, was designed to target the direct repeat (DR sequences of HBV. Tat-PNA-DR effectively inhibited HBV replication in HepG2.2.15 cells. Its anti-HBV effect relied on the binding of Tat-PNA-DR to the DR, whereby it suppressed the translation of hepatitis B e antigen (HBeAg, HBsAg, HBV core, hepatitis B virus x protein, and HBV reverse transcriptase (RT and the reverse transcription of the HBV genome. Furthermore, Tat-PNA-DR administered by intravenous injection efficiently cleared HBeAg and HBsAg in an acute hepatitis B mouse model. Importantly, it induced an 80% decline in HBV DNA in mouse serum, which was similar to the effect of the widely used clinical drug Lamivudine (3TC. Additionally, a long-term hydrodynamics HBV mouse model also demonstrated Tat-PNA-DR's antiviral effect. Interestingly, Tat-PNA-DR displayed low cytotoxicity, low mouse acute toxicity, low immunogenicity, and high serum stability. These data indicate that Tat-PNA-DR is a unique PNA and a promising drug candidate against HBV.
Antimicrobial Peptide-PNA Conjugates Selectively Targeting Bacterial Genes

Science.gov (United States)

2013-07-22

antibacterial therapy. Initial publications suggest that conjugates of cell penetrating peptides and PNA’s can overcome the barrier in transporting ...Zhou, Y., Hou, Z., Meng, J., and Luo, X. Targeting RNA polymerase primary σ70 as a therapeutic strategy against methicillin - resistant ... Staphylococcus aureus by antisense peptide nucleic acid. PLoS One. 2012; 7(1):e29886. 2. Good, L., Sandberg, R., Larsson, O., Nielsen, P.E., and Wahlestedt, C
Sequence-selective targeting of duplex DNA by peptide nucleic acids

DEFF Research Database (Denmark)

Nielsen, Peter E

2010-01-01

Sequence-selective gene targeting constitutes an attractive drug-discovery approach for genetic therapy, with the aim of reducing or enhancing the activity of specific genes at the transcriptional level, or as part of a methodology for targeted gene repair. The pseudopeptide DNA mimic peptide...
Synthetic methodology for the preparation of nucleic acid containing peptides

NARCIS (Netherlands)

Heden-van Noort, Gerbrand Jan van der

2012-01-01

Dit proefschrift beschrijft de ontwikkeling van nieuwe methoden voor de synthese van hybride biomoleculen die samengesteld zijn uit een peptide- en een nucleïnezuurfragment. Zulke hybride moleculen komen in de natuur voor en hebben belangrijke functies. In dit proefschrift wordt aandacht besteed aan
Peptide Nucleic Acid Based Molecular Authentication for Identification of Four Medicinal Paeonia Species Using Melting Array Analysis of the Internal Transcribed Spacer 2 Region

Directory of Open Access Journals (Sweden)

Wook Jin Kim

2017-11-01

Full Text Available Accurate taxonomic identification of plant materials in herbal medicines is important for product quality control. The genus Paeonia (Saxifragales is the source of the herbal preparations Paeoniae Radix (Paeoniae Radix Alba and Paeoniae Radix Rubra and Moutan Radicis Cotex. However, confusion has arisen regarding their contents due to linguistic and taxonomic ambiguities, similar morphologies and different definitions of Paeoniae Radix in the Korean and Chinese national pharmacopoeias, leading to the distribution of adulterated products. To develop a method for identifying the four Paeonia species used in these medicines, three fluorescently-labeled peptide nucleic acid (PNA probes were designed against ITS2 sequences containing single nucleotide polymorphisms (SNPs and used in a real-time PCR melting curve assay. Each of the four Paeonia species was accurately identified using this analysis. The accuracy and analytical stability of the PNA melting curve assay was confirmed using commercially available samples of the four Paeonia species. This assay is a reliable genetic tool to distinguish between different Paeonia-derived herbal medicines and identify the botanical origins of Paeoniae Radix and Moutan Radicis Cortex. This technique may also contribute to quality control and standardization of herbal medicines by providing a reliable authentication tool and preventing the distribution of inauthentic adulterants.
Peptide Nucleic Acid Based Molecular Authentication for Identification of Four Medicinal Paeonia Species Using Melting Array Analysis of the Internal Transcribed Spacer 2 Region.

Science.gov (United States)

Kim, Wook Jin; Yang, Sungyu; Choi, Goya; Moon, Byeong Cheol

2017-11-07

Accurate taxonomic identification of plant materials in herbal medicines is important for product quality control. The genus Paeonia (Saxifragales) is the source of the herbal preparations Paeoniae Radix (Paeoniae Radix Alba and Paeoniae Radix Rubra) and Moutan Radicis Cotex. However, confusion has arisen regarding their contents due to linguistic and taxonomic ambiguities, similar morphologies and different definitions of Paeoniae Radix in the Korean and Chinese national pharmacopoeias, leading to the distribution of adulterated products. To develop a method for identifying the four Paeonia species used in these medicines, three fluorescently-labeled peptide nucleic acid (PNA) probes were designed against ITS2 sequences containing single nucleotide polymorphisms (SNPs) and used in a real-time PCR melting curve assay. Each of the four Paeonia species was accurately identified using this analysis. The accuracy and analytical stability of the PNA melting curve assay was confirmed using commercially available samples of the four Paeonia species. This assay is a reliable genetic tool to distinguish between different Paeonia -derived herbal medicines and identify the botanical origins of Paeoniae Radix and Moutan Radicis Cortex. This technique may also contribute to quality control and standardization of herbal medicines by providing a reliable authentication tool and preventing the distribution of inauthentic adulterants.
Amino acid substrates impose polyamine, eIF5A, or hypusine requirement for peptide synthesis.

Science.gov (United States)

Shin, Byung-Sik; Katoh, Takayuki; Gutierrez, Erik; Kim, Joo-Ran; Suga, Hiroaki; Dever, Thomas E

2017-08-21

Whereas ribosomes efficiently catalyze peptide bond synthesis by most amino acids, the imino acid proline is a poor substrate for protein synthesis. Previous studies have shown that the translation factor eIF5A and its bacterial ortholog EF-P bind in the E site of the ribosome where they contact the peptidyl-tRNA in the P site and play a critical role in promoting the synthesis of polyproline peptides. Using misacylated Pro-tRNAPhe and Phe-tRNAPro, we show that the imino acid proline and not tRNAPro imposes the primary eIF5A requirement for polyproline synthesis. Though most proline analogs require eIF5A for efficient peptide synthesis, azetidine-2-caboxylic acid, a more flexible four-membered ring derivative of proline, shows relaxed eIF5A dependency, indicating that the structural rigidity of proline might contribute to the requirement for eIF5A. Finally, we examine the interplay between eIF5A and polyamines in promoting translation elongation. We show that eIF5A can obviate the polyamine requirement for general translation elongation, and that this activity is independent of the conserved hypusine modification on eIF5A. Thus, we propose that the body of eIF5A functionally substitutes for polyamines to promote general protein synthesis and that the hypusine modification on eIF5A is critically important for poor substrates like proline. Published by Oxford University Press on behalf of Nucleic Acids Research 2017.
DMPD: Plasmacytoid dendritic cells: sensing nucleic acids in viral infection andautoimmune diseases. [Dynamic Macrophage Pathway CSML Database

Lifescience Database Archive (English)

Full Text Available 18641647 Plasmacytoid dendritic cells: sensing nucleic acids in viral infection andautoimmune dise... (.csml) Show Plasmacytoid dendritic cells: sensing nucleic acids in viral infection andautoimmune diseases....iral infection andautoimmune diseases. Authors Gilliet M, Cao W, Liu YJ. Publication Nat Rev Immunol. 2008 A
Nucleic Acid-based Detection of Bacterial Pathogens Using Integrated Microfluidic Platform Systems

Directory of Open Access Journals (Sweden)

Carl A. Batt

2009-05-01

Full Text Available The advent of nucleic acid-based pathogen detection methods offers increased sensitivity and specificity over traditional microbiological techniques, driving the development of portable, integrated biosensors. The miniaturization and automation of integrated detection systems presents a significant advantage for rapid, portable field-based testing. In this review, we highlight current developments and directions in nucleic acid-based micro total analysis systems for the detection of bacterial pathogens. Recent progress in the miniaturization of microfluidic processing steps for cell capture, DNA extraction and purification, polymerase chain reaction, and product detection are detailed. Discussions include strategies and challenges for implementation of an integrated portable platform.
Synthetic oligonucleotide antigens modified with locked nucleic acids detect disease specific antibodies

DEFF Research Database (Denmark)

Samuelsen, Simone V; Solov'yov, Ilia A.; Balboni, Imelda M.

2016-01-01

New techniques to detect and quantify antibodies to nucleic acids would provide a significant advance over current methods, which often lack specificity. We investigate the potential of novel antigens containing locked nucleic acids (LNAs) as targets for antibodies. Particularly, employing...... molecular dynamics we predict optimal nucleotide composition for targeting DNA-binding antibodies. As a proof of concept, we address a problem of detecting anti-DNA antibodies that are characteristic of systemic lupus erythematosus, a chronic autoimmune disease with multiple manifestations. We test the best...... that the novel method is a promising tool to create antigens for research and point-of-care monitoring of anti-DNA antibodies....
Method and apparatus for purifying nucleic acids and performing polymerase chain reaction assays using an immiscible fluid

Energy Technology Data Exchange (ETDEWEB)

Koh, Chung-Yan; Light, Yooli Kim; Piccini, Matthew Ernest; Singh, Anup K.

2017-10-31

Embodiments of the present invention are directed toward devices, systems, and methods for purifying nucleic acids to conduct polymerase chain reaction (PCR) assays. In one example, a method includes generating complexes of silica beads and nucleic acids in a lysis buffer, transporting the complexes through an immiscible fluid to remove interfering compounds from the complexes, further transporting the complexes into a density medium containing components required for PCR where the nucleic acids disassociate from the silica beads, and thermocycling the contents of the density medium to achieve PCR. Signal may be detected from labeling agents in the components required for PCR.

Radiation-induced electron migration in nucleic acids

International Nuclear Information System (INIS)

Fuciarelli, A.F.; Sisk, E.C.; Miller, J.H.; Zimbrick, J.D.

1994-01-01

Radiation-induced electron migration along DNA is a mechanism by which randomly produced stochastic energy deposition events can lead to non-random types of damage along DNA manifested distal to the sites of the initial energy deposition. Radiation-induced electron migration in nucleic acids has been examined using oligonucleotides containing 5-bromouracil (5-BrU). Interaction of 5-BrU with solvated electrons results in release of bromide ions and formation of uracil-5-yl radicals. Monitoring either bromide ion release or uracil formation provides an opportunity to study electron migration processes in model nucleic acid systems. Using this approach we have discovered that electron migration along oligonucleotides is significantly influenced by the base sequence and strandedness. Migration along 7 base pairs in oligonucleotides containing guanine bases was observed for oligonucleotides irradiated in solution, which compares with mean migration distances of 6-10 bp for Escherichia coli DNA irradiated in solution and 5.5 bp for E. coli DNA irradiated in cells. Evidence also suggests that electron migration can occur preferentially in the 5' to 3' direction along a double-stranded oligonucleotide containing a region of purine bases adjacent to the 5-BrU moiety. Our continued efforts will provide information regarding the contribution of electron transfer along DNA to formation of locally multiply damaged sites created in DNA by exposure to ionizing radiation. (Author)
Karyotype and nucleic acid content in Zantedeschia aethiopica Spr ...

African Journals Online (AJOL)

DR. NJ TONUKARI

2012-07-03

Jul 3, 2012 ... Analysis of karyotype, nucleic deoxyribonucleic acid (DNA) content and sodium dodecyl sulfate polyacrylamide ... base pairs) for Z. aethiopica and 1144.26 ± 0.05 picograms (equivalent to 1144.26 mega base pairs) for Z. elliottiana. ... ml ice-cold nuclei-isolation buffer A of the Partec high resolution. DNA kit ...
Modification of nucleic acids by azobenzene derivatives and their applications in biotechnology and nanotechnology.

Science.gov (United States)

Li, Jing; Wang, Xingyu; Liang, Xingguo

2014-12-01

Azobenzene has been widely used as a photoregulator due to its reversible photoisomerization, large structural change between E and Z isomers, high photoisomerization yield, and high chemical stability. On the other hand, some azobenzene derivatives can be used as universal quenchers for many fluorophores. Nucleic acid is a good candidate to be modified because it is not only the template of gene expression but also widely used for building well-organized nanostructures and nanodevices. Because the size and polarity distribution of the azobenzene molecule is similar to a nucleobase pair, the introduction of azobenzene into nucleic acids has been shown to be an ingenious molecular design for constructing light-switching biosystems or light-driven nanomachines. Here we review recent advances in azobenzene-modified nucleic acids and their applications for artificial regulation of gene expression and enzymatic reactions, construction of photoresponsive nanostructures and nanodevices, molecular beacons, as well as obtaining structural information using the introduced azobenzene as an internal probe. In particular, nucleic acids bearing multiple azobenzenes can be used as a novel artificial nanomaterial with merits of high sequence specificity, regular duplex structure, and high photoregulation efficiency. The combination of functional groups with biomolecules may further advance the development of chemical biotechnology and biomolecular engineering. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.
Unlocked Nucleic Acids with a Pyrene-Modified Uracil: Synthesis, Hybridization Studies, Fluorescent Properties and i-Motif Stability

Czech Academy of Sciences Publication Activity Database

Perlíková, Pavla; Karlsen, K. K.; Pedersen, E. B.; Wengel, J.

2014-01-01

Roč. 15, č. 1 (2014), s. 146-156 ISSN 1439-4227 Grant - others:European Research Council(XE) FP7-268776 Institutional support: RVO:61388963 Keywords : fluorescence * i-motifs * nucleic acid hybridization * oligonucleotides * unlocked nucleic acids Subject RIV: CE - Biochemistry Impact factor: 3.088, year: 2014
Development of Temperature Control Solutions for Non-Instrumented Nucleic Acid Amplification Tests (NINAAT

Directory of Open Access Journals (Sweden)

Tamás Pardy

2017-06-01

Full Text Available Non-instrumented nucleic acid amplification tests (NINAAT are a novel paradigm in portable molecular diagnostics. They offer the high detection accuracy characteristic of nucleic acid amplification tests (NAAT in a self-contained device, without the need for any external instrumentation. These Point-of-Care tests typically employ a Lab-on-a-Chip for liquid handling functionality, and perform isothermal nucleic acid amplification protocols that require low power but high accuracy temperature control in a single well-defined temperature range. We propose temperature control solutions based on commercially available heating elements capable of meeting these challenges, as well as demonstrate the process by which such elements can be fitted to a NINAAT system. Self-regulated and thermostat-controlled resistive heating elements were evaluated through experimental characterization as well as thermal analysis using the finite element method (FEM. We demonstrate that the proposed solutions can support various NAAT protocols, as well as demonstrate an optimal solution for the loop-mediated isothermal amplification (LAMP protocol. Furthermore, we present an Arduino-compatible open-source thermostat developed for NINAAT applications.
DMPD: The role of viral nucleic acid recognition in dendritic cells for innate andadaptive antiviral immunity. [Dynamic Macrophage Pathway CSML Database

Lifescience Database Archive (English)

Full Text Available 18086372 The role of viral nucleic acid recognition in dendritic cells for innate a...1-14. Epub 2007 Nov 9. (.png) (.svg) (.html) (.csml) Show The role of viral nucleic acid recognition in dend...e role of viral nucleic acid recognition in dendritic cells for innate andadaptive antiviral immunity. Autho
Design of polymer motifs for nucleic acid recognition and assembly stabilization

Science.gov (United States)

Zhou, Zhun

This dissertation describes the synthesis and assembly of bio-functional polymers and the applications of these polymers to drug encapsulation, delivery, and multivalent biomimetic macromolecular recognition between synthetic polymer and nucleic acids. The main content is divided into three parts: (1) polyacidic domains as strongly stabilizing design elements for aqueous phase polyacrylate diblock assembly; (2) small molecule/polymer recognition triggered macromolecular assembly and drug encapsulation; (3) trizaine derivatized polymer as a novel class of "bifacial polymer nucleic acid" (bPoNA) and applications of bPoNA to nanoparticle loading of DNA/RNA, silencing delivery as well as control of aptamer function. Through the studies in part (1) and part (2), it was demonstrated that well-designed polymer motifs are not only able to enhance assemblies driven by non-specific hydrophobic effect, but are also able to direct assemblies based on specific recognitions. In part (3) of this dissertation, this concept was further extended by the design of polyacrylate polymers that are capable of discrete and robust hybridization with nucleic acids. This surprising finding demonstrated both fundamental and practical applications. Overall, these studies provided insights into the rational design elements for improving the bio-functions of synthetic polymers, and significantly expanded the scope of biological applications in which polymers synthesized via controlled radical polymerization may play a role.
Nucleic acid-binding properties of the RRM-containing protein RDM1

International Nuclear Information System (INIS)

Hamimes, Samia; Bourgeon, Dominique; Stasiak, Alicja Z.; Stasiak, Andrzej; Van Dyck, Eric

2006-01-01

RDM1 (RAD52 Motif 1) is a vertebrate protein involved in the cellular response to the anti-cancer drug cisplatin. In addition to an RNA recognition motif, RDM1 contains a small amino acid motif, named RD motif, which it shares with the recombination and repair protein, RAD52. RDM1 binds to single- and double-stranded DNA, and recognizes DNA distortions induced by cisplatin adducts in vitro. Here, we have performed an in-depth analysis of the nucleic acid-binding properties of RDM1 using gel-shift assays and electron microscopy. We show that RDM1 possesses acidic pH-dependent DNA-binding activity and that it binds RNA as well as DNA, and we present evidence from competition gel-shift experiments that RDM1 may be capable of discrimination between the two nucleic acids. Based on reported studies of RAD52, we have generated an RDM1 variant mutated in its RD motif. We find that the L 119 GF → AAA mutation affects the mode of RDM1 binding to single-stranded DNA
[Genotoxic modification of nucleic acid bases and biological consequences of it. Review and prospects of experimental and computational investigations

Science.gov (United States)

Poltev, V. I.; Bruskov, V. I.; Shuliupina, N. V.; Rein, R.; Shibata, M.; Ornstein, R.; Miller, J.

1993-01-01

The review is presented of experimental and computational data on the influence of genotoxic modification of bases (deamination, alkylation, oxidation) on the structure and biological functioning of nucleic acids. Pathways are discussed for the influence of modification on coding properties of bases, on possible errors of nucleic acid biosynthesis, and on configurations of nucleotide mispairs. The atomic structure of nucleic acid fragments with modified bases and the role of base damages in mutagenesis and carcinogenesis are considered.
Simple Approaches to Minimally-Instrumented, Microfluidic-Based Point-of-Care Nucleic Acid Amplification Tests

Science.gov (United States)

Mauk, Michael G.; Song, Jinzhao; Liu, Changchun; Bau, Haim H.

2018-01-01

Designs and applications of microfluidics-based devices for molecular diagnostics (Nucleic Acid Amplification Tests, NAATs) in infectious disease testing are reviewed, with emphasis on minimally instrumented, point-of-care (POC) tests for resource-limited settings. Microfluidic cartridges (‘chips’) that combine solid-phase nucleic acid extraction; isothermal enzymatic nucleic acid amplification; pre-stored, paraffin-encapsulated lyophilized reagents; and real-time or endpoint optical detection are described. These chips can be used with a companion module for separating plasma from blood through a combined sedimentation-filtration effect. Three reporter types: Fluorescence, colorimetric dyes, and bioluminescence; and a new paradigm for end-point detection based on a diffusion-reaction column are compared. Multiplexing (parallel amplification and detection of multiple targets) is demonstrated. Low-cost detection and added functionality (data analysis, control, communication) can be realized using a cellphone platform with the chip. Some related and similar-purposed approaches by others are surveyed. PMID:29495424
Simple Approaches to Minimally-Instrumented, Microfluidic-Based Point-of-Care Nucleic Acid Amplification Tests

Directory of Open Access Journals (Sweden)

Michael G. Mauk

2018-02-01

Full Text Available Designs and applications of microfluidics-based devices for molecular diagnostics (Nucleic Acid Amplification Tests, NAATs in infectious disease testing are reviewed, with emphasis on minimally instrumented, point-of-care (POC tests for resource-limited settings. Microfluidic cartridges (‘chips’ that combine solid-phase nucleic acid extraction; isothermal enzymatic nucleic acid amplification; pre-stored, paraffin-encapsulated lyophilized reagents; and real-time or endpoint optical detection are described. These chips can be used with a companion module for separating plasma from blood through a combined sedimentation-filtration effect. Three reporter types: Fluorescence, colorimetric dyes, and bioluminescence; and a new paradigm for end-point detection based on a diffusion-reaction column are compared. Multiplexing (parallel amplification and detection of multiple targets is demonstrated. Low-cost detection and added functionality (data analysis, control, communication can be realized using a cellphone platform with the chip. Some related and similar-purposed approaches by others are surveyed.
Molecular polarization potential maps of the nucleic acid bases

International Nuclear Information System (INIS)

Alkorta, I.; Perez, J.J.

1996-01-01

Ab initio calculations at the SCF level were carried out to compute the polarization potential map NM of the nucleic acid bases: cytosine, thymine, uracil, adedine, and guanine. For this purpose, the Dunning's 9s5p basis set contracted to a split-valence, was selected to perform the calculations. The molecular polarization potential (MPP) at each point was evaluated by the difference between the interaction energy of the molecule with a unit point charge and the molecular electrostatic potential (MEP) at that point. MEPS and MPPS for the different molecules were computed with a density of 5 points/Angstrom 2 on the van der Waals surface of each molecule, defined using the van der Waals radii. Due to the symmetry of the molecules, only half the points were computed. The total number of points calculated was 558 for cytosine, 621 for thymine, 526 for uracil, 666 for adenine, and 699 for guanine. The results of these calculations are analyzed in terms of their implications on the molecular interactions between pairs of nucleic acid bases. 23 refs., 5 figs., 1 tab
Evolution of sequence-defined highly functionalized nucleic acid polymers

Science.gov (United States)

Chen, Zhen; Lichtor, Phillip A.; Berliner, Adrian P.; Chen, Jonathan C.; Liu, David R.

2018-03-01

The evolution of sequence-defined synthetic polymers made of building blocks beyond those compatible with polymerase enzymes or the ribosome has the potential to generate new classes of receptors, catalysts and materials. Here we describe a ligase-mediated DNA-templated polymerization and in vitro selection system to evolve highly functionalized nucleic acid polymers (HFNAPs) made from 32 building blocks that contain eight chemically diverse side chains on a DNA backbone. Through iterated cycles of polymer translation, selection and reverse translation, we discovered HFNAPs that bind proprotein convertase subtilisin/kexin type 9 (PCSK9) and interleukin-6, two protein targets implicated in human diseases. Mutation and reselection of an active PCSK9-binding polymer yielded evolved polymers with high affinity (KD = 3 nM). This evolved polymer potently inhibited the binding between PCSK9 and the low-density lipoprotein receptor. Structure-activity relationship studies revealed that specific side chains at defined positions in the polymers are required for binding to their respective targets. Our findings expand the chemical space of evolvable polymers to include densely functionalized nucleic acids with diverse, researcher-defined chemical repertoires.
Nucleic Acid Aptamers Against Proteases

DEFF Research Database (Denmark)

Dupont, D M; Andersen, L M; Bøtkjær, Kenneth Alrø

2011-01-01

, directed against blood coagulation factors, are in clinical trials as anticoagulant drugs. Several of the studies on protease-binding aptamers have been pioneering and trend-setting in the field. The work with protease-binding aptamers also demonstrates many interesting examples of non-standard selection......Proteases are potential or realized therapeutic targets in a wide variety of pathological conditions. Moreover, proteases are classical subjects for studies of enzymatic and regulatory mechanisms. We here review the literature on nucleic acid aptamers selected with proteases as targets. Designing...... small molecule protease inhibitors of sufficient specificity has proved a daunting task. Aptamers seem to represent a promising alternative. In our review, we concentrate on biochemical mechanisms of aptamer selection, proteinaptamer recognition, protease inhibition, and advantages of aptamers...
Rhodium(iii)-catalyzed ortho-olefination of aryl phosphonates.

Science.gov (United States)

Chary, Bathoju Chandra; Kim, Sunggak

2013-09-25

Rhodium(iii)-catalyzed C-H olefination of aryl phosphonic esters is reported for the first time. In this mild and efficient process, the phosphonic ester group is utilized successfully as a new directing group. In addition, mono-olefination for aryl phosphonates is observed using a phosphonic diamide directing group.
Role of SbmA in the Uptake of Peptide Nucleic Acid (PNA)-Peptide Conjugates in E. coli

DEFF Research Database (Denmark)

Ghosal, Anubrata; Vitali, Ally; Stach, James E M

2013-01-01

Antisense PNA oligomers targeting essential genes (acpP or ftsZ) and conjugated to the delivery peptide L((KFF)(3)K) show complete growth inhibition of wild type E. coli strain (MG1655) with submicromolar MIC. In this study we show that resistant mutants generated against such PNA......-peptide conjugates had disruptions in the region of sbmA, a gene encoding an inner membrane peptide transporter. The wild type sensitivity to the PNA conjugates was re-established in the resistance mutants by complementation with sbmA. Furthermore, deletion of sbmA in E. coli AS19, a strain that is sensitive...
Clarithromycin, trimethoprim, and penicillin and oxidative nucleic acid modifications in humans

DEFF Research Database (Denmark)

Larsen, Emil List; Cejvanovic, Vanja; Kjaer, Laura Kofoed

2017-01-01

, phenoxymethylpenicillin (penicillin V), or placebo. Oxidative modifications were measured as 24-h urinary excretion of 8-oxo-7,8-dihydro-2′-deoxyguanosine (8-oxodG) and 8-oxo-7,8-dihydroguanosine (8-oxoGuo), and plasma levels of malondialdehyde before and after treatment as a measurement of DNA oxidation, RNA oxidation.......7% (95% CI: 5.8–37.6%), but did not influence urinary excretion of 8-oxoGuo. Penicillin V did not influence urinary excretion of 8-oxodG or 8-oxoGuo. None of the antibiotic drugs influenced plasma levels of malondialdehyde. Conclusion Clarithromycin significantly increases oxidative nucleic acid...... modifications. Increased oxidative modifications might explain some of clarithromycin's known adverse reactions. Trimethoprim significantly lowers DNA oxidation but not RNA oxidation. Penicillin V had no effect on oxidative nucleic acid modifications....
Nucleic-acid characterization of the identity and activity of subsurface microorganisms

Science.gov (United States)

Madsen, E. L.

Nucleic-acid approaches to characterizing naturally occurring microorganisms in their habitats have risen to prominence during the last decade. Extraction of deoxyribonucleic-acid (DNA) and ribonucleic-acid (RNA) biomarkers directly from environmental samples provides a new means of gathering information in microbial ecology. This review article defines: (1) the subsurface habitat; (2) what nucleic-acid procedures are; and (3) the types of information nucleic-acid procedures can and cannot reveal. Recent literature examining microbial nucleic acids in the terrestrial subsurface is tabulated and reviewed. The majority of effort to date has focused upon insights into the identity and phylogeny of subsurface microorganisms afforded by analysis of their 16S rRNA genes. Given the power of nucleic-acid-based procedures and their limited application to subsurface habitats to date, many future opportunities await exploration. Au cours des derniers dix ans, les approches basées sur les acides nucléiques sont apparues et devenues essentielles pour caractériser dans leurs habitats les microorganismes existant à l'état naturel. L'extraction directe de l'ADN et de l'ARN, qui sont des biomarqueurs, d'échantillons environnementaux a fourni un nouveau moyen d'obtenir des informations sur l'écologie microbienne. Cet article synthétique définit 1) l'habitat souterrain, 2) ce que sont les procédures basées sur les acides nucléiques, 3) quel type d'informations ces procéedures peuvent et ne peuvent pas révéler. Les travaux récemment publiés concernatn les acides nucléiques microbiens dans le milieu souterrain terrestre sont catalogués et passés en revue. La majorité des efforts pour obtenir es données s'est concentrée sur l'identité et la phylogénie des microorganismes souterrains fournies par l'analyse de leurs gènes 16S rRNA. Étant donné la puissance des procédures basées sur les acides nucléiques et leur application limitée aux habitats souterrains
Nucleic acid metabolism in hemopoietic tissues of polycythemic rats during long-term fractionated irradiation

International Nuclear Information System (INIS)

Mushkacheva, G.S.; Murzina, L.D.

1980-01-01

A study was made of the effect of long-term fractionated exposure with a daily dose of 50 R on the nucleic acid metabolism in hemopoietic tissues (bone marrow and spleen) of rats with erythropoiesis selectively inhibited by posttransfusion polycythemia. The comparison of present and previously obtained results enables us to conclude that the pathways of changes in the nucleic acid metabolism, which is responsible for hemopoiesis compensation during long-term exposure, are, in the main, similar for both white and red compartments of hemopoiesis
Novel (Phenylethynyl)pyrene-LNA Constructs for Fluorescence SNP Sensing in Polymorphic Nucleic Acid Targets

DEFF Research Database (Denmark)

Astakhova, Irina Kira; Samokhina, Evgeniya; Babu, B Ravindra

2012-01-01

We describe fluorescent oligonucleotide probes labeled with novel (phenylethynyl)pyrene dyes attached to locked nucleic acids. Furthermore, we prove the utility of these probes for the effective detection of single-nucleotide polymorphisms in natural nucleic acids. High-affinity hybridization......DNA and RNA gene fragments. Target sequences were obtained by analysis of 200 clinical samples from patients currently receiving anti-HIV/AIDS combination therapy at the Russian Federal AIDS Center. Using these fluorescent oligonucleotides, we were able to detect the target mutation despite all the challenges...

Molecular radiobiology of nucleic acids

Energy Technology Data Exchange (ETDEWEB)

Fuciarelli, A F

1987-01-01

In addition to radiolytic adenine release, radiolysis of adenosine 5'-monophosphate, in the absence of oxygen, can result in the formation of 8-hydroxyadenosine 5'-monophosphate and both the (R)- and (S)-epimer of 8,5'-cycloadenosine 5'-monophosphate. The mononucleoside derivatives of these modified nucleotides were also observed in irradiated solutions of adenosine and in the enzyme hydrolysates of irradiated solutions of polyadenylic acid (poly A) using high-performance liquid chromatography (HPLC). In an effort to detect 8,5'-cyclo(deoxy) adenosine formation in irradiated nucleic acids, polyclonal antiserum were raised with specificity to the 8,5'-cycloadenosine 5'-monophosphate moiety and used in an enzyme-linked immunosorbent assay (ELISA). The 8,5'-cyclo(deoxy)adenosine moiety could be detected in nitrous oxide-saturated aqueous solutions containing unhydrolyzed poly A at 10 Gy and DNA at 200 Gy using the colorimetric ELISA. Correlation of product yield measured by ELISA with HPLC analysis of irradiated, enzyme-hydrolyzed solutions of poly A revealed that the ELISA was precisely reflecting changes in the combined yield of (R)- and (S)-8,5'-cycloadenosine.
A nucleic acid dependent chemical photocatalysis in live human cells

DEFF Research Database (Denmark)

Arian, Dumitru; Cló, Emiliano; Gothelf, Kurt V

2010-01-01

Only two nucleic acid directed chemical reactions that are compatible with live cells have been reported to date. Neither of these processes generate toxic species from nontoxic starting materials. Reactions of the latter type could be applied as gene-specific drugs, for example, in the treatment...
Mosaic protein and nucleic acid vaccines against hepatitis C virus

Science.gov (United States)

Yusim, Karina; Korber, Bette T. M.; Kuiken, Carla L.; Fischer, William M.

2013-06-11

The invention relates to immunogenic compositions useful as HCV vaccines. Provided are HCV mosaic polypeptide and nucleic acid compositions which provide higher levels of T-cell epitope coverage while minimizing the occurrence of unnatural and rare epitopes compared to natural HCV polypeptides and consensus HCV sequences.
Nucleic acid probes as a diagnostic method for tick-borne hemoparasites of veterinary importance.

Science.gov (United States)

Figueroa, J V; Buening, G M

1995-03-01

An increased number of articles on the use of nucleic acid-based hybridization techniques for diagnostic purposes have been recently published. This article reviews nucleic acid-based hybridization as an assay to detect hemoparasite infections of economic relevance in veterinary medicine. By using recombinant DNA techniques, selected clones containing inserts of Anaplasma, Babesia, Cowdria or Theileria genomic DNA sequences have been obtained, and they are now available to be utilized as specific, highly sensitive DNA or RNA probes to detect the presence of the hemoparasite DNA in an infected animal. Either in an isotopic or non-isotopic detection system, probes have allowed scientists to test for--originally in samples collected from experimentally infected animals and later in samples collected in the field--the presence of hemoparasites during the prepatent, patent, convalescent, and chronic periods of the infection in the host. Nucleic acid probes have given researchers the opportunity to carry out genomic analysis of parasite DNA to differentiate hemoparasite species and to identify genetically distinct populations among and within isolates, strains and clonal populations. Prevalence of parasite infection in the tick vector can now be accomplished more specifically with the nucleic acid probes. Lately, with the advent of the polymerase chain reaction technique, small numbers of hemoparasites can be positively identified in the vertebrate host and tick vector. These techniques can be used to assess the veterinary epidemiological situation in a particular geographical region for the planning of control measures.
Nucleic acid helix structure determination from NMR proton chemical shifts

Energy Technology Data Exchange (ETDEWEB)

Werf, Ramon M. van der; Tessari, Marco; Wijmenga, Sybren S., E-mail: S.Wijmenga@science.ru.nl [Radboud University Nijmegen, Department of Biophysical Chemistry, Institute of Molecules and Materials (Netherlands)

2013-06-15

We present a method for de novo derivation of the three-dimensional helix structure of nucleic acids using non-exchangeable proton chemical shifts as sole source of experimental restraints. The method is called chemical shift de novo structure derivation protocol employing singular value decomposition (CHEOPS) and uses iterative singular value decomposition to optimize the structure in helix parameter space. The correct performance of CHEOPS and its range of application are established via an extensive set of structure derivations using either simulated or experimental chemical shifts as input. The simulated input data are used to assess in a defined manner the effect of errors or limitations in the input data on the derived structures. We find that the RNA helix parameters can be determined with high accuracy. We finally demonstrate via three deposited RNA structures that experimental proton chemical shifts suffice to derive RNA helix structures with high precision and accuracy. CHEOPS provides, subject to further development, new directions for high-resolution NMR structure determination of nucleic acids.
Adapting capillary gel electrophoresis as a sensitive, high-throughput method to accelerate characterization of nucleic acid metabolic enzymes.

Science.gov (United States)

Greenough, Lucia; Schermerhorn, Kelly M; Mazzola, Laurie; Bybee, Joanna; Rivizzigno, Danielle; Cantin, Elizabeth; Slatko, Barton E; Gardner, Andrew F

2016-01-29

Detailed biochemical characterization of nucleic acid enzymes is fundamental to understanding nucleic acid metabolism, genome replication and repair. We report the development of a rapid, high-throughput fluorescence capillary gel electrophoresis method as an alternative to traditional polyacrylamide gel electrophoresis to characterize nucleic acid metabolic enzymes. The principles of assay design described here can be applied to nearly any enzyme system that acts on a fluorescently labeled oligonucleotide substrate. Herein, we describe several assays using this core capillary gel electrophoresis methodology to accelerate study of nucleic acid enzymes. First, assays were designed to examine DNA polymerase activities including nucleotide incorporation kinetics, strand displacement synthesis and 3'-5' exonuclease activity. Next, DNA repair activities of DNA ligase, flap endonuclease and RNase H2 were monitored. In addition, a multicolor assay that uses four different fluorescently labeled substrates in a single reaction was implemented to characterize GAN nuclease specificity. Finally, a dual-color fluorescence assay to monitor coupled enzyme reactions during Okazaki fragment maturation is described. These assays serve as a template to guide further technical development for enzyme characterization or nucleoside and non-nucleoside inhibitor screening in a high-throughput manner. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.
Structure and behaviour of proteins, nucleic acids and viruses from vibrational Raman optical activity

DEFF Research Database (Denmark)

Barron, L.D.; Blanch, E.W.; McColl, I.H.

2003-01-01

stacking arrangement and the mutual orientation of the sugar and base rings around the C-N glycosidic link. The ROA spectra of intact viruses provide information on the folds of the coat proteins and the nucleic acid structure. The large number of structure-sensitive bands in protein ROA spectra...... is especially favourable for fold determination using pattern recognition techniques. This article gives a brief account of the ROA technique and presents the ROA spectra of a selection of proteins, nucleic acids and viruses that illustrate the applications of ROA spectroscopy in biomolecular research....
Poly(alkylene oxide) Copolymers for Nucleic Acid Delivery

Science.gov (United States)

2012-07-17

Poly(alkylene oxide) Copolymers for Nucleic Acid Delivery Swati Mishra1,#, Lavanya Y. Peddada1,#, David I. Devore3,4, and Charles M. Roth1,2...Neil Raju for assistance with figures. Biographies Swati Mishra received her Ph.D. in Biomedical Engineering and Biotechnology from the University of...Kleiman N, Anderson RD, Gottlieb D, Karlsberg R, Snell J, Rocha- Singh K. Results from a phase II multicenter, double-blind placebo-controlled study of Del
Concentration determination of nucleic acids and proteins using the micro-volume BioSpec-nano-spectrophotometer.

Science.gov (United States)

Sukumaran, Suja

2011-02-17

Nucleic acid quantitation procedures have advanced significantly in the last three decades. More and more, molecular biologists require consistent small-volume analysis of nucleic acid samples for their experiments. The BioSpec-nano provides a potential solution to the problems of inaccurate, non-reproducible results, inherent in current DNA quantitation methods, via specialized optics and a sensitive PDA detector. The BioSpec-nano also has automated functionality such that mounting, measurement, and cleaning are done by the instrument, thereby eliminating tedious, repetitive, and inconsistent placement of the fiber optic element and manual cleaning. In this study, data is presented on the quantification of DNA and protein, as well as on measurement reproducibility and accuracy. Automated sample contact and rapid scanning allows measurement in three seconds, resulting in excellent throughput. Data analysis is carried out using the built-in features of the software. The formula used for calculating DNA concentration is: Sample Concentration = DF · (OD260-OD320)· NACF (1) Where DF = sample dilution factor and NACF = nucleic acid concentration factor. The Nucleic Acid concentration factor is set in accordance with the analyte selected. Protein concentration results can be expressed as μg/mL or as moles/L by entering e280 and molecular weight values respectively. When residue values for Tyr, Trp and Cysteine (S-S bond) are entered in the e280Calc tab, the extinction coefficient values are calculated as e280 = 5500 x (Trp residues) + 1490 x (Tyr residues) + 125 x (cysteine S-S bond). The e280 value is used by the software for concentration calculation. In addition to concentration determination of nucleic acids and protein, the BioSpec-nano can be used as an ultra micro-volume spectrophotometer for many other analytes or as a standard spectrophotometer using 5 mm pathlength cells.
Concentration Determination of Nucleic Acids and Proteins Using the Micro-volume Bio-spec Nano Spectrophotometer

Science.gov (United States)

Sukumaran, Suja

2011-01-01

Nucleic Acid quantitation procedures have advanced significantly in the last three decades. More and more, molecular biologists require consistent small-volume analysis of nucleic acid samples for their experiments. The BioSpec-nano provides a potential solution to the problems of inaccurate, non-reproducible results, inherent in current DNA quantitation methods, via specialized optics and a sensitive PDA detector. The BioSpec-nano also has automated functionality such that mounting, measurement, and cleaning are done by the instrument, thereby eliminating tedious, repetitive, and inconsistent placement of the fiber optic element and manual cleaning. In this study, data is presented on the quantification of DNA and protein, as well as on measurement reproducibility and accuracy. Automated sample contact and rapid scanning allows measurement in three seconds, resulting in excellent throughput. Data analysis is carried out using the built-in features of the software. The formula used for calculating DNA concentration is: Sample Concentration = DF · (OD260-OD320)· NACF (1) Where DF = sample dilution factor and NACF = nucleic acid concentration factor. The Nucleic Acid concentration factor is set in accordance with the analyte selected1. Protein concentration results can be expressed as μg/ mL or as moles/L by entering e280 and molecular weight values respectively. When residue values for Tyr, Trp and Cysteine (S-S bond) are entered in the e280Calc tab, the extinction coefficient values are calculated as e280 = 5500 x (Trp residues) + 1490 x (Tyr residues) + 125 x (cysteine S-S bond). The e280 value is used by the software for concentration calculation. In addition to concentration determination of nucleic acids and protein, the BioSpec-nano can be used as an ultra micro-volume spectrophotometer for many other analytes or as a standard spectrophotometer using 5 mm pathlength cells. PMID:21372788
Nucleic acid detection with surface plasmon resonance using cationic latex

NARCIS (Netherlands)

de Vries, E.F.A.; Schasfoort, Richardus B.M.; van der Plas, J.; Greve, Jan

1994-01-01

An affinity sensor based on Surface Plasmon Resonance (SPR) was used to detect nucleic acids. SPR is an optical technique that is able to detect small changes in the refractive index of the immediate vicinity of a metal surface. After a specific amplification of DNA, achieved using the polymerase
Highly Efficient Synthesis of Allopurinol Locked Nucleic Acid Monomer by C6 Deamination of 8-Aza-7-bromo-7-deazaadenine Locked Nucleic Acid Monomer

DEFF Research Database (Denmark)

Kosbar, Tamer Reda El-Saeed; Sofan, M.; Abou-Zeid, L.

2013-01-01

An allopurinol locked nucleic acid (LNA) monomer was prepared by a novel strategy through C6 deamination of the corresponding 8-aza-7-bromo-7-deazaadenine LNA monomer with aqueous sodium hydroxide. An 8-aza-7-deazaadenine LNA monomer was also synthesized by a modification of the new synthetic...... the required LNA monomers....
A non-covalent peptide-based carrier for in vivo delivery of DNA mimics.

Science.gov (United States)

Morris, May C; Gros, Edwige; Aldrian-Herrada, Gudrun; Choob, Michael; Archdeacon, John; Heitz, Frederic; Divita, Gilles

2007-01-01

The dramatic acceleration in identification of new nucleic-acid-based therapeutic molecules has provided new perspectives in pharmaceutical research. However, their development is limited by their poor cellular uptake and inefficient trafficking. Here we describe a short amphipathic peptide, Pep-3, that combines a tryptophan/phenylalanine domain with a lysine/arginine-rich hydrophilic motif. Pep-3 forms stable nano-size complexes with peptide-nucleic acid analogues and promotes their efficient delivery into a wide variety of cell lines, including primary and suspension lines, without any associated cytotoxicity. We demonstrate that Pep-3-mediated delivery of antisense-cyclin B1-charged-PNA blocks tumour growth in vivo upon intratumoral and intravenous injection. Moreover, we show that PEGylation of Pep-3 significantly improves complex stability in vivo and consequently the efficiency of antisense cyclin B1 administered intravenously. Given the biological characteristics of these vectors, we believe that peptide-based delivery technologies hold a true promise for therapeutic applications of DNA mimics.
Rapid Bedside Inactivation of Ebola Virus for Safe Nucleic Acid Tests

DEFF Research Database (Denmark)

Rosenstierne, Maiken Worsøe; Karlberg, Helen; Bragstad, Karoline

2016-01-01

Rapid bedside inactivation of Ebola virus would be a solution for the safety of medical and technical staff, risk containment, sample transport, and high-throughput or rapid diagnostic testing during an outbreak. We show that the commercially available Magna Pure lysis/binding buffer used...... for nucleic acid extraction inactivates Ebola virus. A rapid bedside inactivation method for nucleic acid tests is obtained by simply adding Magna Pure lysis/binding buffer directly into vacuum blood collection EDTA tubes using a thin needle and syringe prior to sampling. The ready-to-use inactivation vacuum...... tubes are stable for more than 4 months, and Ebola virus RNA is preserved in the Magna Pure lysis/binding buffer for at least 5 weeks independent of the storage temperature. We also show that Ebola virus RNA can be manually extracted from Magna Pure lysis/binding buffer-inactivated samples using...
Isothermal amplification detection of nucleic acids by a double-nicked beacon.

Science.gov (United States)

Shi, Chao; Zhou, Meiling; Pan, Mei; Zhong, Guilin; Ma, Cuiping

2016-03-01

Isothermal and rapid amplification detection of nucleic acids is an important technology in environmental monitoring, foodborne pathogen detection, and point-of-care clinical diagnostics. Here we have developed a novel method of isothermal signal amplification for single-stranded DNA (ssDNA) detection. The ssDNA target could be used as an initiator, coupled with a double-nicked molecular beacon, to originate amplification cycles, achieving cascade signal amplification. In addition, the method showed good specificity and strong anti-jamming capability. Overall, it is a one-pot and isothermal strand displacement amplification method without the requirement of a stepwise procedure, which greatly simplifies the experimental procedure and decreases the probability of contamination of samples. With its advantages, the method would be very useful to detect nucleic acids in point-of-care or field use. Copyright © 2015 Elsevier Inc. All rights reserved.
Methods of combined bioprocessing and related microorganisms, thermophilic and/or acidophilic enzymes, and nucleic acids encoding said enzymes

Energy Technology Data Exchange (ETDEWEB)

Thompson, David N.; Apel, William A.; Thompson, Vicki S.; Ward, Thomas E.

2017-08-15

A genetically modified organism comprising: at least one nucleic acid sequence and/or at least one recombinant nucleic acid isolated from Alicyclobacillus acidocaldarius and encoding a polypeptide involved in at least partially degrading, cleaving, transporting, metabolizing, or removing polysaccharides, cellulose, lignocellulose, hemicellulose, lignin, starch, sugars, sugar oligomers, carbohydrates, complex carbohydrates, chitin, heteroxylans, glycosides, xylan-, glucan-, galactan-, or mannan-decorating groups; and at least one nucleic acid sequence and/or at least one recombinant nucleic acid encoding a polypeptide involved in fermenting sugar molecules to a product. Additionally, enzymatic and/or proteinaceous extracts may be isolated from one or more genetically modified organisms. The extracts are utilized to convert biomass into a product. Further provided are methods of converting biomass into products comprising: placing the genetically modified organism and/or enzymatic extracts thereof in fluid contact with polysaccharides, cellulose, lignocellulose, hemicellulose, lignin, starch, sugars, sugar oligomers, carbohydrates, complex carbohydrates, chitin, heteroxylans, glycosides, and/or xylan-, glucan-, galactan-, or mannan-decorating groups.
Relationship between growth and total nucleic acids in juvenile pink salmon, Oncorhynchus gorbuscha, fed crude oil contaminated food

International Nuclear Information System (INIS)

Wang Shiao, Y.; Lum, J.L.; Carls, M.G.; Rice, S.D.

1993-01-01

Total nucleic acids of junvenile pink salmon fed crude oil contaminated food were analyzed to deteremine if nucleic acid measurements can be used to evaluate growth of fish collected at oil spill sites. In general, the nucleic acid concentration (μg per mg dry weight) of salmon fry fed food contaminated with either 0.37 or 2.78 mg crude oil/g food was not significantly affected. However, RNA concentration of fry fed food contaminated with 34.83 mg/g was reduced whereas DNA concentration increased. Results over 8 weeks indicate decreased protein synthesis and cell content but maintenance of cell integrity in these fish. Growth was inversely related to the level of crude oil contamination in the food. The significant correlations between measured growth and RNA/DNA ratios and RNA contents (mg RNA per mm fork length) suggest that nucleic acid measurement can be used to compare growth of fish collected from the field. 23 refs., 4 figs
Methods of combined bioprocessing and related microorganisms, thermophilic and/or acidophilic enzymes, and nucleic acids encoding said enzymes

Science.gov (United States)

Thompson, David N; Apel, William A; Thompson, Vicki S; Ward, Thomas E

2013-07-23

A genetically modified organism comprising: at least one nucleic acid sequence and/or at least one recombinant nucleic acid isolated from Alicyclobacillus acidocaldarius and encoding a polypeptide involved in at least partially degrading, cleaving, transporting, metabolizing, or removing polysaccharides, cellulose, lignocellulose, hemicellulose, lignin, starch, sugars, sugar oligomers, carbohydrates, complex carbohydrates, chitin, heteroxylans, glycosides, xylan-, glucan-, galactan-, or mannan-decorating groups; and at least one nucleic acid sequence and/or at least one recombinant nucleic acid encoding a polypeptide involved in fermenting sugar molecules to a product. Additionally, enzymatic and/or proteinaceous extracts may be isolated from one or more genetically modified organisms. The extracts are utilized to convert biomass into a product. Further provided are methods of converting biomass into products comprising: placing the genetically modified organism and/or enzymatic extracts thereof in fluid contact with polysaccharides, cellulose, lignocellulose, hemicellulose, lignin, starch, sugars, sugar oligomers, carbohydrates, complex carbohydrates, chitin, heteroxylans, glycosides, and/or xylan-, glucan-, galactan-, or mannan-decorating groups.
Methods of combined bioprocessing and related microorganisms, thermophilic and/or acidophilic enzymes, and nucleic acids encoding said enzymes

Energy Technology Data Exchange (ETDEWEB)

Thompson, David N.; Apel, William A.; Thompson, Vicki S.; Ward, Thomas E.

2016-03-22

A genetically modified organism comprising: at least one nucleic acid sequence and/or at least one recombinant nucleic acid isolated from Alicyclobacillus acidocaldarius and encoding a polypeptide involved in at least partially degrading, cleaving, transporting, metabolizing, or removing polysaccharides, cellulose, lignocellulose, hemicellulose, lignin, starch, sugars, sugar oligomers, carbohydrates, complex carbohydrates, chitin, heteroxylans, glycosides, xylan-, glucan-, galactan-, or mannan-decorating groups; and at least one nucleic acid sequence and/or at least one recombinant nucleic acid encoding a polypeptide involved in fermenting sugar molecules to a product. Additionally, enzymatic and/or proteinaceous extracts may be isolated from one or more genetically modified organisms. The extracts are utilized to convert biomass into a product. Further provided are methods of converting biomass into products comprising: placing the genetically modified organism and/or enzymatic extracts thereof in fluid contact with polysaccharides, cellulose, lignocellulose, hemicellulose, lignin, starch, sugars, sugar oligomers, carbohydrates, complex carbohydrates, chitin, heteroxylans, glycosides, and/or xylan-, glucan-, galactan-, or mannan-decorating groups.
Hepatitis B virus X protein (HBx)-induced abnormalities of nucleic acid metabolism revealed by (1)H-NMR-based metabonomics.

Science.gov (United States)

Dan Yue; Zhang, Yuwei; Cheng, Liuliu; Ma, Jinhu; Xi, Yufeng; Yang, Liping; Su, Chao; Shao, Bin; Huang, Anliang; Xiang, Rong; Cheng, Ping

2016-04-14

Hepatitis B virus X protein (HBx) plays an important role in HBV-related hepatocarcinogenesis; however, mechanisms underlying HBx-mediated carcinogenesis remain unclear. In this study, an NMR-based metabolomics approach was applied to systematically investigate the effects of HBx on cell metabolism. EdU incorporation assay was conducted to examine the effects of HBx on DNA synthesis, an important feature of nucleic acid metabolism. The results revealed that HBx disrupted metabolism of glucose, lipids, and amino acids, especially nucleic acids. To understand the potential mechanism of HBx-induced abnormalities of nucleic acid metabolism, gene expression profiles of HepG2 cells expressing HBx were investigated. The results showed that 29 genes involved in DNA damage and DNA repair were differentially expressed in HBx-expressing HepG2 cells. HBx-induced DNA damage was further demonstrated by karyotyping, comet assay, Western blotting, immunofluorescence and immunohistochemistry analyses. Many studies have previously reported that DNA damage can induce abnormalities of nucleic acid metabolism. Thus, our results implied that HBx initially induces DNA damage, and then disrupts nucleic acid metabolism, which in turn blocks DNA repair and induces the occurrence of hepatocellular carcinoma (HCC). These findings further contribute to our understanding of the occurrence of HCC.

Single-Labeled Oligonucleotides Showing Fluorescence Changes upon Hybridization with Target Nucleic Acids

Directory of Open Access Journals (Sweden)

Gil Tae Hwang

2018-01-01

Full Text Available Sequence-specific detection of nucleic acids has been intensively studied in the field of molecular diagnostics. In particular, the detection and analysis of single-nucleotide polymorphisms (SNPs is crucial for the identification of disease-causing genes and diagnosis of diseases. Sequence-specific hybridization probes, such as molecular beacons bearing the fluorophore and quencher at both ends of the stem, have been developed to enable DNA mutation detection. Interestingly, DNA mutations can be detected using fluorescently labeled oligonucleotide probes with only one fluorophore. This review summarizes recent research on single-labeled oligonucleotide probes that exhibit fluorescence changes after encountering target nucleic acids, such as guanine-quenching probes, cyanine-containing probes, probes containing a fluorophore-labeled base, and microenvironment-sensitive probes.
First improvements in the detection and quantification of label-free nucleic acids by laser-induced breakdown spectroscopy: Application to the deoxyribonucleic acid micro-array technology

International Nuclear Information System (INIS)

Le Meur, Julien; Menut, Denis; Wodling, Pascal; Salmon, Laurent; Thro, Pierre-Yves; Chevillard, Sylvie; Ugolin, Nicolas

2008-01-01

The accurate quantification of nucleic acids is essential in many fields of modern biology and industry, and in some cases requires the use of fluorescence labeling. Yet, in addition to standardization problems and quantification reproducibility, labeling can modify the physicochemical properties of molecules or affect their stability. To address these limitations, we have developed a novel method to detect and quantify label-free nucleic acids. This method is based on stoichiometric proportioning of phosphorus in the nucleic acid skeleton, using laser-induced breakdown spectroscopy, and a specific statistical analysis, which indicates the error probability for each measurement. The results obtained appear to be quantitative, with a limit of detection of 10 5 nucleotides/μm 2 (i.e. 2 x 10 13 phosphorus atoms/cm 2 ). Initial micro-array analysis has given very encouraging results, which point to new ways of quantifying hybridized nucleic acids. This is essential when comparing molecules of different sequences, which is presently very difficult with fluorescence labeling
First improvements in the detection and quantification of label-free nucleic acids by laser-induced breakdown spectroscopy: Application to the deoxyribonucleic acid micro-array technology

Energy Technology Data Exchange (ETDEWEB)

Le Meur, Julien [Laboratoire de Cancerologie Experimentale, Commissariat a l' Energie Atomique de Fontenay-aux-Roses, Direction des Sciences du Vivant, Departement de Radiobiologie et Radiopathologie, Fontenay-aux-Roses (France); Menut, Denis [Laboratoire de Reactivite des Surfaces et des Interfaces, Commissariat a l' Energie Atomique de Saclay, Direction de l' Energie Nucleaire, Departement de Physico-Chimie, Gif sur Yvette (France); Wodling, Pascal [Laboratoire d' Interaction Laser-Matiere, Commissariat a l' Energie Atomique de Saclay, Direction de l' Energie Nucleaire, Departement de Physico-Chimie, Gif sur Yvette (France); Salmon, Laurent [Laboratoire de Reactivite des Surfaces et des Interfaces, Commissariat a l' Energie Atomique de Saclay, Direction de l' Energie Nucleaire, Departement de Physico-Chimie, Gif sur Yvette (France); Thro, Pierre-Yves [Laboratoire d' Interaction Laser-Matiere, Commissariat a l' Energie Atomique de Saclay, Direction de l' Energie Nucleaire, Departement de Physico-Chimie, Gif sur Yvette (France); Chevillard, Sylvie [Laboratoire de Cancerologie Experimentale, Commissariat a l' Energie Atomique de Fontenay-aux-Roses, Direction des Sciences du Vivant, Departement de Radiobiologie et Radiopathologie, Fontenay-aux-Roses (France); Ugolin, Nicolas [Laboratoire de Cancerologie Experimentale, Commissariat a l' Energie Atomique de Fontenay-aux-Roses, Direction des Sciences du Vivant, Departement de Radiobiologie et Radiopathologie, Fontenay-aux-Roses (France)], E-mail: nugolin@cea.fr

2008-04-15

The accurate quantification of nucleic acids is essential in many fields of modern biology and industry, and in some cases requires the use of fluorescence labeling. Yet, in addition to standardization problems and quantification reproducibility, labeling can modify the physicochemical properties of molecules or affect their stability. To address these limitations, we have developed a novel method to detect and quantify label-free nucleic acids. This method is based on stoichiometric proportioning of phosphorus in the nucleic acid skeleton, using laser-induced breakdown spectroscopy, and a specific statistical analysis, which indicates the error probability for each measurement. The results obtained appear to be quantitative, with a limit of detection of 10{sup 5} nucleotides/{mu}m{sup 2} (i.e. 2 x 10{sup 13} phosphorus atoms/cm{sup 2}). Initial micro-array analysis has given very encouraging results, which point to new ways of quantifying hybridized nucleic acids. This is essential when comparing molecules of different sequences, which is presently very difficult with fluorescence labeling.
The neuroprotective efficacy of cell-penetrating peptides TAT, penetratin, Arg-9, and Pep-1 in glutamic acid, kainic acid, and in vitro ischemia injury models using primary cortical neuronal cultures.

Science.gov (United States)

Meloni, Bruno P; Craig, Amanda J; Milech, Nadia; Hopkins, Richard M; Watt, Paul M; Knuckey, Neville W

2014-03-01

Cell-penetrating peptides (CPPs) are small peptides (typically 5-25 amino acids), which are used to facilitate the delivery of normally non-permeable cargos such as other peptides, proteins, nucleic acids, or drugs into cells. However, several recent studies have demonstrated that the TAT CPP has neuroprotective properties. Therefore, in this study, we assessed the TAT and three other CPPs (penetratin, Arg-9, Pep-1) for their neuroprotective properties in cortical neuronal cultures following exposure to glutamic acid, kainic acid, or in vitro ischemia (oxygen-glucose deprivation). Arg-9, penetratin, and TAT-D displayed consistent and high level neuroprotective activity in both the glutamic acid (IC50: 0.78, 3.4, 13.9 μM) and kainic acid (IC50: 0.81, 2.0, 6.2 μM) injury models, while Pep-1 was ineffective. The TAT-D isoform displayed similar efficacy to the TAT-L isoform in the glutamic acid model. Interestingly, Arg-9 was the only CPP that displayed efficacy when washed-out prior to glutamic acid exposure. Neuroprotection following in vitro ischemia was more variable with all peptides providing some level of neuroprotection (IC50; Arg-9: 6.0 μM, TAT-D: 7.1 μM, penetratin/Pep-1: >10 μM). The positive control peptides JNKI-1D-TAT (JNK inhibitory peptide) and/or PYC36L-TAT (AP-1 inhibitory peptide) were neuroprotective in all models. Finally, in a post-glutamic acid treatment experiment, Arg-9 was highly effective when added immediately after, and mildly effective when added 15 min post-insult, while the JNKI-1D-TAT control peptide was ineffective when added post-insult. These findings demonstrate that different CPPs have the ability to inhibit neurodamaging events/pathways associated with excitotoxic and ischemic injuries. More importantly, they highlight the need to interpret neuroprotection studies when using CPPs as delivery agents with caution. On a positive note, the cytoprotective properties of CPPs suggests they are ideal carrier molecules to
A non-covalent peptide-based strategy for ex vivo and in vivo oligonucleotide delivery.

Science.gov (United States)

Crombez, Laurence; Morris, May C; Heitz, Frederic; Divita, Gilles

2011-01-01

The dramatic acceleration in identification of new nucleic acid-based therapeutic molecules such as short interfering RNA (siRNA) and peptide-nucleic acid (PNA) analogues has provided new perspectives for therapeutic targeting of specific genes responsible for pathological disorders. However, the poor cellular uptake of nucleic acids together with the low permeability of the cell membrane to negatively charged molecules remain major obstacles to their clinical development. Several non-viral strategies have been proposed to improve the delivery of synthetic short oligonucleotides both in cultured cells and in vivo. Cell-penetrating peptides constitute very promising tools for non-invasive cellular import of oligonucleotides and analogs. We recently described a non-covalent strategy based on short amphiphatic peptides (MPG8/PEP3) that have been successfully applied ex vivo and in vivo for the delivery of therapeutic siRNA and PNA molecules. PEP3 and MPG8 form stable nanoparticles with PNA analogues and siRNA, respectively, and promote their efficient cellular uptake, independently of the endosomal pathway, into a wide variety of cell lines, including primary and suspension lines, without any associated cytotoxicity. This chapter describes easy-to-handle protocols for the use of MPG-8 or PEP-3-nanoparticle technologies for PNA and siRNA delivery into adherent and suspension cell lines as well as in vivo into cancer mouse models.
Polyvinyl-alcohol-based magnetic beads for rapid and efficient separation of specific or unspecific nucleic acid sequences

International Nuclear Information System (INIS)

Oster, J.; Parker, Jeffrey; Brassard, Lothar

2001-01-01

The versatile application of polyvinyl-alcohol-based magnetic M-PVA beads is demonstrated in the separation of genomic DNA, sequence specific nucleic acid purification, and binding of bacteria for subsequent DNA extraction and detection. It is shown that nucleic acids can be obtained in high yield and purity using M-PVA beads, making sample preparation efficient, fast and highly adaptable for automation processes
"Customizable" units in di- and tripeptides: selective conversion into substituted dehydroamino acids.

Science.gov (United States)

Saavedra, Carlos J; Boto, Alicia; Hernández, Rosendo

2012-07-20

The selective conversion of serine or threonine units of di- and tripeptides into substituted dehydroamino acids is reported. Thus, these common α-amino acids undergo a scission-phosphorylation process to give α-amino phosphonate residues. A Horner-Wadsworth-Emmons reaction with aldehydes or ketones follows to afford the final products with excellent Z-stereoselectivity (Z:E > 98:2). In this way, a single peptide precursor can selectively be transformed into a variety of derivatives.
Phosphonic Acids for Interfacial Engineering of Transparent Conductive Oxides

KAUST Repository

Paniagua, Sergio A.; Giordano, Anthony J.; Smith, O’ Neil L.; Barlow, Stephen; Li, Hong; Armstrong, Neal R.; Pemberton, Jeanne E.; Bredas, Jean-Luc; Ginger, David; Marder, Seth R.

2016-01-01

Transparent conducting oxides (TCOs), such as indium tin oxide and zinc oxide, play an important role as electrode materials in organic-semiconductor devices. The properties of the inorganic-organic interface - the offset between the TCO Fermi level and the relevant transport level, the extent to which the organic semiconductor can wet the oxide surface, and the influence of the surface on semiconductor morphology - significantly affect device performance. This review surveys the literature on TCO modification with phosphonic acids (PAs), which has increasingly been used to engineer these interfacial properties. The first part outlines the relevance of TCO surface modification to organic electronics, surveys methods for the synthesis of PAs, discusses the modes by which they can bind to TCO surfaces, and compares PAs to alternative organic surface modifiers. The next section discusses methods of PA monolayer deposition, the kinetics of monolayer formation, and structural evidence regarding molecular orientation on TCOs. The next sections discuss TCO work-function modification using PAs, tuning of TCO surface energy using PAs, and initiation of polymerizations from TCO-tethered PAs. Finally, studies that examine the use of PA-modified TCOs in organic light-emitting diodes and organic photovoltaics are compared. © 2016 American Chemical Society.
Phosphonic Acids for Interfacial Engineering of Transparent Conductive Oxides

KAUST Repository

Paniagua, Sergio A.

2016-05-26

Transparent conducting oxides (TCOs), such as indium tin oxide and zinc oxide, play an important role as electrode materials in organic-semiconductor devices. The properties of the inorganic-organic interface - the offset between the TCO Fermi level and the relevant transport level, the extent to which the organic semiconductor can wet the oxide surface, and the influence of the surface on semiconductor morphology - significantly affect device performance. This review surveys the literature on TCO modification with phosphonic acids (PAs), which has increasingly been used to engineer these interfacial properties. The first part outlines the relevance of TCO surface modification to organic electronics, surveys methods for the synthesis of PAs, discusses the modes by which they can bind to TCO surfaces, and compares PAs to alternative organic surface modifiers. The next section discusses methods of PA monolayer deposition, the kinetics of monolayer formation, and structural evidence regarding molecular orientation on TCOs. The next sections discuss TCO work-function modification using PAs, tuning of TCO surface energy using PAs, and initiation of polymerizations from TCO-tethered PAs. Finally, studies that examine the use of PA-modified TCOs in organic light-emitting diodes and organic photovoltaics are compared. © 2016 American Chemical Society.
Caged molecular beacons: controlling nucleic acid hybridization with light.

Science.gov (United States)

Wang, Chunming; Zhu, Zhi; Song, Yanling; Lin, Hui; Yang, Chaoyong James; Tan, Weihong

2011-05-28

We have constructed a novel class of light-activatable caged molecular beacons (cMBs) that are caged by locking two stems with a photo-labile biomolecular interaction or covalent bond. With the cMBs, the nucleic acid hybridization process can be easily controlled with light, which offers the possibility for a high spatiotemporal resolution study of intracellular mRNAs. © The Royal Society of Chemistry 2011
Effects of ultraviolet and visible radiation on nucleic acids and proteins

International Nuclear Information System (INIS)

Loeber, G.

1977-01-01

Three possible photochemical reaction mechanisms have been discussed which might cause changes in biological materials: 1) Photoreactions induced in that constituents of cell substrates absorbing UV-light by themselves, i.e. heteroaromatic moieties of nucleic acids and proteins. 2) Photoreactions induced in that constituents not belonging to the natural biological system and absorbing UV-light, i.e. furocoumarins or cancer producing hydrocarbons. 3) Photoreactions induced in that proper sensitizer molecules absorbing UV-light or visible light. The latter type of photoreactions consumes molecular oxygen but does not consume sensitizer molecules (photodynamic action). Examples have been given for the three possibilities concerning photochemistry of nucleic acids and proteins. Damages of biopolymers were discussed with respect to their biological consequences. Photodynamic changes in the blood system might be caused either after addition of sensitizers to blood or by sensitizers which are constituents of blood itself, i.e. porphytines. (author)
Copper-Catalyzed Synthesis of Mixed Alkyl Aryl Phosphonates

NARCIS (Netherlands)

Fañanás-Mastral, Martín; Feringa, Ben L

2014-01-01

Copper-catalysis allows the direct oxygenarylation of dialkyl phosphonates with diaryliodonium salts. This novel methodology proceeds with a wide range of phosphonates and phosphoramidates under mild conditions and gives straightforward access to valuable mixed alkyl aryl phosphonates in very good
Electrochemical detection of human papillomavirus DNA type 16 using a pyrrolidinyl peptide nucleic acid probe immobilized on screen-printed carbon electrodes.

Science.gov (United States)

Jampasa, Sakda; Wonsawat, Wanida; Rodthongkum, Nadnudda; Siangproh, Weena; Yanatatsaneejit, Pattamawadee; Vilaivan, Tirayut; Chailapakul, Orawon

2014-04-15

An electrochemical biosensor based on an immobilized anthraquinone-labeled pyrrolidinyl peptide nucleic acid (acpcPNA) probe was successfully developed for the selective detection of human papillomavirus (HPV) type 16 DNA. A 14-mer acpcPNA capture probe was designed to recognize a specific 14 nucleotide region of HPV type 16 L1 gene. The redox-active label anthraquinone (AQ) was covalently attached to the N-terminus of the acpcPNA probe through an amide bond. The probe was immobilized onto a chitosan-modified disposable screen-printed carbon electrode via a C-terminal lysine residue using glutaraldehyde as a cross-linking agent. Hybridization with the target DNA was studied by measuring the electrochemical signal response of the AQ label using square-wave voltammetric analysis. The calibration curve exhibited a linear range between 0.02 and 12.0 µM with a limit of detection and limit of quantitation of 4 and 14 nM, respectively. This DNA sensing platform was successfully applied to detect the HPV type 16 DNA from a PCR amplified (240 bp fragment of the L1 gene) sample derived from the HPV type 16 positive human cancer cell line (SiHa), and failed to detect the HPV-negative c33a cell line. The sensor probe exhibited very high selectivity for the complementary 14 base oligonucleotide over the non-complementary oligonucleotides with sequences derived from HPV types 18, 31 and 33. The proposed sensor provides an inexpensive tool for the early stage detection of HPV type 16, which is an important biomarker for cervical cancer. © 2013 Elsevier B.V. All rights reserved.
An integrated paper-based sample-to-answer biosensor for nucleic acid testing at the point of care.

Science.gov (United States)

Choi, Jane Ru; Hu, Jie; Tang, Ruihua; Gong, Yan; Feng, Shangsheng; Ren, Hui; Wen, Ting; Li, XiuJun; Wan Abas, Wan Abu Bakar; Pingguan-Murphy, Belinda; Xu, Feng

2016-02-07

With advances in point-of-care testing (POCT), lateral flow assays (LFAs) have been explored for nucleic acid detection. However, biological samples generally contain complex compositions and low amounts of target nucleic acids, and currently require laborious off-chip nucleic acid extraction and amplification processes (e.g., tube-based extraction and polymerase chain reaction (PCR)) prior to detection. To the best of our knowledge, even though the integration of DNA extraction and amplification into a paper-based biosensor has been reported, a combination of LFA with the aforementioned steps for simple colorimetric readout has not yet been demonstrated. Here, we demonstrate for the first time an integrated paper-based biosensor incorporating nucleic acid extraction, amplification and visual detection or quantification using a smartphone. A handheld battery-powered heating device was specially developed for nucleic acid amplification in POC settings, which is coupled with this simple assay for rapid target detection. The biosensor can successfully detect Escherichia coli (as a model analyte) in spiked drinking water, milk, blood, and spinach with a detection limit of as low as 10-1000 CFU mL(-1), and Streptococcus pneumonia in clinical blood samples, highlighting its potential use in medical diagnostics, food safety analysis and environmental monitoring. As compared to the lengthy conventional assay, which requires more than 5 hours for the entire sample-to-answer process, it takes about 1 hour for our integrated biosensor. The integrated biosensor holds great potential for detection of various target analytes for wide applications in the near future.
Contributions of the Histidine Side Chain and the N-terminal α-Amino Group to the Binding Thermodynamics of Oligopeptides to Nucleic Acids as a Function of pH

Science.gov (United States)

Ballin, Jeff D.; Prevas, James P.; Ross, Christina R.; Toth, Eric A.; Wilson, Gerald M.; Record, M. Thomas

2010-01-01

Interactions of histidine with nucleic acid phosphates and histidine pKa shifts make important contributions to many protein-nucleic acid binding processes. To characterize these phenomena in simplified systems, we quantified binding of a histidine-containing model peptide HWKK (+NH3-His-Trp-Lys-Lys-NH2) and its lysine analog KWKK (+NH3-Lys-Trp-Lys-Lys-NH2) to a single-stranded RNA model, polyuridylate (polyU), by changes in tryptophan fluorescence as a function of salt concentration and pH. For both HWKK and KWKK, equilibrium binding constants, Kobs, and magnitudes of log-log salt derivatives SKobs ≡ (∂logKobs/∂log[Na+]), decreased with increasing pH in the manner expected for a titration curve model in which deprotonation of the histidine and α-amino groups weakens binding and reduces its salt-dependence. Fully protonated HWKK and KWKK exhibit the same Kobs and SKobs within uncertainty, and these SKobs values are consistent with limiting-law polyelectrolyte theory for +4 cationic oligopeptides binding to single-stranded nucleic acids. The pH-dependence of HWKK binding to polyU provides no evidence for pKa shifts nor any requirement for histidine protonation, in stark contrast to the thermodynamics of coupled protonation often seen for these cationic residues in the context of native protein structure where histidine protonation satisfies specific interactions (e.g., salt-bridge formation) within highly complementary binding interfaces. The absence of pKa shifts in our studies indicates that additional Coulombic interactions across the nonspecific-binding interface between RNA and protonated histidine or the α-amino group are not sufficient to promote proton uptake for these oligopeptides. We present our findings in the context of hydration models for specific versus nonspecific nucleic acid binding. PMID:20108951
A versatile method for the preparation of conjugates of peptides with DNA/PNA/analog by employing chemo-selective click reaction in water

Science.gov (United States)

Gogoi, Khirud; Mane, Meenakshi V.; Kunte, Sunita S.; Kumar, Vaijayanti A.

2007-01-01

The specific 1,3 dipolar Hüisgen cycloaddition reaction known as ‘click-reaction’ between azide and alkyne groups is employed for the synthesis of peptide–oligonucleotide conjugates. The peptide nucleic acids (PNA)/DNA and peptides may be appended either by azide or alkyne groups. The cycloaddition reaction between the azide and alkyne appended substrates allows the synthesis of the desired conjugates in high purity and yields irrespective of the sequence and functional groups on either of the two substrates. The versatile approach could also be employed to generate the conjugates of peptides with thioacetamido nucleic acid (TANA) analog. The click reaction is catalyzed by Cu (I) in either water or in organic medium. In water, ∼3-fold excess of the peptide-alkyne/azide drives the reaction to completion in 2 h with no side products. PMID:17981837
A bench-top automated workstation for nucleic acid isolation from clinical sample types.

Science.gov (United States)

Thakore, Nitu; Garber, Steve; Bueno, Arial; Qu, Peter; Norville, Ryan; Villanueva, Michael; Chandler, Darrell P; Holmberg, Rebecca; Cooney, Christopher G

2018-04-18

Systems that automate extraction of nucleic acid from cells or viruses in complex clinical matrices have tremendous value even in the absence of an integrated downstream detector. We describe our bench-top automated workstation that integrates our previously-reported extraction method - TruTip - with our newly-developed mechanical lysis method. This is the first report of this method for homogenizing viscous and heterogeneous samples and lysing difficult-to-disrupt cells using "MagVor": a rotating magnet that rotates a miniature stir disk amidst glass beads confined inside of a disposable tube. Using this system, we demonstrate automated nucleic acid extraction from methicillin-resistant Staphylococcus aureus (MRSA) in nasopharyngeal aspirate (NPA), influenza A in nasopharyngeal swabs (NPS), human genomic DNA from whole blood, and Mycobacterium tuberculosis in NPA. The automated workstation yields nucleic acid with comparable extraction efficiency to manual protocols, which include commercially-available Qiagen spin column kits, across each of these sample types. This work expands the scope of applications beyond previous reports of TruTip to include difficult-to-disrupt cell types and automates the process, including a method for removal of organics, inside a compact bench-top workstation. Copyright © 2018 Elsevier B.V. All rights reserved.
Silver ions-mediated conformational switch: facile design of structure-controllable nucleic acid probes.

Science.gov (United States)

Wang, Yongxiang; Li, Jishan; Wang, Hao; Jin, Jianyu; Liu, Jinhua; Wang, Kemin; Tan, Weihong; Yang, Ronghua

2010-08-01

Conformationally constraint nucleic acid probes were usually designed by forming an intramolecular duplex based on Watson-Crick hydrogen bonds. The disadvantages of these approaches are the inflexibility and instability in complex environment of the Watson-Crick-based duplex. We report that this hydrogen bonding pattern can be replaced by metal-ligation between specific metal ions and the natural bases. To demonstrate the feasibility of this principle, two linear oligonucleotides and silver ions were examined as models for DNA hybridization assay and adenosine triphosphate detection. The both nucleic acids contain target binding sequences in the middle and cytosine (C)-rich sequences at the lateral portions. The strong interaction between Ag(+) ions and cytosines forms stable C-Ag(+)-C structures, which promises the oligonucleotides to form conformationally constraint formations. In the presence of its target, interaction between the loop sequences and the target unfolds the C-Ag(+)-C structures, and the corresponding probes unfolding can be detected by a change in their fluorescence emission. We discuss the thermodynamic and kinetic opportunities that are provided by using Ag(+) ion complexes instead of traditional Watson-Crick-based duplex. In particular, the intrinsic feature of the metal-ligation motif facilitates the design of functional nucleic acids probes by independently varying the concentration of Ag(+) ions in the medium.
Codes in the codons: construction of a codon/amino acid periodic table and a study of the nature of specific nucleic acid-protein interactions.

Science.gov (United States)

Benyo, B; Biro, J C; Benyo, Z

2004-01-01

The theory of "codon-amino acid coevolution" was first proposed by Woese in 1967. It suggests that there is a stereochemical matching - that is, affinity - between amino acids and certain of the base triplet sequences that code for those amino acids. We have constructed a common periodic table of codons and amino acids, where the nucleic acid table showed perfect axial symmetry for codons and the corresponding amino acid table also displayed periodicity regarding the biochemical properties (charge and hydrophobicity) of the 20 amino acids and the position of the stop signals. The table indicates that the middle (2/sup nd/) amino acid in the codon has a prominent role in determining some of the structural features of the amino acids. The possibility that physical contact between codons and amino acids might exist was tested on restriction enzymes. Many recognition site-like sequences were found in the coding sequences of these enzymes and as many as 73 examples of codon-amino acid co-location were observed in the 7 known 3D structures (December 2003) of endonuclease-nucleic acid complexes. These results indicate that the smallest possible units of specific nucleic acid-protein interaction are indeed the stereochemically compatible codons and amino acids.
A survey of advancements in nucleic acid-based logic gates and computing for applications in biotechnology and biomedicine.

Science.gov (United States)

Wu, Cuichen; Wan, Shuo; Hou, Weijia; Zhang, Liqin; Xu, Jiehua; Cui, Cheng; Wang, Yanyue; Hu, Jun; Tan, Weihong

2015-03-04

Nucleic acid-based logic devices were first introduced in 1994. Since then, science has seen the emergence of new logic systems for mimicking mathematical functions, diagnosing disease and even imitating biological systems. The unique features of nucleic acids, such as facile and high-throughput synthesis, Watson-Crick complementary base pairing, and predictable structures, together with the aid of programming design, have led to the widespread applications of nucleic acids (NA) for logic gate and computing in biotechnology and biomedicine. In this feature article, the development of in vitro NA logic systems will be discussed, as well as the expansion of such systems using various input molecules for potential cellular, or even in vivo, applications.

The effects of borate minerals on the synthesis of nucleic acid bases, amino acids and biogenic carboxylic acids from formamide.

Science.gov (United States)

Saladino, Raffaele; Barontini, Maurizio; Cossetti, Cristina; Di Mauro, Ernesto; Crestini, Claudia

2011-08-01

The thermal condensation of formamide in the presence of mineral borates is reported. The products afforded are precursors of nucleic acids, amino acids derivatives and carboxylic acids. The efficiency and the selectivity of the reaction was studied in relation to the elemental composition of the 18 minerals analyzed. The possibility of synthesizing at the same time building blocks of both genetic and metabolic apparatuses, along with the production of amino acids, highlights the interest of the formamide/borate system in prebiotic chemistry.
Long-term stability assessment of AlGaN/GaN field effect transistors modified with peptides: Device characteristics vs. surface properties

Energy Technology Data Exchange (ETDEWEB)

Rohrbaugh, Nathaniel; Bryan, Isaac; Bryan, Zachary; Collazo, Ramon; Ivanisevic, Albena, E-mail: ivanisevic@ncsu.edu [Department of Materials Science and Engineering, North Carolina State University, Raleigh, NC 27695 (United States)

2015-09-15

AlGaN/GaN Field Effect Transistors (FETs) are promising biosensing devices. Functionalization of these devices is explored in this study using an in situ approach with phosphoric acid etchant and a phosphonic acid derivative. Devices are terminated on peptides and soaked in water for up to 168 hrs to examine FETs for both device responses and surface chemistry changes. Measurements demonstrated threshold voltage shifting after the functionalization and soaking processes, but demonstrated stable FET behavior throughout. X-ray photoelectron spectroscopy and atomic force microscopy confirmed peptides attachment to device surfaces before and after water soaking. Results of this work point to the stability of peptide coated functionalized AlGaN/GaN devices in solution and support further research of these devices as disposable, long term, in situ biosensors.
Isolation of nucleic acids and cultures from fossil ice and permafrost

DEFF Research Database (Denmark)

Willerslev, E.; Hansen, Anders J.; Poinar, H. N.

2004-01-01

Owing to their constant low temperatures, glacial ice and permafrost might contain the oldest nucleic acids and microbial cells on Earth, which could prove key to reconstructing past ecosystems and for the planning of missions to other planets. However, recent claims concerning viable cells and m...
Sodium sulphite inhibition of potato and cherry polyphenolics in nucleic acid extraction for virus detection by RT-PCR.

Science.gov (United States)

Singh, R P; Nie, X; Singh, M; Coffin, R; Duplessis, P

2002-01-01

Phenolic compounds from plant tissues inhibit reverse transcription-polymerase chain reaction (RT-PCR). Multiple-step protocols using several additives to inhibit polyphenolic compounds during nucleic acid extraction are common, but time consuming and laborious. The current research highlights that the inclusion of 0.65 to 0.70% of sodium sulphite in the extraction buffer minimizes the pigmentation of nucleic acid extracts and improves the RT-PCR detection of Potato virus Y (PVY) and Potato leafroll virus (PLRV) in potato (Solanum tuberosum) tubers and Prune dwarf virus (PDV) and Prunus necrotic ringspot virus (PNRSV) in leaves and bark in the sweet cherry (Prunus avium) tree. Substituting sodium sulphite in the nucleic acid extraction buffer eliminated the use of proteinase K during extraction. Reagents phosphate buffered saline (PBS)-Tween 20 and polyvinylpyrrolidone (PVP) were also no longer required during RT or PCR phase. The resultant nucleic acid extracts were suitable for both duplex and multiplex RT-PCR. This simple and less expensive nucleic acid extraction protocol has proved very effective for potato cv. Russet Norkotah, which contains a high amount of polyphenolics. Comparing commercially available RNA extraction kits (Catrimox and RNeasy), the sodium sulphite based extraction protocol yielded two to three times higher amounts of RNA, while maintaining comparable virus detection by RT-PCR. The sodium sulphite based extraction protocol was equally effective in potato tubers, and in leaves and bark from the cherry tree.
Experimental Warming Decreases the Average Size and Nucleic Acid Content of Marine Bacterial Communities

KAUST Repository

Huete-Stauffer, Tamara M.

2016-05-23

Organism size reduction with increasing temperature has been suggested as a universal response to global warming. Since genome size is usually correlated to cell size, reduction of genome size in unicells could be a parallel outcome of warming at ecological and evolutionary time scales. In this study, the short-term response of cell size and nucleic acid content of coastal marine prokaryotic communities to temperature was studied over a full annual cycle at a NE Atlantic temperate site. We used flow cytometry and experimental warming incubations, spanning a 6°C range, to analyze the hypothesized reduction with temperature in the size of the widespread flow cytometric bacterial groups of high and low nucleic acid content (HNA and LNA bacteria, respectively). Our results showed decreases in size in response to experimental warming, which were more marked in 0.8 μm pre-filtered treatment rather than in the whole community treatment, thus excluding the role of protistan grazers in our findings. Interestingly, a significant effect of temperature on reducing the average nucleic acid content (NAC) of prokaryotic cells in the communities was also observed. Cell size and nucleic acid decrease with temperature were correlated, showing a common mean decrease of 0.4% per °C. The usually larger HNA bacteria consistently showed a greater reduction in cell and NAC compared with their LNA counterparts, especially during the spring phytoplankton bloom period associated to maximum bacterial growth rates in response to nutrient availability. Our results show that the already smallest planktonic microbes, yet with key roles in global biogeochemical cycling, are likely undergoing important structural shrinkage in response to rising temperatures.
Experimental Warming Decreases the Average Size and Nucleic Acid Content of Marine Bacterial Communities

KAUST Repository

Huete-Stauffer, Tamara M.; Arandia-Gorostidi, Nestor; Alonso-Sá ez, Laura; Moran, Xose Anxelu G.

2016-01-01

Organism size reduction with increasing temperature has been suggested as a universal response to global warming. Since genome size is usually correlated to cell size, reduction of genome size in unicells could be a parallel outcome of warming at ecological and evolutionary time scales. In this study, the short-term response of cell size and nucleic acid content of coastal marine prokaryotic communities to temperature was studied over a full annual cycle at a NE Atlantic temperate site. We used flow cytometry and experimental warming incubations, spanning a 6°C range, to analyze the hypothesized reduction with temperature in the size of the widespread flow cytometric bacterial groups of high and low nucleic acid content (HNA and LNA bacteria, respectively). Our results showed decreases in size in response to experimental warming, which were more marked in 0.8 μm pre-filtered treatment rather than in the whole community treatment, thus excluding the role of protistan grazers in our findings. Interestingly, a significant effect of temperature on reducing the average nucleic acid content (NAC) of prokaryotic cells in the communities was also observed. Cell size and nucleic acid decrease with temperature were correlated, showing a common mean decrease of 0.4% per °C. The usually larger HNA bacteria consistently showed a greater reduction in cell and NAC compared with their LNA counterparts, especially during the spring phytoplankton bloom period associated to maximum bacterial growth rates in response to nutrient availability. Our results show that the already smallest planktonic microbes, yet with key roles in global biogeochemical cycling, are likely undergoing important structural shrinkage in response to rising temperatures.
FTA Cards for Preservation of Nucleic Acids for Molecular Assays: A Review on the Use of Cytologic/Tissue Samples.

Science.gov (United States)

da Cunha Santos, Gilda

2018-03-01

- Traditional methods for storing histologic and cytologic specimens for future use in molecular assays have consisted of either snap-freezing with cryopreservation or formalin-fixing, paraffin-embedding the samples. Although snap-freezing with cryopreservation is recommended for better preservation of nucleic acids, the infrastructure and space required for archiving impose challenges for high-volume pathology laboratories. Cost-effective, long-term storage at room temperature; relatively easy shipment; and standardized handling can be achieved with formalin-fixed, paraffin-embedded samples, but formalin fixation induces fragmentation and chemical modification of nucleic acids. Advances in next-generation sequencing platforms, coupled with an increase in diagnostic, prognostic, and predictive molecular biomarkers have created a demand for high-quality nucleic acids. To address issues of the quality of nucleic acid and logistics in sample acquisition, alternatives for specimen preservation and long-term storage have been described and include novel universal tissue fixatives, stabilizers, and technologies. - To collect, retrieve, and review information from studies describing the use of nucleic acids recovered from cytologic/tissue specimens stored on Flinders Technology Associates (FTA, GE Whatman, Maidstone, Kent, United Kingdom) cards for downstream molecular applications. - An electronic literature search in the PubMed (National Center for Biotechnology Information, Bethesda, Maryland) database allowed the selection of manuscripts addressing the use of FTA cards for storage of cytologic samples for molecular analysis. Only articles published in English were retrieved. - The use of FTA cards is a versatile method for fostering multicenter, international collaborations and clinical trials that require centralized testing, long-distance shipment, and high-quality nucleic acids for molecular techniques. Studies with controlled temperature are required to test the
Mapping photothermally induced gene expression in living cells and tissues by nanorod-locked nucleic acid complexes.

Science.gov (United States)

Riahi, Reza; Wang, Shue; Long, Min; Li, Na; Chiou, Pei-Yu; Zhang, Donna D; Wong, Pak Kin

2014-04-22

The photothermal effect of plasmonic nanostructures has numerous applications, such as cancer therapy, photonic gene circuit, large cargo delivery, and nanostructure-enhanced laser tweezers. The photothermal operation can also induce unwanted physical and biochemical effects, which potentially alter the cell behaviors. However, there is a lack of techniques for characterizing the dynamic cell responses near the site of photothermal operation with high spatiotemporal resolution. In this work, we show that the incorporation of locked nucleic acid probes with gold nanorods allows photothermal manipulation and real-time monitoring of gene expression near the area of irradiation in living cells and animal tissues. The multimodal gold nanorod serves as an endocytic delivery reagent to transport the probes into the cells, a fluorescence quencher and a binding competitor to detect intracellular mRNA, and a plasmonic photothermal transducer to induce cell ablation. We demonstrate the ability of the gold nanorod-locked nucleic acid complex for detecting the spatiotemporal gene expression in viable cells and tissues and inducing photothermal ablation of single cells. Using the gold nanorod-locked nucleic acid complex, we systematically characterize the dynamic cellular heat shock responses near the site of photothermal operation. The gold nanorod-locked nucleic acid complex enables mapping of intracellular gene expressions and analyzes the photothermal effects of nanostructures toward various biomedical applications.
The pattern recognition molecule deleted in malignant brain tumors 1 (DMBT1) and synthetic mimics inhibit liposomal nucleic acid delivery

DEFF Research Database (Denmark)

Lund Hansen, Pernille; Blaich, Stephanie; End, Caroline

2011-01-01

Liposomal nucleic acid delivery is a preferred option for therapeutic settings. The cellular pattern recognition molecule DMBT1, secreted at high levels in various diseases, and synthetic mimics efficiently inhibit liposomal nucleic acid delivery to human cells. These findings may have relevance...
Nanomedicine-based combination anticancer therapy between nucleic acids and small-molecular drugs.

Science.gov (United States)

Huang, Wei; Chen, Liqing; Kang, Lin; Jin, Mingji; Sun, Ping; Xin, Xin; Gao, Zhonggao; Bae, You Han

2017-06-01

Anticancer therapy has always been a vital challenge for the development of nanomedicine. Repeated single therapeutic agent may lead to undesirable and severe side effects, unbearable toxicity and multidrug resistance due to complex nature of tumor. Nanomedicine-based combination anticancer therapy can synergistically improve antitumor outcomes through multiple-target therapy, decreasing the dose of each therapeutic agent and reducing side effects. There are versatile combinational anticancer strategies such as chemotherapeutic combination, nucleic acid-based co-delivery, intrinsic sensitive and extrinsic stimulus combinational patterns. Based on these combination strategies, various nanocarriers and drug delivery systems were engineered to carry out the efficient co-delivery of combined therapeutic agents for combination anticancer therapy. This review focused on illustrating nanomedicine-based combination anticancer therapy between nucleic acids and small-molecular drugs for synergistically improving anticancer efficacy. Copyright © 2017 Elsevier B.V. All rights reserved.
Clustered Regularly Interspaced Short Palindromic Repeats/Cas9 Triggered Isothermal Amplification for Site-Specific Nucleic Acid Detection.

Science.gov (United States)

Huang, Mengqi; Zhou, Xiaoming; Wang, Huiying; Xing, Da

2018-02-06

A novel CRISPR/Cas9 triggered isothermal exponential amplification reaction (CAS-EXPAR) strategy based on CRISPR/Cas9 cleavage and nicking endonuclease (NEase) mediated nucleic acids amplification was developed for rapid and site-specific nucleic acid detection. CAS-EXPAR was primed by the target DNA fragment produced by cleavage of CRISPR/Cas9, and the amplification reaction performed cyclically to generate a large number of DNA replicates which were detected using a real-time fluorescence monitoring method. This strategy that combines the advantages of CRISPR/Cas9 and exponential amplification showed high specificity as well as rapid amplification kinetics. Unlike conventional nucleic acids amplification reactions, CAS-EXPAR does not require exogenous primers, which often cause target-independent amplification. Instead, primers are first generated by Cas9/sgRNA directed site-specific cleavage of target and accumulated during the reaction. It was demonstrated this strategy gave a detection limit of 0.82 amol and showed excellent specificity in discriminating single-base mismatch. Moreover, the applicability of this method to detect DNA methylation and L. monocytogenes total RNA was also verified. Therefore, CAS-EXPAR may provide a new paradigm for efficient nucleic acid amplification and hold the potential for molecular diagnostic applications.
THE VARIATION OF NUCLEIC ACIDS CONTENT AFTER SIMAZIN TREATMENT ON VICIA SATIVA L.

Directory of Open Access Journals (Sweden)

Odeta Grama-Tiganasu

2005-08-01

Full Text Available Simazin has in certain conditions stimulatory effects on nucleic acids biosynthese. The biosyntese and mitotic division stimulation sugest the possibility to use simazin like growing and germination stimulator.
Intumescent features of nucleic acids and proteins

Energy Technology Data Exchange (ETDEWEB)

Alongi, Jenny, E-mail: jenny.alongi@polito.it; Cuttica, Fabio; Blasio, Alessandro Di; Carosio, Federico; Malucelli, Giulio

2014-09-10

Highlights: • The combustion resistance of DNA and caseins to different heat fluxes was studied. • Upon heating, DNA and caseins exhibited an intumescent behaviour. • The char derived from DNA was more stable and coherent than that from caseins. - Abstract: Are nucleic acids and proteins intumescent molecules? In order to get an answer, in the present manuscript, powders of deoxyribose nucleic acids (DNA) and caseins have been exposed to different heat fluxes under a cone calorimeter source and to the direct application of a propane flame. Under these conditions, DNA and caseins exhibited a typical intumescent behaviour, generating a coherent expanded cellular carbonaceous residue (char), extremely resistant to heat exposure. The resulting volumetric expansion as well as the resistance of the formed char turned out to be dependent on (i) the chemical structure of the chosen biomacromolecule, (ii) the evolution of ammonia and (iii) the adopted heat flux in cone calorimetry tests (namely, 25, 35, 50 and 75 kW/m{sup 2}). The presence of ribose units within the DNA backbone determined the formation of highly expanded and coherent residues as compared to those obtained from caseins. Indeed, under a heat flux of 35 kW/m{sup 2}, when a carbon source (i.e. common cane sugar) was added to caseins, the resulting char was similar to that formed by DNA. Furthermore, the char expansion was ascribed to the evolution of ammonia released by these biomacromolecules upon heating, as detected by thermogravimetry coupled to infrared spectroscopy, and confirmed by scanning electron microscopy experiments performed on the bubbles present in the residues of flammability tests.
Intumescent features of nucleic acids and proteins

International Nuclear Information System (INIS)

Alongi, Jenny; Cuttica, Fabio; Blasio, Alessandro Di; Carosio, Federico; Malucelli, Giulio

2014-01-01

Highlights: • The combustion resistance of DNA and caseins to different heat fluxes was studied. • Upon heating, DNA and caseins exhibited an intumescent behaviour. • The char derived from DNA was more stable and coherent than that from caseins. - Abstract: Are nucleic acids and proteins intumescent molecules? In order to get an answer, in the present manuscript, powders of deoxyribose nucleic acids (DNA) and caseins have been exposed to different heat fluxes under a cone calorimeter source and to the direct application of a propane flame. Under these conditions, DNA and caseins exhibited a typical intumescent behaviour, generating a coherent expanded cellular carbonaceous residue (char), extremely resistant to heat exposure. The resulting volumetric expansion as well as the resistance of the formed char turned out to be dependent on (i) the chemical structure of the chosen biomacromolecule, (ii) the evolution of ammonia and (iii) the adopted heat flux in cone calorimetry tests (namely, 25, 35, 50 and 75 kW/m 2 ). The presence of ribose units within the DNA backbone determined the formation of highly expanded and coherent residues as compared to those obtained from caseins. Indeed, under a heat flux of 35 kW/m 2 , when a carbon source (i.e. common cane sugar) was added to caseins, the resulting char was similar to that formed by DNA. Furthermore, the char expansion was ascribed to the evolution of ammonia released by these biomacromolecules upon heating, as detected by thermogravimetry coupled to infrared spectroscopy, and confirmed by scanning electron microscopy experiments performed on the bubbles present in the residues of flammability tests
Computational Approaches to Nucleic Acid Origami.

Science.gov (United States)

Jabbari, Hosna; Aminpour, Maral; Montemagno, Carlo

2015-10-12

Recent advances in experimental DNA origami have dramatically expanded the horizon of DNA nanotechnology. Complex 3D suprastructures have been designed and developed using DNA origami with applications in biomaterial science, nanomedicine, nanorobotics, and molecular computation. Ribonucleic acid (RNA) origami has recently been realized as a new approach. Similar to DNA, RNA molecules can be designed to form complex 3D structures through complementary base pairings. RNA origami structures are, however, more compact and more thermodynamically stable due to RNA's non-canonical base pairing and tertiary interactions. With all these advantages, the development of RNA origami lags behind DNA origami by a large gap. Furthermore, although computational methods have proven to be effective in designing DNA and RNA origami structures and in their evaluation, advances in computational nucleic acid origami is even more limited. In this paper, we review major milestones in experimental and computational DNA and RNA origami and present current challenges in these fields. We believe collaboration between experimental nanotechnologists and computer scientists are critical for advancing these new research paradigms.
Preparative concentration of nucleic acids fragments by capillary isotachophoretic analyzer

Czech Academy of Sciences Publication Activity Database

Datinská, Vladimíra; Voráčová, Ivona; Berka, J.; Foret, František

2018-01-01

Roč. 1548 (2018), s. 100-103 ISSN 0021-9673 R&D Projects: GA MŠk(CZ) LQ1601; GA ČR(CZ) GBP206/12/G014; GA MŠk(CZ) 8F17003 Institutional support: RVO:68081715 Keywords : DNA * isotachophoresis * nucleic acids * sample preparation Subject RIV: CB - Analytical Chemistry, Separation OBOR OECD: Analytical chemistry Impact factor: 3.981, year: 2016
Preparative concentration of nucleic acids fragments by capillary isotachophoretic analyzer

Czech Academy of Sciences Publication Activity Database

Datinská, Vladimíra; Voráčová, Ivona; Berka, J.; Foret, František

2018-01-01

Roč. 1548, MAY (2018), s. 100-103 ISSN 0021-9673 R&D Projects: GA MŠk(CZ) LQ1601; GA ČR(CZ) GBP206/12/G014; GA MŠk(CZ) 8F17003 Institutional support: RVO:68081715 Keywords : DNA * isotachophoresis * nucleic acids * sample preparation Subject RIV: CB - Analytical Chemistry, Separation OBOR OECD: Analytical chemistry Impact factor: 3.981, year: 2016
Glutamic Acid Selective Chemical Cleavage of Peptide Bonds.

Science.gov (United States)

Nalbone, Joseph M; Lahankar, Neelam; Buissereth, Lyssa; Raj, Monika

2016-03-04

Site-specific hydrolysis of peptide bonds at glutamic acid under neutral aqueous conditions is reported. The method relies on the activation of the backbone amide chain at glutamic acid by the formation of a pyroglutamyl (pGlu) imide moiety. This activation increases the susceptibility of a peptide bond toward hydrolysis. The method is highly specific and demonstrates broad substrate scope including cleavage of various bioactive peptides with unnatural amino acid residues, which are unsuitable substrates for enzymatic hydrolysis.
Removal of phosphonates from industrial wastewater with UV/FeII, Fenton and UV/Fenton treatment.

Science.gov (United States)

Rott, Eduard; Minke, Ralf; Bali, Ulusoy; Steinmetz, Heidrun

2017-10-01

Phosphonates are an important group of phosphorus-containing compounds due to their increasing industrial use and possible eutrophication potential. This study involves investigations into the methods UV/Fe II , Fenton and UV/Fenton for their removal from a pure water matrix and industrial wastewaters. It could be shown that the degradability of phosphonates by UV/Fe II (6 kWh/m 3 ) in pure water crucially depended on the pH and was higher the less phosphonate groups a phosphonate contains. The UV/Fe II method is recommended in particular for the treatment of concentrates with nitrogen-free phosphonates, only little turbidity and a low content of organic compounds. Using Fenton reagent, the degradation of polyphosphonates was relatively weak in a pure water matrix (Fenton method (6 kWh/m 3 ), those phosphonates with the smallest numbers of phosphonate groups were easier degraded as well at pH 3.5 in a pure water matrix (o-PO 4 3- formation rates of up to 80%). Despite an incomplete transformation of organically bound phosphorus to o-PO 4 3- with Fenton reagent in an organically highly polluted wastewater (max. 15%), an almost total removal of the total P occurred. The most efficient total P elimination rates were achieved in accordance with the following Fenton implementation: reaction → sludge separation (acidic) → neutralization of the supernatant → sludge separation (neutral). Accordingly, a neutralization directly after the reaction phase led to a lower total P removal extent. Copyright © 2017 Elsevier Ltd. All rights reserved.
Production of extracellular nucleic acids by genetically altered bacteria in aquatic-environment microcosms

International Nuclear Information System (INIS)

Paul, J.H.; David, A.W.

1989-01-01

The factors which affect the production of extracellular DNA by genetically altered strains of Escherichia coli, Pseudomonas aeruginosa, Pseudomonas cepacia, and Bradyrhizobium japonicum in aquatic environments were investigated. Cellular nucleic acids were labeled in vivo by incubation with [ 3 H]thymidine or [ 3 H]adenine, and production of extracellular DNA in marine waters, artificial seawater, or minimal salts media was determined by detecting radiolabeled macromolecules in incubation filtrates. The presence or absence of the ambient microbial community had little effect on the production of extracellular DNA. Three of four organisms produced the greatest amounts of extracellular nucleic acids when incubated in low-salinity media (2% artificial seawater) rather than high-salinity media (10 to 50% artificial seawater). The greatest production of extracellular nucleic acids by P. cepacia occurred at pH 7 and 37 degree C, suggesting that extracellular-DNA production may be a normal physiologic function of the cell. Incubation of labeled P. cepacia cells in water from Bimini Harbor, Bahamas, resulted in labeling of macromolecules of the ambient microbial population. Collectively these results indicate that (i) extracellular-DNA production by genetically altered bacteria released into aquatic environments is more strongly influenced by physicochemical factors than biotic factors, (ii) extracellular-DNA production rates are usually greater for organisms released in freshwater than marine environments, and (iii) ambient microbial populations can readily utilize materials released by these organisms

Rapid amplification/detection of nucleic acid targets utilizing a HDA/thin film biosensor.

Science.gov (United States)

Jenison, Robert; Jaeckel, Heidi; Klonoski, Joshua; Latorra, David; Wiens, Jacinta

2014-08-07

Thin film biosensors exploit a flat, optically coated silicon-based surface whereupon formation of nucleic acid hybrids are enzymatically transduced in a molecular thin film that can be detected by the unaided human eye under white light. While the limit of sensitivity for detection of nucleic acid targets is at sub-attomole levels (60 000 copies) many clinical specimens containing bacterial pathogens have much lower levels of analyte present. Herein, we describe a platform, termed HDA/thin film biosensor, which performs helicase-dependant nucleic acid amplification on a thin film biosensor surface to improve the limit of sensitivity to 10 copies of the mecA gene present in methicillin-resistant strains of Staphylococcus. As double-stranded DNA is unwound by helicase it was either bound by solution-phase DNA primers to be copied by DNA polymerase or hybridized to surface immobilized probe on the thin film biosensor surface to be detected. Herein, we show that amplification reactions on the thin film biosensor are equivalent to in standard thin wall tubes, with detection at the limit of sensitivity of the assay occurring after 30 minutes of incubation time. Further we validate the approach by detecting the presence of the mecA gene in methicillin-resistant Staphylococcus aureus (MRSA) from positive blood culture aliquots with high specificity (signal/noise ratio of 105).
Sensitvie life detection strategies for low-biomass environments: optimizing extraction of nucleic acids adsorbing to terrestrial and Mars analogue minerals.

NARCIS (Netherlands)

Direito, S.O.L.; Marees, A.; Roling, W.F.M.

2012-01-01

The adsorption of nucleic acids to mineral matrixes can result in low extraction yields and negatively influences molecular microbial ecology studies, in particular for low-biomass environments on Earth and Mars. We determined the recovery of nucleic acids from a range of minerals relevant to Earth
Modulation of Cellular Transcription Factor Activity

DEFF Research Database (Denmark)

2003-01-01

A novel class of compounds, known as peptide nucleic acids, form double-stranded structures with one another and with ssDNA. The peptide nucleic acids generally comprise ligands such as naturally occurring DNA bases attached to a peptide backbone through a suitable linker.......A novel class of compounds, known as peptide nucleic acids, form double-stranded structures with one another and with ssDNA. The peptide nucleic acids generally comprise ligands such as naturally occurring DNA bases attached to a peptide backbone through a suitable linker....
Cation–Anion Interactions within the Nucleic Acid Ion Atmosphere Revealed by Ion Counting

Science.gov (United States)

Gebala, Magdalena; Giambasu, George M.; Lipfert, Jan; Bisaria, Namita; Bonilla, Steve; Li, Guangchao; York, Darrin M.; Herschlag, Daniel

2016-01-01

The ion atmosphere is a critical structural, dynamic, and energetic component of nucleic acids that profoundly affects their interactions with proteins and ligands. Experimental methods that “count” the number of ions thermodynamically associated with the ion atmosphere allow dissection of energetic properties of the ion atmosphere, and thus provide direct comparison to theoretical results. Previous experiments have focused primarily on the cations that are attracted to nucleic acid polyanions, but have also showed that anions are excluded from the ion atmosphere. Herein, we have systematically explored the properties of anion exclusion, testing the zeroth-order model that anions of different identity are equally excluded due to electrostatic repulsion. Using a series of monovalent salts, we find, surprisingly, that the extent of anion exclusion and cation inclusion significantly depends on salt identity. The differences are prominent at higher concentrations and mirror trends in mean activity coefficients of the electrolyte solutions. Salts with lower activity coefficients exhibit greater accumulation of both cations and anions within the ion atmosphere, strongly suggesting that cation–anion correlation effects are present in the ion atmosphere and need to be accounted for to understand electrostatic interactions of nucleic acids. To test whether the effects of cation–anion correlations extend to nucleic acid kinetics and thermodynamics, we followed the folding of P4–P6, a domain of the Tetrahymena group I ribozyme, via single-molecule fluorescence resonance energy transfer in solutions with different salts. Solutions of identical concentration but lower activity gave slower and less favorable folding. Our results reveal hitherto unknown properties of the ion atmosphere and suggest possible roles of oriented ion pairs or anion-bridged cations in the ion atmosphere for electrolyte solutions of salts with reduced activity. Consideration of these new
Comparison of femtosecond laser and continuous wave UV sources for protein-nucleic acid crosslinking.

Science.gov (United States)

Fecko, Christopher J; Munson, Katherine M; Saunders, Abbie; Sun, Guangxing; Begley, Tadhg P; Lis, John T; Webb, Watt W

2007-01-01

Crosslinking proteins to the nucleic acids they bind affords stable access to otherwise transient regulatory interactions. Photochemical crosslinking provides an attractive alternative to formaldehyde-based protocols, but irradiation with conventional UV sources typically yields inadequate product amounts. Crosslinking with pulsed UV lasers has been heralded as a revolutionary technique to increase photochemical yield, but this method had only been tested on a few protein-nucleic acid complexes. To test the generality of the yield enhancement, we have investigated the benefits of using approximately 150 fs UV pulses to crosslink TATA-binding protein, glucocorticoid receptor and heat shock factor to oligonucleotides in vitro. For these proteins, we find that the quantum yields (and saturating yields) for forming crosslinks using the high-peak intensity femtosecond laser do not improve on those obtained with low-intensity continuous wave (CW) UV sources. The photodamage to the oligonucleotides and proteins also has comparable quantum yields. Measurements of the photochemical reaction yields of several small molecules selected to model the crosslinking reactions also exhibit nearly linear dependences on UV intensity instead of the previously predicted quadratic dependence. Unfortunately, these results disprove earlier assertions that femtosecond pulsed laser sources provide significant advantages over CW radiation for protein-nucleic acid crosslinking.
Nucleic acid components from strontium-90 exposed miniature swine

International Nuclear Information System (INIS)

Frazier, M.E.

1976-01-01

The reverse transcriptase associated with porcine type C virus particles was used to generate a tritium-labeled DNA product complementary to the viral RNA template. Results of nucleic acid hybridization experiments indicate this 3 H-DNA (probe) was copied from heteropolymeric regions of the procine viral RNA. Probe prepared from this porcine type C virus contains sequences that possess some homology in sequence to RNA isolated from viruses known to cause similar diseases in other animals
Nucleic acid reactivity : challenges for next-generation semiempirical quantum models

OpenAIRE

Huang, Ming; Giese, Timothy J.; York, Darrin M.

2015-01-01

Semiempirical quantum models are routinely used to study mechanisms of RNA catalysis and phosphoryl transfer reactions using combined quantum mechanical/molecular mechanical methods. Herein, we provide a broad assessment of the performance of existing semiempirical quantum models to describe nucleic acid structure and reactivity in order to quantify their limitations and guide the development of next-generation quantum models with improved accuracy. Neglect of diatomic diffierential overlap (...
Recent advances in H-phosphonate chemistry. Part 1. H-phosphonate esters: synthesis and basic reactions.

Science.gov (United States)

Sobkowski, Michal; Kraszewski, Adam; Stawinski, Jacek

2015-01-01

This review covers recent progress in the preparation of H-phosphonate mono- and diesters, basic studies on mechanistic and stereochemical aspects of this class of phosphorus compounds, and their fundamental chemistry in terms of transformation of P-H bonds into P-heteroatom bonds. Selected recent applications of H-phosphonate derivatives in basic organic phosphorus chemistry and in the synthesis of biologically important phosphorus compounds are also discussed.
Polyethylene glycol and octa-arginine dual-functionalized nanographene oxide: an optimization for efficient nucleic acid delivery.

Science.gov (United States)

Imani, Rana; Prakash, Satya; Vali, Hojatollah; Faghihi, Shahab

2018-05-29

The successful application of nucleic acid-based therapy for the treatment of various cancers is largely dependent on a safe and efficient delivery system. A dual-functionalized graphene oxide (GO)-based nanocarrier with the conjugation of aminated-polyethylene glycol (PEG-diamine) and octa-arginine (R8) for the intracellular delivery of nucleic acids is proposed. The functionalized sites are covalently co-conjugated and the PEG : R8 molar ratio is optimized at 10 : 1 to achieve a hydrocolloidally stable size of 252 ± 2.0 nm with an effective charge of +40.97 ± 1.05 and an amine-rich content of 10.87 ± 0.4 μmol g-1. The uptake of the nanocarrier in breast cancer cell lines, MCF-7 and MDA-MB 231, is investigated. The siRNA and pDNA condensation ability in the presence and absence of enzymes and the endosomal buffering capacity, as well as the intracellular localization of the gene/nanocarrier complex are also evaluated. Furthermore, the delivery of functional genes associated with the nanocarrier is assessed using c-Myc protein knockdown and EGFP gene expression. The effective uptake of the nanocarrier by the cells shows superior cytocompatibility, and protects the siRNA and pDNA against enzyme degradation while inhibiting their migration with N : P ratios of 10 and 5, respectively. The co-conjugation of PEG-diamine and the cationic cell-penetrating peptide (CPP) into the GO nanocarrier also provides a superior internalization efficacy of 85% in comparison with a commercially available transfection reagent. The c-Myc protein knockdown and EGFP expression, which are induced by the nanocarrier, confirm that the optimized PEG-diamine/R8-functionalized GO could effectively deliver pDNA and siRNA into the cells and interfere with gene expression.
A novel glucose biosensor based on phosphonic acid-functionalized silica nanoparticles for sensitive detection of glucose in real samples

International Nuclear Information System (INIS)

Zhao, Wenbo; Fang, Yi; Zhu, Qinshu; Wang, Kuai; Liu, Min; Huang, Xiaohua; Shen, Jian

2013-01-01

An effective strategy for preparation amperometric biosensor by using the phosphonic acid-functionalized silica nanoparticles (PFSi NPs) as special modified materials is proposed. In such a strategy, glucose oxidase (GOD) was selected as model protein to fabricate glucose biosensor in the presence of phosphonic acid-functionalized silica nanoparticles (PFSi NPs). The PFSi NPs were first modified on the surface of glassy carbon (GC) electrode, then, GOD was adsorbed onto the PFSi NPs film by drop-coating. The PFSi NPs were characterized by transmission electron microscopy (TEM) and nuclear magnetic resonance (NMR) spectra. The interaction of PFSi NPs with GOD was investigated by the circular dicroism spectroscopy (CD). The results showed PFSi NPs could essentially maintain the native conformation of GOD. The direct electron transfer of GOD on (PFSi NPs)/GCE electrode exhibited excellent electrocatalytic activity for the oxidation of glucose. The proposed biosensor modified with PFSi NPs displayed a fast amperometric response (5 s) to glucose, a good linear current–time relation over a wide range of glucose concentrations from 5.00 × 10 −4 to 1.87 × 10 −1 M, and a low detection limit of 2.44 × 10 −5 M (S/N = 3). Moreover, the biosensor can be used for assessment of the concentration of glucose in many real samples (relative error < 3%). The GOD biosensor modified with PFSi NPs will have essential meaning and practical application in future that attributed to the simple method of fabrication and good performance
Nucleic acid-based vaccines targeting respiratory syncytial virus: Delivering the goods.

Science.gov (United States)

Smith, Trevor R F; Schultheis, Katherine; Broderick, Kate E

2017-11-02

Respiratory syncytial virus (RSV) is a massive medical burden on a global scale. Infants, children and the elderly represent the vulnerable populations. Currently there is no approved vaccine to protect against the disease. Vaccine development has been hindered by several factors including vaccine enhanced disease (VED) associated with formalin-inactivated RSV vaccines, inability of target populations to raise protective immune responses after vaccination or natural viral infection, and a lack of consensus concerning the most appropriate virus-associated target antigen. However, with recent advances in the molecular understanding of the virus, and design of highly characterized vaccines with enhanced immunogenicity there is new belief a RSV vaccine is possible. One promising approach is nucleic acid-based vaccinology. Both DNA and mRNA RSV vaccines are showing promising results in clinically relevant animal models, supporting their transition into humans. Here we will discuss this strategy to target RSV, and the ongoing studies to advance the nucleic acid vaccine platform as a viable option to protect vulnerable populations from this important disease.
Increasing the Fill Factor of Inverted P3HT:PCBM Solar Cells Through Surface Modification of Al-Doped ZnO via Phosphonic Acid-Anchored C60 SAMs

DEFF Research Database (Denmark)

Stubhan, Tobias; Salinas, Michael; Ebel, Alexander

2012-01-01

The influence of aluminum-doped zinc oxide (AZO) electron extraction layers modified with self-assembled monolayers (SAMs) on inverted polymer solar cells is investigated. It is found that AZO modification with phosphonic acid-anchored Fullerene–SAMs leads to a reduction of the series resistance,...
Oxygen isotope signature of UV degradation of glyphosate and phosphonoacetate: Tracing sources and cycling of phosphonates

Energy Technology Data Exchange (ETDEWEB)

Sandy, Edward H. [State Key Laboratory of Biogeology and Environmental Geology, China University of Geosciences, Wuhan 430074 (China); Department of Geology and Geophysics, Yale University, P.O. Box 208109, New Haven, CT 06520-8109 (United States); Department of Chemistry, University of Sierra Leone, Freetown (Sierra Leone); Blake, Ruth E., E-mail: ruth.blake@yale.edu [Department of Geology and Geophysics, Yale University, P.O. Box 208109, New Haven, CT 06520-8109 (United States); School of Civil and Environmental Engineering, and National “International Cooperation Base on Environment and Energy”, University of Science and Technology Beijing, Beijing 100084 (China); Chang, Sae Jung [Department of Geology and Geophysics, Yale University, P.O. Box 208109, New Haven, CT 06520-8109 (United States); Jun, Yao, E-mail: yaojun@ustb.edu.cn [State Key Laboratory of Biogeology and Environmental Geology, China University of Geosciences, Wuhan 430074 (China); Department of Geology and Geophysics, Yale University, P.O. Box 208109, New Haven, CT 06520-8109 (United States); School of Civil and Environmental Engineering, and National “International Cooperation Base on Environment and Energy”, University of Science and Technology Beijing, Beijing 100084 (China); Yu, Chan [Department of Geology and Geophysics, Yale University, P.O. Box 208109, New Haven, CT 06520-8109 (United States); School of Civil and Environmental Engineering, and National “International Cooperation Base on Environment and Energy”, University of Science and Technology Beijing, Beijing 100084 (China)

2013-09-15

Highlights: • Phosphonate (phon) hydrolysis by UVR (1.2 kW) attained ≥90% completion in 84 h. • Isotope study reveals both ambient H{sub 2}O and O{sub 2} involvements in phon C-P bond cleavage. • Mechanistic models proposed for phon C-P bond cleavage based on O-isotope analysis. • Model equations used to calculate δ{sup 18}O{sub P-org} of original phon P-moiety-useful as a tracer. • Study shows relevance in tracing phon sources and cycling in the environment. -- Abstract: The degradation of phosphonates in the natural environment constitutes a major route by which orthophosphate (Pi) is regenerated from organic phosphorus and recently implicated in marine methane production, with ramifications to environmental pollution issues and global climate change concerns. This work explores the application of stable oxygen isotope analysis in elucidating the C-P bond cleavage mechanism(s) of phosphonates by UV photo-oxidation and for tracing their sources in the environment. The two model phosphonates used, glyphosate and phosphonoacetic acid were effectively degraded after exposure to UV irradiation. The isotope results indicate the involvement of both ambient water and atmospheric oxygen in the C-P bond cleavage and generally consistent with previously posited mechanisms of UV-photon excitation reactions. A model developed to calculate the oxygen isotopic composition of the original phosphonate P-moiety, shows both synthetic phosphonates having distinctly lower values compared to naturally derived organophosphorus compounds. Such mechanistic models, based on O-isotope probing, are useful for tracing the sources and reactions of phosphonates in the environment.
Interactions of hydrated divalent metal cations with nucleic acid bases. How to relate the gas phase data to solution situation and binding selectivity in nucleic acids

Czech Academy of Sciences Publication Activity Database

Šponer, Judit E.; Sychrovský, Vladimír; Hobza, Pavel; Šponer, Jiří

2004-01-01

Roč. 6, č. 10 (2004), s. 2772-2780 ISSN 1463-9076 R&D Projects: GA MŠk LN00A016; GA MŠk LN00A032 Grant - others:Wellcome Trust(GB) GR067507MF Institutional research plan: CEZ:AV0Z5004920 Keywords : nucleic acids * gas phase * guanine Subject RIV: BO - Biophysics Impact factor: 2.076, year: 2004
Evaluation of three automated nucleic acid extraction systems for identification of respiratory viruses in clinical specimens by multiplex real-time PCR.

Science.gov (United States)

Kim, Yoonjung; Han, Mi-Soon; Kim, Juwon; Kwon, Aerin; Lee, Kyung-A

2014-01-01

A total of 84 nasopharyngeal swab specimens were collected from 84 patients. Viral nucleic acid was extracted by three automated extraction systems: QIAcube (Qiagen, Germany), EZ1 Advanced XL (Qiagen), and MICROLAB Nimbus IVD (Hamilton, USA). Fourteen RNA viruses and two DNA viruses were detected using the Anyplex II RV16 Detection kit (Seegene, Republic of Korea). The EZ1 Advanced XL system demonstrated the best analytical sensitivity for all the three viral strains. The nucleic acids extracted by EZ1 Advanced XL showed higher positive rates for virus detection than the others. Meanwhile, the MICROLAB Nimbus IVD system was comprised of fully automated steps from nucleic extraction to PCR setup function that could reduce human errors. For the nucleic acids recovered from nasopharyngeal swab specimens, the QIAcube system showed the fewest false negative results and the best concordance rate, and it may be more suitable for detecting various viruses including RNA and DNA virus strains. Each system showed different sensitivity and specificity for detection of certain viral pathogens and demonstrated different characteristics such as turnaround time and sample capacity. Therefore, these factors should be considered when new nucleic acid extraction systems are introduced to the laboratory.
Prolinol-based nucleoside phosphonic acids: new isosteric conformationally flexible nucleotide analogues

Czech Academy of Sciences Publication Activity Database

Vaněk, Václav; Buděšínský, Miloš; Rinnová, Markéta; Rosenberg, Ivan

2009-01-01

Roč. 65, č. 4 (2009), s. 862-876 ISSN 0040-4020 R&D Projects: GA ČR GA203/05/0827; GA MŠk(CZ) LC06077; GA MŠk LC512; GA MŠk(CZ) LC06061 Institutional research plan: CEZ:AV0Z40550506 Keywords : nucleotide analogues * phosphonates * prolinol derivatives * N- alkylation Subject RIV: CC - Organic Chemistry Impact factor: 3.219, year: 2009
Paper-based solid-phase nucleic acid hybridization assay using immobilized quantum dots as donors in fluorescence resonance energy transfer.

Science.gov (United States)

Noor, M Omair; Shahmuradyan, Anna; Krull, Ulrich J

2013-02-05

A paper-based solid-phase assay is presented for transduction of nucleic acid hybridization using immobilized quantum dots (QDs) as donors in fluorescence resonance energy transfer (FRET). The surface of paper was modified with imidazole groups to immobilize QD-probe oligonucleotide conjugates that were assembled in solution. Green-emitting QDs (gQDs) were FRET-paired with Cy3 acceptor. Hybridization of Cy3-labeled oligonucleotide targets provided the proximity required for FRET-sensitized emission from Cy3, which served as an analytical signal. The assay exhibited rapid transduction of nucleic acid hybridization within minutes. Without any amplification steps, the limit of detection of the assay was found to be 300 fmol with the upper limit of the dynamic range at 5 pmol. The implementation of glutathione-coated QDs for the development of nucleic acid hybridization assay integrated on a paper-based platform exhibited excellent resistance to nonspecific adsorption of oligonucleotides and showed no reduction in the performance of the assay in the presence of large quantities of noncomplementary DNA. The selectivity of nucleic acid hybridization was demonstrated by single-nucleotide polymorphism (SNP) detection at a contrast ratio of 19 to 1. The reuse of paper over multiple cycles of hybridization and dehybridization was possible, with less than 20% reduction in the performance of the assay in five cycles. This work provides an important framework for the development of paper-based solid-phase QD-FRET nucleic acid hybridization assays that make use of a ratiometric approach for detection and analysis.
RBscore&NBench: a high-level web server for nucleic acid binding residues prediction with a large-scale benchmarking database.

Science.gov (United States)

Miao, Zhichao; Westhof, Eric

2016-07-08

RBscore&NBench combines a web server, RBscore and a database, NBench. RBscore predicts RNA-/DNA-binding residues in proteins and visualizes the prediction scores and features on protein structures. The scoring scheme of RBscore directly links feature values to nucleic acid binding probabilities and illustrates the nucleic acid binding energy funnel on the protein surface. To avoid dataset, binding site definition and assessment metric biases, we compared RBscore with 18 web servers and 3 stand-alone programs on 41 datasets, which demonstrated the high and stable accuracy of RBscore. A comprehensive comparison led us to develop a benchmark database named NBench. The web server is available on: http://ahsoka.u-strasbg.fr/rbscorenbench/. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.
BIGNASim: a NoSQL database structure and analysis portal for nucleic acids simulation data.

Science.gov (United States)

Hospital, Adam; Andrio, Pau; Cugnasco, Cesare; Codo, Laia; Becerra, Yolanda; Dans, Pablo D; Battistini, Federica; Torres, Jordi; Goñi, Ramón; Orozco, Modesto; Gelpí, Josep Ll

2016-01-04

Molecular dynamics simulation (MD) is, just behind genomics, the bioinformatics tool that generates the largest amounts of data, and that is using the largest amount of CPU time in supercomputing centres. MD trajectories are obtained after months of calculations, analysed in situ, and in practice forgotten. Several projects to generate stable trajectory databases have been developed for proteins, but no equivalence exists in the nucleic acids world. We present here a novel database system to store MD trajectories and analyses of nucleic acids. The initial data set available consists mainly of the benchmark of the new molecular dynamics force-field, parmBSC1. It contains 156 simulations, with over 120 μs of total simulation time. A deposition protocol is available to accept the submission of new trajectory data. The database is based on the combination of two NoSQL engines, Cassandra for storing trajectories and MongoDB to store analysis results and simulation metadata. The analyses available include backbone geometries, helical analysis, NMR observables and a variety of mechanical analyses. Individual trajectories and combined meta-trajectories can be downloaded from the portal. The system is accessible through http://mmb.irbbarcelona.org/BIGNASim/. Supplementary Material is also available on-line at http://mmb.irbbarcelona.org/BIGNASim/SuppMaterial/. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.
[MEASUREMENT OF HISTONES AND CIRCULATING EXTRACELLULAR NUCLEIC ACIDS IN PATIENTS' WITH COMPLICATED FORMS OF PEPTIC ULCER].

Science.gov (United States)

Yerznkyan, G; Kultanov, B; Shakeev, K; Tatina, Ye

2017-04-01

We studied 135 people (24 people, apparently healthy, 39 uncomplicated peptic ulcer disease, 42 people with complex forms peptic ulcer, 30 and after the treatment of complicated forms of peptic ulcer disease, both sexes (18-45 y.). In all patients, the diagnosis was confirmed fibrogastroduodenoscopy (EGD). Determination of histones and acid soluble fraction (ASF), RNA, DNA, in blood was performed by the method of L. Markusheva. Studies have led to the conclusion that the change in the blood concentration of extracellular nucleic acids in patients with uncomplicated disease and complex shapes can be caused by oxidative stress products and can be a signal for elimination of nucleic acids from cells. We have registered various dynamics of the studied parameters histones in the blood of patients with various forms of peptic ulcer disease, which reflects the degree of metabolic abnormalities that occur in the body, associated with changes in the structure of the nucleus. According to the results of our research in the study of the role of extracellular nucleic acids, histones to assess the extent of violations of metabolic processes at a peptic ulcer, complicated and uncomplicated form, the obtained results can be used as predictors of complications of a stomach ulcer.

Advances in nucleic acid-based diagnostics of bacterial infections

DEFF Research Database (Denmark)

Barken, Kim Bundvig; Haagensen, Janus Anders Juul; Tolker-Nielsen, Tim

2007-01-01

Methods for rapid detection of infectious bacteria and antimicrobial-resistant pathogens have evolved significantly over the last decade. Many of the new procedures are nucleic acid-based and replace conventional diagnostic methods like culturing which is time consuming especially with fastidious...... of these pathogens is important to isolate patients and prevent further spreading of the diseases. Newly developed diagnostic procedures are superior with respect to turnaround time, sensitivity and specificity. Methods like multiplex real time PCR and different array-based technologies offer the possibility...
Nucleic acid constructs containing orthogonal site selective recombinases (OSSRs)

Energy Technology Data Exchange (ETDEWEB)

Gilmore, Joshua M.; Anderson, J. Christopher; Dueber, John E.

2017-08-29

The present invention provides for a recombinant nucleic acid comprising a nucleotide sequence comprising a plurality of constructs, wherein each construct independently comprises a nucleotide sequence of interest flanked by a pair of recombinase recognition sequences. Each pair of recombinase recognition sequences is recognized by a distinct recombinase. Optionally, each construct can, independently, further comprise one or more genes encoding a recombinase capable of recognizing the pair of recombinase recognition sequences of the construct. The recombinase can be an orthogonal (non-cross reacting), site-selective recombinase (OSSR).
Unlocked nucleic acids with a pyrene-modified uracil: Synthesis, hybridization studies, fluorescent properties and i-motif stability

DEFF Research Database (Denmark)

Perlíková, P.; Karlsen, K.K.; Pedersen, E.B.

2014-01-01

The synthesis of two new phosphoramidite building blocks for the incorporation of 5-(pyren-1-yl)uracilyl unlocked nucleic acid (UNA) monomers into oligonucleotides has been developed. Monomers containing a pyrene-modified nucleobase component were found to destabilize an i-motif structure at pH 5...... intensities upon hybridization to DNA or RNA. Efficient quenching of fluorescence of pyrene-modified UNA monomers was observed after formation of i-motif structures at pH 5.2. The stabilizing/destabilizing effect of pyrene-modified nucleic acids might be useful for designing antisense oligonucleotides...
Synthesis and configurational analysis of phosphonate cavitands

NARCIS (Netherlands)

Jacopozzi, Paola; Dalcanale, Enrico; Spera, Silvia; Chrisstoffels, L.A.J.; Reinhoudt, David; Lippmann, Tino; Mann, Gerhard

1998-01-01

Synthesis, separation and configurational analysis of phosphonated and partially phosphonated cavitands derived from resorcinarenes are described. The configuration of all diastereomers has been elucidated by their 1H, 31P NMR spectra and 13C relaxation times. In all cases the course of the bridging
Extraction of nucleic acids from yeast cells and plant tissues using ethanol as medium for sample preservation and cell disruption.

Science.gov (United States)

Linke, Bettina; Schröder, Kersten; Arter, Juliane; Gasperazzo, Tatiana; Woehlecke, Holger; Ehwald, Rudolf

2010-09-01

Here we report that dehydrated ethanol is an excellent medium for both in situ preservation of nucleic acids and cell disruption of plant and yeast cells. Cell disruption was strongly facilitated by prior dehydration of the ethanol using dehydrated zeolite. Following removal of ethanol, nucleic acids were extracted from the homogenate pellet using denaturing buffers. The method provided DNA and RNA of high yield and integrity. Whereas cell wall disruption was essential for extraction of DNA and large RNA molecules, smaller molecules such as tRNAs could be selectively extracted from undisrupted, ethanol-treated yeast cells. Our results demonstrate the utility of absolute ethanol for sample fixation, cell membrane and cell wall disruption, as well as preservation of nucleic acids during sample storage.
Nucleic Acid Aptamers Against Biotoxins: A New Paradigm Toward the Treatment and Diagnostic Approach

DEFF Research Database (Denmark)

Lauridsen, Lasse Holm; Veedu, Rakesh N.

2012-01-01

Nucleic acid aptamers are short single-stranded DNA or RNA oligonucleotides that can bind to their targets with very high affinity and specificity, and are generally selected by a process referred to as systematic evolution of ligands by exponential enrichment. Conventional antibody-based therape......Nucleic acid aptamers are short single-stranded DNA or RNA oligonucleotides that can bind to their targets with very high affinity and specificity, and are generally selected by a process referred to as systematic evolution of ligands by exponential enrichment. Conventional antibody......-based therapeutic and diagnostic approach currently employed against biotoxins pose major limitations such as the requirement of a live animal for the in vivo enrichment of the antibody species, decreased stability, high production cost, and side effects. Aptamer technology is a viable alternative that can be used...
One window-period donation in two years of individual donor-nucleic acid test screening for hepatitis B, hepatitis C and human immunodeficiency virus

Directory of Open Access Journals (Sweden)

Jose Eduardo Levi

2013-06-01

Full Text Available Objective: To describe general data on nucleic acid/serology testing and report the first hepatitis B-nucleic acid testing yield case of an immunized donor in Brazil. Methods: A total of 24,441 donations collected in 2010 and 2011 were submitted to individual nucleic acid testing for hepatitis B, hepatitis C and human immunodeficiency virus using the TaqMan® MPX kit (Roche on the Cobas s201 platform, in addition to routine screening for serological markers. Nucleic acid testing-reactive donations were further evaluated by real-time polymerase chain reaction using Cobas AmpliPrep/Cobas TaqMan hepatitis B virus, hepatitis C virus and human immunodeficiency virus tests. Results: Thirty-two donations were reactive by nucleic acid testing, 31 were also serologically reactive and one first-time donor was identified as having hepatitis B in the window period. Follow-up samples showed increasing titers of anti-HBs rising from 19 UI/mL in the index donation to 109 IU/mL seven months later attributable to his vaccination history. Curiously, this donor was never reactive for HbsAg nor for anti-HBc. In the yield donation, he was concomitantly reactive for syphilis (enzyme immunoassay and fluorescent treponemal antibody-absorption; venereal disease research laboratory non-reactive. Overall, six donors (0.02% were characterized as occult hepatitis B. A total of 35% of the confirmed (recombinant immunoblot assay positive hepatitis C donations were nucleic acid testing non-reactive and no human immunodeficiency virus "elite controller" was identified. Conclusion: The yield rate (1:24,441; 95% confidence interval: 1:9,537 - 1:89,717 contrasts to the North American rate (1:410,540 donations and strongly advocates the adoption of nucleic acid testing for hepatitis B in Brazil despite the increasing rate of anti-HBs reactive subjects due to the successful immunization program.
Hydrophobic and electrostatic interactions between cell penetrating peptides and plasmid DNA are important for stable non-covalent complexation and intracellular delivery.

Science.gov (United States)

Upadhya, Archana; Sangave, Preeti C

2016-10-01

Cell penetrating peptides are useful tools for intracellular delivery of nucleic acids. Delivery of plasmid DNA, a large nucleic acid, poses a challenge for peptide mediated transport. The paper investigates and compares efficacy of five novel peptide designs for complexation of plasmid DNA and subsequent delivery into cells. The peptides were designed to contain reported DNA condensing agents and basic cell penetrating sequences, octa-arginine (R 8 ) and CHK 6 HC coupled to cell penetration accelerating peptides such as Bax inhibitory mutant peptide (KLPVM) and a peptide derived from the Kaposi fibroblast growth factor (kFGF) membrane translocating sequence. A tryptophan rich peptide, an analogue of Pep-3, flanked with CH 3 on either ends was also a part of the study. The peptides were analysed for plasmid DNA complexation, protection of peptide-plasmid DNA complexes against DNase I, serum components and competitive ligands by simple agarose gel electrophoresis techniques. Hemolysis of rat red blood corpuscles (RBCs) in the presence of the peptides was used as a measure of peptide cytotoxicity. Plasmid DNA delivery through the designed peptides was evaluated in two cell lines, human cervical cancer cell line (HeLa) and (NIH/3 T3) mouse embryonic fibroblasts via expression of the secreted alkaline phosphatase (SEAP) reporter gene. The importance of hydrophobic sequences in addition to cationic sequences in peptides for non-covalent plasmid DNA complexation and delivery has been illustrated. An alternative to the employment of fatty acid moieties for enhanced gene transfer has been proposed. Comparison of peptides for plasmid DNA complexation and delivery of peptide-plasmid DNA complexes to cells estimated by expression of a reporter gene, SEAP. Copyright © 2016 European Peptide Society and John Wiley & Sons, Ltd. Copyright © 2016 European Peptide Society and John Wiley & Sons, Ltd.
Recent applications of the combination of mesoporous silica nanoparticles with nucleic acids: development of bioresponsive devices, carriers and sensors.

Science.gov (United States)

Castillo, Rafael R; Baeza, Alejandro; Vallet-Regí, María

2017-02-28

The discovery and control of the biological roles mediated by nucleic acids have turned them into a powerful tool for the development of advanced biotechnological materials. Such is the importance of these gene-keeping biomacromolecules that even nanomaterials have succumbed to the claimed benefits of DNA and RNA. Currently, there could be found in the literature a practically intractable number of examples reporting the use of combination of nanoparticles with nucleic acids, so boundaries are demanded. Following this premise, this review will only cover the most recent and powerful strategies developed to exploit the possibilities of nucleic acids as biotechnological materials when in combination with mesoporous silica nanoparticles. The extensive research done on nucleic acids has significantly incremented the technological possibilities for those biomacromolecules, which could be employed in many different applications, where substrate or sequence recognition or modulation of biological pathways due to its coding role in living cells are the most promising. In the present review, the chosen counterpart, mesoporous silica nanoparticles, also with unique properties, became a reference material for drug delivery and biomedical applications due to their high biocompatibility and porous structure suitable for hosting and delivering small molecules. Although most of the reviews dealt with significant advances in the use of nucleic acid and mesoporous silica nanoparticles in biotechnological applications, a rational classification of these new generation hybrid materials is still uncovered. In this review, there will be covered promising strategies for the development of living cell and biological sensors, DNA-based molecular gates with targeting, transfection or silencing properties, which could provide a significant advance in current nanomedicine.
Effects of nucleic acid local structure and magnesium ions on minus-strand transfer mediated by the nucleic acid chaperone activity of HIV-1 nucleocapsid protein

OpenAIRE

Wu, Tiyun; Heilman-Miller, Susan L.; Levin, Judith G.

2007-01-01

HIV-1 nucleocapsid protein (NC) is a nucleic acid chaperone, which is required for highly specific and efficient reverse transcription. Here, we demonstrate that local structure of acceptor RNA at a potential nucleation site, rather than overall thermodynamic stability, is a critical determinant for the minus-strand transfer step (annealing of acceptor RNA to (−) strong-stop DNA followed by reverse transcriptase (RT)-catalyzed DNA extension). In our system, destabilization of a stem-loop stru...
An integrated portable hand-held analyser for real-time isothermal nucleic acid amplification

International Nuclear Information System (INIS)

Smith, Matthew C.; Steimle, George; Ivanov, Stan; Holly, Mark; Fries, David P.

2007-01-01

A compact hand-held heated fluorometric instrument for performing real-time isothermal nucleic acid amplification and detection is described. The optoelectronic instrument combines a Printed Circuit Board/Micro Electro Mechanical Systems (PCB/MEMS) reaction detection/chamber containing an integrated resistive heater with attached miniature LED light source and photo-detector and a disposable glass waveguide capillary to enable a mini-fluorometer. The fluorometer is fabricated and assembled in planar geometry, rolled into a tubular format and packaged with custom control electronics to form the hand-held reactor. Positive or negative results for each reaction are displayed to the user using an LED interface. Reaction data is stored in FLASH memory for retrieval via an in-built USB connection. Operating on one disposable 3 V lithium battery >12, 60 min reactions can be performed. Maximum dimensions of the system are 150 mm (h) x 48 mm (d) x 40 mm (w), the total instrument weight (with battery) is 140 g. The system produces comparable results to laboratory instrumentation when performing a real-time nucleic acid sequence-based amplification (NASBA) reaction, and also displayed comparable precision, accuracy and resolution to laboratory-based real-time nucleic acid amplification instrumentation. A good linear response (R 2 = 0.948) to fluorescein gradients ranging from 0.5 to 10 μM was also obtained from the instrument indicating that it may be utilized for other fluorometric assays. This instrument enables an inexpensive, compact approach to in-field genetic screening, providing results comparable to laboratory equipment with rapid user feedback as to the status of the reaction
An integrated portable hand-held analyser for real-time isothermal nucleic acid amplification

Energy Technology Data Exchange (ETDEWEB)

Smith, Matthew C. [College of Marine Science, University of South Florida, St Petersburg, FL (United States)], E-mail: msmith@marine.usf.edu; Steimle, George; Ivanov, Stan; Holly, Mark; Fries, David P. [College of Marine Science, University of South Florida, St Petersburg, FL (United States)

2007-08-29

A compact hand-held heated fluorometric instrument for performing real-time isothermal nucleic acid amplification and detection is described. The optoelectronic instrument combines a Printed Circuit Board/Micro Electro Mechanical Systems (PCB/MEMS) reaction detection/chamber containing an integrated resistive heater with attached miniature LED light source and photo-detector and a disposable glass waveguide capillary to enable a mini-fluorometer. The fluorometer is fabricated and assembled in planar geometry, rolled into a tubular format and packaged with custom control electronics to form the hand-held reactor. Positive or negative results for each reaction are displayed to the user using an LED interface. Reaction data is stored in FLASH memory for retrieval via an in-built USB connection. Operating on one disposable 3 V lithium battery >12, 60 min reactions can be performed. Maximum dimensions of the system are 150 mm (h) x 48 mm (d) x 40 mm (w), the total instrument weight (with battery) is 140 g. The system produces comparable results to laboratory instrumentation when performing a real-time nucleic acid sequence-based amplification (NASBA) reaction, and also displayed comparable precision, accuracy and resolution to laboratory-based real-time nucleic acid amplification instrumentation. A good linear response (R{sup 2} = 0.948) to fluorescein gradients ranging from 0.5 to 10 {mu}M was also obtained from the instrument indicating that it may be utilized for other fluorometric assays. This instrument enables an inexpensive, compact approach to in-field genetic screening, providing results comparable to laboratory equipment with rapid user feedback as to the status of the reaction.
Nucleic acid nanostructures: bottom-up control of geometry on the nanoscale

International Nuclear Information System (INIS)

Seeman, Nadrian C; Lukeman, Philip S

2005-01-01

DNA may seem an unlikely molecule from which to build nanostructures, but this is not correct. The specificity of interaction that enables DNA to function so successfully as genetic material also enables its use as a smart molecule for construction on the nanoscale. The key to using DNA for this purpose is the design of stable branched molecules, which expand its ability to interact specifically with other nucleic acid molecules. The same interactions used by genetic engineers can be used to make cohesive interactions with other DNA molecules that lead to a variety of new species. Branched DNA molecules are easy to design, and they can assume a variety of structural motifs. These can be used for purposes both of specific construction, such as polyhedra, and for the assembly of topological targets. A variety of two-dimensional periodic arrays with specific patterns have been made. DNA nanomechanical devices have been built with a series of different triggers, small molecules, nucleic acid molecules and proteins. Recently, progress has been made in self-replication of DNA nanoconstructs, and in the scaffolding of other species into DNA arrangements
Localization of proteins and nucleic acids using soft x-ray microscopy

International Nuclear Information System (INIS)

Larabell, Carolyn A.; Yager, Deborah; Meyer-Ilse, Werner

2000-01-01

The high-resolution soft x-ray microscope (XM-1) at the Advanced Light Source was used to examine whole, hydrated mammalian cells, both chemically fixed and rapidly frozen and viewed in a cryostage. Using x-ray microscopy, high contrast information about the organization of the cytoplasm and nucleus of these cells was revealed at unsurpassed resolution. It is important to note that cryo-fixed cells have been examined in a state that most closely resembles their natural environment in that the cells were not exposed to chemical fixatives or chemical contrast enhancement reagents. We also used the power of soft x-ray microscopy to examine the localization of proteins and nucleic acids in whole, hydrated cells using silver-enhanced, immunogold labeling techniques. With this approach, we have obtained information about the distribution of such molecules with respect to cellular ultrastructure at five times better resolution than light microscopy. The power of soft x-ray microscopy to provide superb resolution information about the subcellular localization of proteins and nucleic acids places it in a commanding position to contribute to our understanding of the numerous molecules being identified through modern molecular biology techniques
Phosphorus sorption on marine carbonate sediment: phosphonate as model organic compounds.

Science.gov (United States)

Huang, Xiao-Lan; Zhang, Jia-Zhong

2011-11-01

Organophosphonate, characterized by the presence of a stable, covalent, carbon to phosphorus (C-P) bond, is a group of synthetic or biogenic organophosphorus compounds. The fate of these organic phosphorus compounds in the environment is not well studied. This study presents the first investigation on the sorption of phosphorus (P) in the presence of two model phosphonate compounds, 2-aminothylphosphonoic acid (2-AEP) and phosphonoformic acid (PFA), on marine carbonate sediments. In contrast to other organic P compounds, no significant inorganic phosphate exchange was observed in seawater. P was found to adsorb on the sediment only in the presence of PFA, not 2-AEP. This indicated that sorption of P from phosphonate on marine sediment was compound specific. Compared with inorganic phosphate sorption on the same sediments, P sorption from organic phosphorus is much less in the marine environment. Further study is needed to understand the potential role of the organophosphonate compounds in biogeochemical cycle of phosphorus in the environment. Copyright © 2011 Elsevier Ltd. All rights reserved.
Analysis of protein-nucleic acid interactions by photochemical cross-linking and mass spectrometry

DEFF Research Database (Denmark)

Steen, Hanno; Jensen, Ole Nørregaard

2002-01-01

. Mass spectrometry (MS) has emerged as a sensitive and efficient analytical technique for determination of such cross-linking sites in proteins. The present review of the field describes a number of MS-based approaches for the characterization of cross-linked protein-nucleic acid complexes...
Survivin mRNA antagonists using locked nucleic acid, potential for molecular cancer therapy

DEFF Research Database (Denmark)

Fisker, Niels; Westergaard, Majken; Hansen, Henrik Frydenlund

2007-01-01

We have investigated the effects of different locked nucleic acid modified antisense mRNA antagonists against Survivin in a prostate cancer model. These mRNA antagonists were found to be potent inhibitors of Survivin expression at low nanomolar concentrations. Additionally there was a pronounced ...
Synthetic Nucleic Acid Analogues in Gene Therapy: An Update for Peptide–Oligonucleotide Conjugates

DEFF Research Database (Denmark)

Taskova, Maria; Mantsiou, Anna; Astakhova, Kira

2017-01-01

The main objective of this work is to provide an update on synthetic nucleic acid analogues and nanoassemblies as tools in gene therapy. In particular, the synthesis and properties of peptide–oligonucleotide conjugates (POCs), which have high potential in research and as therapeutics, are described...
Silicon Dioxide Thin Film Mediated Single Cell Nucleic Acid Isolation

Science.gov (United States)

Bogdanov, Evgeny; Dominova, Irina; Shusharina, Natalia; Botman, Stepan; Kasymov, Vitaliy; Patrushev, Maksim

2013-01-01

A limited amount of DNA extracted from single cells, and the development of single cell diagnostics make it necessary to create a new highly effective method for the single cells nucleic acids isolation. In this paper, we propose the DNA isolation method from biomaterials with limited DNA quantity in sample, and from samples with degradable DNA based on the use of solid-phase adsorbent silicon dioxide nanofilm deposited on the inner surface of PCR tube. PMID:23874571
Switchable reconfiguration of nucleic acid nanostructures by stimuli-responsive DNA machines.

Science.gov (United States)

Liu, Xiaoqing; Lu, Chun-Hua; Willner, Itamar

2014-06-17

CONSPECTUS: The base sequence in DNA dictates structural and reactivity features of the biopolymer. These properties are implemented to use DNA as a unique material for developing the area of DNA nanotechnology. The design of DNA machines represents a rapidly developing research field in the area of DNA nanotechnology. The present Account discusses the switchable reconfiguration of nucleic acid nanostructures by stimuli-responsive DNA machines, and it highlights potential applications and future perspectives of the area. Programmed switchable DNA machines driven by various fuels and antifuels, such as pH, Hg(2+) ions/cysteine, or nucleic acid strands/antistrands, are described. These include the assembly of DNA tweezers, walkers, a rotor, a pendulum, and more. Using a pH-oscillatory system, the oscillatory mechanical operation of a DNA pendulum is presented. Specifically, the synthesis and "mechanical" properties of interlocked DNA rings are described. This is exemplified with the preparation of interlocked DNA catenanes and a DNA rotaxane. The dynamic fuel-driven reconfiguration of the catenane/rotaxane structures is followed by fluorescence spectroscopy. The use of DNA machines as functional scaffolds to reconfigurate Au nanoparticle assemblies and to switch the fluorescence features within fluorophore/Au nanoparticle conjugates between quenching and surface-enhanced fluorescence states are addressed. Specifically, the fluorescence features of the different DNA machines are characterized as a function of the spatial separation between the fluorophore and Au nanoparticles. The experimental results are supported by theoretical calculations. The future development of reconfigurable stimuli-responsive DNA machines involves fundamental challenges, such as the synthesis of molecular devices exhibiting enhanced complexities, the introduction of new fuels and antifuels, and the integration of new payloads being reconfigured by the molecular devices, such as enzymes or

Urinary markers of nucleic acid oxidation in Danish overweight/obese children and youths

DEFF Research Database (Denmark)

Kloppenborg, Julie Tonsgaard; Fonvig, Cilius Esman; Johannesen, Jesper

2016-01-01

study we investigated the relationships between urinary markers of nucleic acid oxidation concentrations and the degree of obesity and glucose metabolism in overweight compared to lean children. 42 (24 girls) and 35 lean (19 girls) children and adolescents were recruited from the Registry of the Danish...... or glucose metabolism in lean and obese children. However, sub-analyses adjusted for age, sex and the degree of obesity showed positive associations between the two hour glucose (2 h glucose) and the urinary markers 8-oxoGuo (p=0.02, r(2)= 0.63) and 8-oxodG (p=0.046, r(2)= 0.48) and between the insulinogenic...... index and 8-oxoGuo (p=0.03, r(2)=0.60) in the 12 obese children exhibiting impaired glucose tolerance. Excretion of the urinary markers of nucleic acid oxidation and the degree of obesity or the glucose metabolism were not associated in this study. Nevertheless, obese children with impaired glucose...
The Obesity-Associated FTO Gene Encodes a 2-Oxoglutarate–Dependent Nucleic Acid Demethylase

Science.gov (United States)

Gerken, Thomas; Girard, Christophe A.; Tung, Yi-Chun Loraine; Webby, Celia J.; Saudek, Vladimir; Hewitson, Kirsty S.; Yeo, Giles S. H.; McDonough, Michael A.; Cunliffe, Sharon; McNeill, Luke A.; Galvanovskis, Juris; Rorsman, Patrik; Robins, Peter; Prieur, Xavier; Coll, Anthony P.; Ma, Marcella; Jovanovic, Zorica; Farooqi, I. Sadaf; Sedgwick, Barbara; Barroso, Inês; Lindahl, Tomas; Ponting, Chris P.; Ashcroft, Frances M.; O'Rahilly, Stephen; Schofield, Christopher J.

2009-01-01

Variants in the FTO (fat mass and obesity associated) gene are associated with increased body mass index in humans. Here, we show by bioinformatics analysis that FTO shares sequence motifs with Fe(II)- and 2-oxoglutarate–dependent oxygenases. We find that recombinant murine Fto catalyzes the Fe(II)- and 2OG-dependent demethylation of 3-methylthymine in single-stranded DNA, with concomitant production of succinate, formaldehyde, and carbon dioxide. Consistent with a potential role in nucleic acid demethylation, Fto localizes to the nucleus in transfected cells. Studies of wild-type mice indicate that Fto messenger RNA (mRNA) is most abundant in the brain, particularly in hypothalamic nuclei governing energy balance, and that Fto mRNA levels in the arcuate nucleus are regulated by feeding and fasting. Studies can now be directed toward determining the physiologically relevant FTO substrate and how nucleic acid methylation status is linked to increased fat mass. PMID:17991826
Phosphonate removal from discharged circulating cooling water using iron-carbon micro-electrolysis.

Science.gov (United States)

Zhou, Zhen; Qiao, Weimin; Lin, Yangbo; Shen, Xuelian; Hu, Dalong; Zhang, Jianqiao; Jiang, Lu-Man; Wang, Luochun

2014-01-01

Phosphonate is a commonly used corrosion and scale inhibitor for a circulating cooling water (CCW) system. Its discharge could cause eutrophication of receiving waters. The iron-carbon (Fe/C) micro-electrolysis technology was used to degrade and remove phosphonate from discharged CCW. The influences of initial pH, Fe/C ratio (FCR) and temperature on phosphonate removal were investigated in a series of batch tests and optimized by response surface methodology. The quadratic model of phosphonate removal was obtained with satisfactory degrees of fitness. The optimum conditions with total phosphorus removal efficiency of 95% were obtained at pH 7.0, FCR of 1.25, and temperature of 45 °C. The phosphonate removal mechanisms were also studied. Phosphonate removal occurred predominantly via two consecutive reactive phases: the degradation of phosphonate complexes (Ca-phosphonate) and the precipitation of Fe/C micro-electrolysis products (PO₄(3-), Ca²⁺ and Fe³⁺).
Safety profile of the intravenous administration of brain-targeted stable nucleic acid lipid particles

Directory of Open Access Journals (Sweden)

Mariana Conceição

2016-03-01

Full Text Available In a clinical setting, where multiple administrations of the therapeutic agent are usually required to improve the therapeutic outcome, it is crucial to assess the immunogenicity of the administered nanoparticles. In this data work, we investigated the safety profile of the repeated intravenous administration of brain-targeted stable nucleic acid lipid particles (RVG-9r-targeted SNALPs. To evaluate local activation of the immune system, we performed analysis of mouse tissue homogenates and sections from cerebellum. To investigate peripheral activation of the immune system, we used serum of mice that were intravenously injected with RVG-9r-targeted SNALPs. These data are related and were discussed in the accompanying research article entitled “Intravenous administration of brain-targeted stable nucleic acid lipid particles alleviates Machado–Joseph disease neurological phenotype” (Conceição et al., in press [1].
Experimental approaches to the interaction of the prion protein with nucleic acids and glycosaminoglycans: Modulators of the pathogenic conversion.

Science.gov (United States)

Silva, Jerson L; Vieira, Tuane C R G; Gomes, Mariana P B; Rangel, Luciana P; Scapin, Sandra M N; Cordeiro, Yraima

2011-03-01

The concept that transmissible spongiform encephalopathies (TSEs) are caused only by proteins has changed the traditional paradigm that disease transmission is due solely to an agent that carries genetic information. The central hypothesis for prion diseases proposes that the conversion of a cellular prion protein (PrP(C)) into a misfolded, β-sheet-rich isoform (PrP(Sc)) accounts for the development of (TSE). There is substantial evidence that the infectious material consists chiefly of a protein, PrP(Sc), with no genomic coding material, unlike a virus particle, which has both. However, prions seem to have other partners that chaperone their activities in converting the PrP(C) into the disease-causing isoform. Nucleic acids (NAs) and glycosaminoglycans (GAGs) are the most probable accomplices of prion conversion. Here, we review the recent experimental approaches that have been employed to characterize the interaction of prion proteins with nucleic acids and glycosaminoglycans. A PrP recognizes many nucleic acids and GAGs with high affinities, and this seems to be related to a pathophysiological role for this interaction. A PrP binds nucleic acids and GAGs with structural selectivity, and some PrP:NA complexes can become proteinase K-resistant, undergoing amyloid oligomerization and conversion to a β-sheet-rich structure. These results are consistent with the hypothesis that endogenous polyanions (such as NAs and GAGs) may accelerate the rate of prion disease progression by acting as scaffolds or lattices that mediate the interaction between PrP(C) and PrP(Sc) molecules. In addition to a still-possible hypothesis that nucleic acids and GAGs, especially those from the host, may modulate the conversion, the recent structural characterization of the complexes has raised the possibility of developing new diagnostic and therapeutic strategies. Copyright © 2010 Elsevier Inc. All rights reserved.
A C-terminal truncated hepatitis C virus core protein variant assembles in vitro into virus-like particles in the absence of structured nucleic acids

International Nuclear Information System (INIS)

Acosta-Rivero, Nelson; Rodriguez, Armando; Mussachio, Alexis; Poutu, Johana; Falcon, Viviana; Torres, Dinorah; Aguilar, Julio C.; Linares, Marbelis; Alonso, Mabel; Perez, Angel; Menendez, Ivon; Morales-Grillo, Juan; Marquez, Gabriel; Duenas-Carrera, Santiago

2005-01-01

Little is known about the assembly pathway or structure of the hepatitis C virus (HCV). In this work a truncated HCcAg variant covering the first 120 aa (HCcAg.120) with a 32 aa N-terminal fusion peptide (6x Histag-Xpress epitope) was purified as a monomer under strong denaturing conditions. In addition, minor HCcAg.120 peaks exhibiting little different molecular mass by SDS-PAGE which possibly represents alternative forms harboring the N-termini of HCcAg.120 were detected. Analysis using gel filtration chromatography showed that HCcAg.120 assembled into high molecular weight structures in vitro in the absence of structured nucleic acids. The negative-stain electron microscopy analysis revealed that these structures correspond with spherical VLPs of uniform morphology and size distribution. The diameters of these particles ranged from 20 to 43 nm with an average diameter of approximately 30 nm and were specifically immunolabelled with a mouse monoclonal antibody against the residues 5-35 of HCcAg. Results presented in this work showed that HCcAg.120 assembled in vitro into VLPs in the absence of structured nucleic acids with similar morphology and size distribution to those found in sera and hepatocytes from HCV-infected patients. Therefore, these VLPs would be important to elucidate the mechanisms behind the ability of HCcAg to assemble into a nucleocapsid structure
Does the high nucleic acid content of individual bacterial cells allow us to discriminate between active cells and inactive cells in aquatic systems?

Science.gov (United States)

Lebaron, P; Servais, P; Agogué, H; Courties, C; Joux, F

2001-04-01

The nucleic acid contents of individual bacterial cells as determined with three different nucleic acid-specific fluorescent dyes (SYBR I, SYBR II, and SYTO 13) and flow cytometry were compared for different seawater samples. Similar fluorescence patterns were observed, and bacteria with high apparent nucleic acid contents (HNA) could be discriminated from bacteria with low nucleic acid contents (LNA). The best discrimination between HNA and LNA cells was found when cells were stained with SYBR II. Bacteria in different water samples collected from seven freshwater, brackish water, and seawater ecosystems were prelabeled with tritiated leucine and then stained with SYBR II. After labeling and staining, HNA, LNA, and total cells were sorted by flow cytometry, and the specific activity of each cellular category was determined from leucine incorporation rates. The HNA cells were responsible for most of the total bacterial production, and the specific activities of cells in the HNA population varied between samples by a factor of seven. We suggest that nucleic acid content alone can be a better indicator of the fraction of growing cells than total counts and that this approach should be combined with other fluorescent physiological probes to improve detection of the most active cells in aquatic systems.
Fanconi anemia complementation group A (FANCA) protein has intrinsic affinity for nucleic acids with preference for single-stranded forms.

Science.gov (United States)

Yuan, Fenghua; Qian, Liangyue; Zhao, Xinliang; Liu, Jesse Y; Song, Limin; D'Urso, Gennaro; Jain, Chaitanya; Zhang, Yanbin

2012-02-10

The Fanconi anemia complementation group A (FANCA) gene is one of 15 disease-causing genes and has been found to be mutated in ∼60% of Fanconi anemia patients. Using purified protein, we report that human FANCA has intrinsic affinity for nucleic acids. FANCA binds to both single-stranded (ssDNA) and double-stranded (dsDNA) DNAs; however, its affinity for ssDNA is significantly higher than for dsDNA in an electrophoretic mobility shift assay. FANCA also binds to RNA with an intriguingly higher affinity than its DNA counterpart. FANCA requires a certain length of nucleic acids for optimal binding. Using DNA and RNA ladders, we determined that the minimum number of nucleotides required for FANCA recognition is ∼30 for both DNA and RNA. By testing the affinity between FANCA and a variety of DNA structures, we found that a 5'-flap or 5'-tail on DNA facilitates its interaction with FANCA. A patient-derived FANCA truncation mutant (Q772X) has diminished affinity for both DNA and RNA. In contrast, the complementing C-terminal fragment of Q772X, C772-1455, retains the differentiated nucleic acid-binding activity (RNA > ssDNA > dsDNA), indicating that the nucleic acid-binding domain of FANCA is located primarily at its C terminus, where most disease-causing mutations are found.
Fanconi Anemia Complementation Group A (FANCA) Protein Has Intrinsic Affinity for Nucleic Acids with Preference for Single-stranded Forms*

Science.gov (United States)

Yuan, Fenghua; Qian, Liangyue; Zhao, Xinliang; Liu, Jesse Y.; Song, Limin; D'Urso, Gennaro; Jain, Chaitanya; Zhang, Yanbin

2012-01-01

The Fanconi anemia complementation group A (FANCA) gene is one of 15 disease-causing genes and has been found to be mutated in ∼60% of Fanconi anemia patients. Using purified protein, we report that human FANCA has intrinsic affinity for nucleic acids. FANCA binds to both single-stranded (ssDNA) and double-stranded (dsDNA) DNAs; however, its affinity for ssDNA is significantly higher than for dsDNA in an electrophoretic mobility shift assay. FANCA also binds to RNA with an intriguingly higher affinity than its DNA counterpart. FANCA requires a certain length of nucleic acids for optimal binding. Using DNA and RNA ladders, we determined that the minimum number of nucleotides required for FANCA recognition is ∼30 for both DNA and RNA. By testing the affinity between FANCA and a variety of DNA structures, we found that a 5′-flap or 5′-tail on DNA facilitates its interaction with FANCA. A patient-derived FANCA truncation mutant (Q772X) has diminished affinity for both DNA and RNA. In contrast, the complementing C-terminal fragment of Q772X, C772–1455, retains the differentiated nucleic acid-binding activity (RNA > ssDNA > dsDNA), indicating that the nucleic acid-binding domain of FANCA is located primarily at its C terminus, where most disease-causing mutations are found. PMID:22194614
Products obtained after in vitro reaction of 7,12-dimethylbenz[alpha]anthracene 5,6-oxide with nucleic acids.

Science.gov (United States)

Blobstein, S H; Weinstein, I B; Grunberger, D; Weisgras, J; Harvey, R G

1975-07-29

Several lines of evidence suggest that oxide derivatives of carcinogenic polycyclic hydrocarbons are the reactive intermediates for in vivo binding to cellular nucleic acids. In the present study the covalent binding of 7,12-dimethylbenz[alpha]anthracene 5,6-oxide to synthetic homopolymers and nucleic acids in aqueous-acetone solutions has been investigated. Poly(G) was found to be the most reactive nucleic acid and underwent approximately 7-10% modification. Alkaline hydrolysis of the poly(G)-dimethylbenzathracene conjugate yielded chromatographically distinct polycyclic hydrocarbon-modified nucleotides which were further characterized by spectral analyses and enzymatic and chemical degradation. When the oxide was allowed to react with GMP or dGMP, at least two products were obtained in about 1% yield. Acid hydrolysis of the dGMP-dimethylbenzanthracene conjugates liberated the corresponding guanine-dimethylbenzathracene products. Mass spectral analysis of the modified bases provided direct evidence that we had obtained covalent binding of the poly-cyclic hydrocarbon to guanine. The mass spectral cleavage pattern suggest that one of these products is a hydroxydihydro derivative of dimethylbenzanthracene bound to guanine and the other is a dimethylbenzanthracene-guanine conjugate. Additional structural aspects of these guanine derivatives are discussed.
Continuously tunable nucleic acid hybridization probes.

Science.gov (United States)

Wu, Lucia R; Wang, Juexiao Sherry; Fang, John Z; Evans, Emily R; Pinto, Alessandro; Pekker, Irena; Boykin, Richard; Ngouenet, Celine; Webster, Philippa J; Beechem, Joseph; Zhang, David Yu

2015-12-01

In silico-designed nucleic acid probes and primers often do not achieve favorable specificity and sensitivity tradeoffs on the first try, and iterative empirical sequence-based optimization is needed, particularly in multiplexed assays. We present a novel, on-the-fly method of tuning probe affinity and selectivity by adjusting the stoichiometry of auxiliary species, which allows for independent and decoupled adjustment of the hybridization yield for different probes in multiplexed assays. Using this method, we achieved near-continuous tuning of probe effective free energy. To demonstrate our approach, we enforced uniform capture efficiency of 31 DNA molecules (GC content, 0-100%), maximized the signal difference for 11 pairs of single-nucleotide variants and performed tunable hybrid capture of mRNA from total RNA. Using the Nanostring nCounter platform, we applied stoichiometric tuning to simultaneously adjust yields for a 24-plex assay, and we show multiplexed quantitation of RNA sequences and variants from formalin-fixed, paraffin-embedded samples.
Haematoxylin and eosin staining identifies medium to large bacterial aggregates with a reliable specificity: A comparative analysis of follicular bacterial aggregates in axillary biopsies using peptide nucleic acid-fluorescence in situ hybridization and haematoxylin and eosin staining.

Science.gov (United States)

Ring, Hans Christian; Theut Riis, Peter; Bay, Lene; Kallenbach, Klaus; Bjarnsholt, Thomas; Jemec, Gregor B E

2017-10-01

Although peptide nucleic acid (PNA), fluorescence in situ hybridization (FISH) and confocal laser scanning microscopy (CLSM) are the reference tools in the study of bacterial aggregates/biofilms, it may also be rather time-consuming. This study aimed to investigate the sensitivity and specificity between bacterial aggregates identified by haematoxylin and eosin (HE) staining vs bacterial aggregates in corresponding PNA-FISH samples. Axillary biopsies were obtained in 24 healthy controls. HE-stained and PNA-FISH samples were investigated using traditional light microscopy and CLSM, respectively. The data demonstrate that HE staining identifies large bacterial aggregates (>10 μm) with a sensitivity of 0.43 and specificity of 1. The methods, however, are not equivalent as demonstrated by a McNemar's test (P=.04). Where bacterial aggregates >10 μm in diameter, HE staining may offer a rapid and practical low-cost tool to evaluate bacterial aggregates. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.
Formation of taste-active amino acids, amino acid derivatives and peptides in food fermentations - A review.

Science.gov (United States)

Zhao, Cindy J; Schieber, Andreas; Gänzle, Michael G

2016-11-01

Fermented foods are valued for their rich and complex odour and taste. The metabolic activity of food-fermenting microorganisms determines food quality and generates odour and taste compounds. This communication reviews the formation of taste-active amino acids, amino acid derivatives and peptides in food fermentations. Pathways of the generation of taste compounds are presented for soy sauce, cheese, fermented meats, and bread. Proteolysis or autolysis during food fermentations generates taste-active amino acids and peptides; peptides derived from proteolysis particularly impart umami taste (e.g. α-glutamyl peptides) or bitter taste (e.g. hydrophobic peptides containing proline). Taste active peptide derivatives include pyroglutamyl peptides, γ-glutamyl peptides, and succinyl- or lactoyl amino acids. The influence of fermentation microbiota on proteolysis, and peptide hydrolysis, and the metabolism of glutamate and arginine is well understood, however, the understanding of microbial metabolic activities related to the formation of taste-active peptide derivatives is incomplete. Improved knowledge of the interactions between taste-active compounds will enable the development of novel fermentation strategies to develop tastier, less bitter, and low-salt food products, and may provide novel and "clean label" ingredients to improve the taste of other food products. Copyright © 2016 Elsevier Ltd. All rights reserved.
The nucleic acid metabolism in rat liver after single and long-term administration of tritium oxide

International Nuclear Information System (INIS)

Shorokhova, V.B.

1984-01-01

It was shown that after a single administration of tritiUm oxide in a dose of 22.2 MBq/g body mass the liver mass increased, the concentration of nucleic acids decreased and the biosynthesjs rate increased dUring a one-month observation. By the end of the observation period (the first year) the parameters under study were normalized. The long-term administration of tritium oxide in daily doses of 0.37, 0.925 and 1.85 MBq/g body mass caused changes in the nucleac acid metabolism which were less manifest (at early times), than in the case of a single injection. At the same time, the long-term administration of tritium oxide in the dose of 0.925 MBq/g caused a substantial disturbance of the nucleic acid metabolism at later times (after 2-9 months)
A Prebiotic Chemistry Experiment on the Adsorption of Nucleic Acids Bases onto a Natural Zeolite.

Science.gov (United States)

Anizelli, Pedro R; Baú, João Paulo T; Gomes, Frederico P; da Costa, Antonio Carlos S; Carneiro, Cristine E A; Zaia, Cássia Thaïs B V; Zaia, Dimas A M

2015-09-01

There are currently few mechanisms that can explain how nucleic acid bases were synthesized, concentrated from dilute solutions, and/or protected against degradation by UV radiation or hydrolysis on the prebiotic Earth. A natural zeolite exhibited the potential to adsorb adenine, cytosine, thymine, and uracil over a range of pH, with greater adsorption of adenine and cytosine at acidic pH. Adsorption of all nucleic acid bases was decreased in artificial seawater compared to water, likely due to cation complexation. Furthermore, adsorption of adenine appeared to protect natural zeolite from thermal degradation. The C=O groups from thymine, cytosine and uracil appeared to assist the dissolution of the mineral while the NH2 group from adenine had no effect. As shown by FT-IR spectroscopy, adenine interacted with a natural zeolite through the NH2 group, and cytosine through the C=O group. A pseudo-second-order model best described the kinetics of adenine adsorption, which occurred faster in artificial seawaters.
Colorimetric Nucleic Acid Detection on Paper Microchip Using Loop Mediated Isothermal Amplification and Crystal Violet Dye.

Science.gov (United States)

Roy, Sharmili; Mohd-Naim, Noor Faizah; Safavieh, Mohammadali; Ahmed, Minhaz Uddin

2017-11-22

Nucleic acid detection is of paramount importance in monitoring of microbial pathogens in food safety and infectious disease diagnostic applications. To address these challenges, a rapid, cost-effective label-free technique for nucleic acid detection with minimal instrumentations is highly desired. Here, we present paper microchip to detect and quantify nucleic acid using colorimetric sensing modality. The extracted DNA from food samples of meat as well as microbial pathogens was amplified utilizing loop-mediated isothermal amplification (LAMP). LAMP amplicon was then detected and quantified on a paper microchip fabricated in a cellulose paper and a small wax chamber utilizing crystal violet dye. The affinity of crystal violet dye toward dsDNA and positive signal were identified by changing the color from colorless to purple. Using this method, detection of Sus scrofa (porcine) and Bacillus subtilis (bacteria) DNA was possible at concentrations as low as 1 pg/μL (3.43 × 10 -1 copies/μL) and 10 pg/μL (2.2 × 10 3 copies/μL), respectively. This strategy can be adapted for detection of other DNA samples, with potential for development of a new breed of simple and inexpensive paper microchip at the point-of-need.
Locked vs. unlocked nucleic acids (LNA vs. UNA): contrasting structures work towards common therapeutic goals

DEFF Research Database (Denmark)

Campbell, Meghan A; Wengel, Jesper

2011-01-01

Oligonucleotide chemistry has been developed greatly over the past three decades, with many advances in increasing nuclease resistance, enhancing duplex stability and assisting with cellular uptake. Locked nucleic acid (LNA) is a structurally rigid modification that increases the binding affinity...... of a modified-oligonucleotide. In contrast, unlocked nucleic acid (UNA) is a highly flexible modification, which can be used to modulate duplex characteristics. In this tutorial review, we will compare the synthetic routes to both of these modifications, contrast the structural features, examine...... the hybridization properties of LNA and UNA modified duplexes, and discuss how they have been applied within biotechnology and drug research. LNA has found widespread use in antisense oligonucleotide technology, where it can stabilize interactions with target RNA and protect from cellular nucleases. The newly...
DMPD: Nucleic acid-sensing TLRs as modifiers of autoimmunity. [Dynamic Macrophage Pathway CSML Database

Lifescience Database Archive (English)

Full Text Available S. J Immunol. 2006 Nov 15;177(10):6573-8. (.png) (.svg) (.html) (.csml) Show Nucleic acid-sensing TLRs as mo...way - PNG File (.png) SVG File (.svg) HTML File (.html) CSML File (.csml) Open .csml file with CIOPlayer Ope
77 FR 16126 - Microbiology Devices; Reclassification of Nucleic Acid-Based Systems for Mycobacterium tuberculosis

Science.gov (United States)

2012-03-19

.... FDA-2012-N-0159] Microbiology Devices; Reclassification of Nucleic Acid-Based Systems for... convened a meeting of the Microbiology Devices Panel of the Medical Devices Advisory Committee (Microbiology Devices Panel) on June 29, 2011 (Ref. 2). Although not a formal reclassification meeting, panel...
Spectrofluorometric study on photochemical interaction between chlorpromazine and nucleic acids

International Nuclear Information System (INIS)

Fujita, H.; Hayashi, H.; Suzuki, K.

1981-01-01

Near-UV irradiation of a mixture of chlorpromazine and single-stranded nucleic acids produced a non-dialyzable photoproduct which emitted characteristic fluorescence at around 520 nm. The same fluorescent species was also formed by the photoreaction with purine nucleotides but not with pyrimidine nucleotide. The highest photoreactivity was observed with GMP. Smaller amounts of the species were formed in a solution with a high salt concentration than in that with a low salt concentration. A higher rate was observed under anaerobic conditions than under aerobic conditions. (author)

Photobiological behavior of bacteria and phages supplemented with aza-analogues of nucleic acid bases

Energy Technology Data Exchange (ETDEWEB)

Kittler, L; Hradecna, Z; Jacob, H E; Loeber, G

1975-01-01

The photochemical stability of anomalous nucleic acid bases of the azatype, 5-azacytosine (1), 5-azacytidine (II), 6-azacytosine (III), 6-azacytidine (IV), 6-azathymine (V), 6-azauracil (VI), and 8-aza-adenine (VII) to uv light of the wavelength 254 nm differs from the uv stability of the normal constituents. Changes of the uv inactivation of Escherichia coli K12 C600, E. coli B, Bacillus cereus, as well as E. coli phages lambda cb/sub 2/ and lambda b/sub 2/b/sub 5/ supplemented with azaderivatives prior to irradiation were investigated. It was found that I, II, III, IV, and VII are more, V and VI less sensitive to uv light compared with corresponding natural nucleic acid bases. Their changed uv sensitivities are reflected in the survival curves after uv irradiation in as far as azabases are incorporated into the nucleic acids in vivo. This explains the increase in uv sensitivity of E. coli K 12 C600, E. coli B, and B. cereus supplemented with I, II, III, IV, and VII and the decrease in uv sensitivity of Streptococcus faecalis supplemented with V (the latter information was taken from Gunther and Prusoff 1967). The lack of any significant influence of inactivation curves of E. coli K 12 C600 by V and VI, and on E. coli phages lambda cb/sub 2/ and lambda c/sub 2/b/sub 5/ by II, is discussed in terms of too small incorporation rates. No discrimination was put forward with respect to DNA and RNA incorporation.
Effect of gamma rays on nucleic acids content (RNA and DNA) of the cotton leaf worm Spodoptera Littoralis (BOISD). Vol. 4

International Nuclear Information System (INIS)

Sallam, H.A.; El-Shall, S.A.; Sobeiha, A.K.; El-Bamby, M.A.

1996-01-01

Full grown pupae of the cotton leaf worm Spodoptera Littoralis (Boisd) were exposed to exposed to sub sterilizing doses of 100, 200 and 300 Gy gamma radiation. The changes in nucleic acids content (RNA and DNA) of irradiated pupae, after 24 hours from irradiation, and also in 3 days old adults resulting from irradiated pupae were investigated. The total nucleic acids content in either pupae or adults was progressively reduced as the dose was increased. The reduction of both RNA and DNA in females was greater than in males. DNA was more radiosensitive than RNA. The destructive action of irradiation on nucleic acids was more pronounced in adult stage. Irradiation increased the RNA/DNA ratio than control at all treatments for female pupae at 200 Gy. 2 tabs
Effect of gamma rays on nucleic acids content (RNA and DNA) of the cotton leaf worm Spodoptera Littoralis (BOISD). Vol. 4.

Energy Technology Data Exchange (ETDEWEB)

Sallam, H A; El-Shall, S A [Biological Applications Department, Nuclear Research Center, Atomic Energy Authority, Cairo (Egypt); Sobeiha, A K; El-Bamby, M A [Plant Protection Department, Faculty of Agriculture, Ain Shams University, Cairo (Egypt)

1996-03-01

Full grown pupae of the cotton leaf worm Spodoptera Littoralis (Boisd) were exposed to exposed to sub sterilizing doses of 100, 200 and 300 Gy gamma radiation. The changes in nucleic acids content (RNA and DNA) of irradiated pupae, after 24 hours from irradiation, and also in 3 days old adults resulting from irradiated pupae were investigated. The total nucleic acids content in either pupae or adults was progressively reduced as the dose was increased. The reduction of both RNA and DNA in females was greater than in males. DNA was more radiosensitive than RNA. The destructive action of irradiation on nucleic acids was more pronounced in adult stage. Irradiation increased the RNA/DNA ratio than control at all treatments for female pupae at 200 Gy. 2 tabs.
Variation, differential reproduction and oscillation: the evolution of nucleic acid hybridization.

Science.gov (United States)

Suárez-Díaz, Edna

2013-01-01

This paper builds upon Hans-Jörg Rheinberger ideas on the oscillation and intercalation of epistemic things and technical objects in experimental systems, to give a fine-grained analysis of what here is called the problems of "adaptation" between our material and cognitive tools and the phenomena of the material world. To do so, it relies on the case-study of the evolution of nucleic acid hybridization and the stabilization of satellite DNA.
Analysis of Peptides and Conjugates by Amino Acid Analysis

DEFF Research Database (Denmark)

Højrup, Peter

2015-01-01

Amino acid analysis is a highly accurate method for characterization of the composition of synthetic peptides. Together with mass spectrometry, it gives a reliable control of peptide quality and quantity before conjugation and immunization.Peptides are hydrolyzed, preferably in gas phase, with 6 M...... HCl at 110 °C for 20-24 h and the resulting amino acids analyzed by ion-exchange chromatography with post-column ninhydrin derivatization. Depending on the hydrolysis conditions, tryptophan is destroyed, and cysteine also, unless derivatized, and the amides, glutamine and asparagine, are deamidated...... to glutamic acid and aspartic acid, respectively. Three different ways of calculating results are suggested, and taking the above limitations into account, a quantitation better than 5 % can usually be obtained....
External quality assurance of malaria nucleic acid testing for clinical trials and eradication surveillance

NARCIS (Netherlands)

Murphy, S.C.; Hermsen, C.C.; Douglas, A.D.; Edwards, N.J.; Petersen, I.; Fahle, G.A.; Adams, M.; Berry, A.A.; Billman, Z.P.; Gilbert, S.C.; Laurens, M.B.; Leroy, O.; Lyke, K.E.; Plowe, C.V.; Seilie, A.M.; Strauss, K.A.; Teelen, K.; Hill, A.V.; Sauerwein, R.W.

2014-01-01

Nucleic acid testing (NAT) for malaria parasites is an increasingly recommended diagnostic endpoint in clinical trials of vaccine and drug candidates and is also important in surveillance of malaria control and elimination efforts. A variety of reported NAT assays have been described, yet no formal
78 FR 36698 - Microbiology Devices; Reclassification of Nucleic Acid-Based Systems for Mycobacterium tuberculosis

Science.gov (United States)

2013-06-19

.... FDA-2013-N-0544] Microbiology Devices; Reclassification of Nucleic Acid-Based Systems for... workshop, FDA agreed to consider this issue further and subsequently convened a meeting of the Microbiology... Health After considering the information discussed by the Microbiology Devices Panel during the June 29...
Modulation of mdm2 pre-mRNA splicing by 9-aminoacridine-PNA (peptide nucleic acid conjugates targeting intron-exon junctions

Directory of Open Access Journals (Sweden)

Nielsen Peter E

2010-06-01

Full Text Available Abstract Background Modulation of pre-mRNA splicing by antisense molecules is a promising mechanism of action for gene therapeutic drugs. In this study, we have examined the potential of peptide nucleic acid (PNA 9-aminoacridine conjugates to modulate the pre-mRNA splicing of the mdm2 human cancer gene in JAR cells. Methods We screened 10 different 15 mer PNAs targeting intron2 at both the 5' - and the 3'-splice site for their effects on the splicing of mdm2 using RT-PCR analysis. We also tested a PNA (2512 targeting the 3'-splice site of intron3 with a complementarity of 4 bases to intron3 and 11 bases to exon4 for its splicing modulation effect. This PNA2512 was further tested for the effects on the mdm2 protein level as well as for inhibition of cell growth in combination with the DNA damaging agent camptothecin (CPT. Results We show that several of these PNAs effectively inhibit the splicing thereby producing a larger mRNA still containing intron2, while skipping of exon3 was not observed by any of these PNAs. The most effective PNA (PNA2406 targeting the 3'-splice site of intron2 had a complementarity of 4 bases to intron2 and 11 bases to exon3. PNA (2512 targeting the 3'-splice site of intron3 induced both splicing inhibition (intron3 skipping and skipping of exon4. Furthermore, treatment of JAR cells with this PNA resulted in a reduction in the level of MDM2 protein and a concomitant increase in the level of tumor suppressor p53. In addition, a combination of this PNA with CPT inhibited cell growth more than CPT alone. Conclusion We have identified several PNAs targeting the 5'- or 3'-splice sites in intron2 or the 3'-splice site of intron3 of mdm2 pre-mRNA which can inhibit splicing. Antisense targeting of splice junctions of mdm2 pre-mRNA may be a powerful method to evaluate the cellular function of MDM2 splice variants as well as a promising approach for discovery of mdm2 targeted anticancer drugs.
Sustained release of nucleic acids from polymeric nanoparticles using microemulsion precipitation in supercritical carbon dioxide.

Science.gov (United States)

Ge, Jun; Jacobson, Gunilla B; Lobovkina, Tatsiana; Holmberg, Krister; Zare, Richard N

2010-12-21

A general approach for producing biodegradable nanoparticles for sustained nucleic acid release is presented. The nanoparticles are produced by precipitating a water-in-oil microemulsion in supercritical CO(2). The microemulsion consists of a transfer RNA aqueous solution (water phase), dichloromethane containing poly(l-lactic acid)-poly(ethylene glycol) (oil phase), the surfactant n-octyl β-D-glucopyranoside, and the cosurfactant n-butanol.
Pyrene-Phosphonate Conjugate: Aggregation-Induced Enhanced Emission, and Selective Fe3+ Ions Sensing Properties

Directory of Open Access Journals (Sweden)

Sachin D. Padghan

2017-08-01

Full Text Available A new pyrene-phosphonate colorimetric receptor 1 has been designed and synthesized in a one-step process via amide bond formation between pyrene butyric acid chloride and phosphonate-appended aniline. The pyrene-phosphonate receptor 1 showed aggregation-induced enhanced emission (AIEE properties in water/acetonitrile (ACN solutions. Dynamic light scattering (DLS characterization revealed that the aggregates of receptor 1 at 80% water fraction have an average size of ≈142 nm. Field emission scanning electron microscopy (FE-SEM analysis confirmed the formation of spherical aggregates upon solvent evaporation. The sensing properties of receptor 1 were investigated by UV-vis, fluorescence emission spectroscopy, and other optical methods. Among the tested metal ions, receptor 1 is capable of recognizing the Fe3+ ion selectively. The changes in spectral measurements were explained on the basis of complex formation. The composition of receptor 1 and Fe3+ ions was determined by using Job’s plot and found to be 1:1. The receptor 1–Fe3+ complex showed a reversible UV-vis response in the presence of EDTA.
Nanomaterials for delivery of nucleic acid to the central nervous system (CNS)

DEFF Research Database (Denmark)

Wang, Danyang; Wu, Lin-Ping

2017-01-01

-related disease, such as neurodegeneration and disorders, suitable, safe and effective drug delivery nanocarriers have to been developed to overcome the blood brain barrier (BBB), which is the most inflexible barrier in human body. Here, we highlight the structure and function of barriers in the central nervous...... system (CNS) and summary several types of nanomaterials which can be potentially used in the brain delivery nucleic acid....
Nucleic acid and protein extraction from electropermeabilized E. coli cells on a microfluidic chip

DEFF Research Database (Denmark)

Matos, T.; Senkbeil, Silja; Mendonça, A.

2013-01-01

technique has been developed which is based on exposing E. coli cells to low voltages to allow extraction of nucleic acids and proteins. The flow-through electropermeability chip used consists of a microfluidic channel with integrated gold electrodes that promote cell envelope channel formation at low...
Hydrothermal syntheses, characterizations and crystal structures of a new lead(II) carboxylate-phosphonate with a double layer structure and a new nickel(II) carboxylate-phosphonate containing a hydrogen-bonded 2D layer with intercalation of ethylenediamines

International Nuclear Information System (INIS)

Song, J.-L.; Mao, J.-G.; Sun, Y.-Q.; Zeng, H.-Y.; Kremer, R.K.; Clearfield, Abraham

2004-01-01

Hydrothermal reactions of N,N-bis(phosphonomethyl)aminoacetic acid (HO 2 CCH 2 N(CH 2 PO 3 H 2 ) 2 ) with metal(II) salts afforded two new metal carboxylate-phosphonates, namely, Pb 2 [O 2 CCH 2 N(CH 2 PO 3 )(CH 2 PO 3 H)]·H 2 O (1) and {NH 3 CH 2 CH 2 NH 3 }{Ni[O 2 CCH 2 N(CH 2 PO 3 H) 2 ](H 2 O) 2 } 2 (2). Among two unique lead(II) ions in the asymmetric unit of complex 1, one is five coordinated by five phosphonate oxygen atoms from 5 ligands, whereas the other one is five-coordinated by a tridentate chelating ligand (1 N and 2 phosphonate O atoms) and two phosphonate oxygen atoms from two other ligands. The carboxylate group of the ligand remains non-coordinated. The bridging of above two types of lead(II) ions through phosphonate groups resulted in a double layer with the carboxylate group of the ligand as a pendant group. These double layers are further interlinked via hydrogen bonds between the carboxylate groups into a 3D network. The nickel(II) ion in complex 2 is octahedrally coordinated by a tetradentate chelating ligand (two phosphonate oxygen atoms, one nitrogen and one carboxylate oxygen atoms) and two aqua ligands. These {Ni[O 2 CCH 2 N(CH 2 PO 3 H) 2 ][H 2 O] 2 } - anions are further interlinked via hydrogen bonds between non-coordinated phosphonate oxygen atoms to form a hydrogen bonded 2D layer. The 2H-protonated ethylenediamine cations are intercalated between two layers, forming hydrogen bonds with the non-coordinated carboxylate oxygen atoms. Results of magnetic measurements for complex 2 indicate that there is weak Curie-Weiss behavior with θ=-4.4 K indicating predominant antiferromagnetic interaction between the Ni(II) ions. Indication for magnetic low-dimension magnetism could not be detected
Different ratios of docosahexaenoic and eicosapentaenoic acids do not alter growth, nucleic acid and fatty acids of juvenile cobia (Rachycentron canadum).

Science.gov (United States)

Xu, Youqing; Ding, Zhaokun; Zhang, Haizhu; Liu, Liang; Wang, Shuqi; Gorge, John

2009-12-01

An experiment was performed to study the effect of different ratios of docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA) on the growth, nucleic acid and fatty acids of cobia (Rachycentron canadum) juveniles. The juveniles were fed for 8 weeks using seven treatment diets (D-1-D-7) with the same amount of DHA and EPA (1.50 +/- 0.1% of dried diet), but varying ratios of DHA to EPA (0.90, 1.10, 1.30, 1.50, 1.70, 1.90, 2.10, respectively) and a control diet (D-0, DHA + EPA = 0.8% of dried diet, DHA/EPA = 1.30). At the end of the experiment, the mean body weight (BW) of juveniles fed D-0-D-7 increased significantly (from 6.86 +/- 1.64 in the week 0 to 58.52 +/- 16.45 g at the end of week 8, P cobia juveniles fed D-0-D-7 were significantly higher at the end of 8-week experiment than initially (P cobia juveniles increased with their growth and appeared an obvious positive relationship, especially in the muscle, based on regression analysis. The mean lipid content increased significantly in the liver (from 29.82 +/- 0.99 to 37.47 +/- 3.25% totally) and muscle (from 6.74 +/- 0.25 to 10.63 +/- 0.23% totally) of cobia juveniles (P 0.05). In the muscle and liver of juveniles, EPA decreased with its reduction in the diet; DHA, DHA/EPA ratio and poly unsaturated fatty acids (PUFAs) generally increased with their increment in the diet. The conclusion was drawn that the growth, nucleic acid and fatty acids of cobia juveniles were not significantly affected by different DHA/EPA ratios in our experiments.
Nucleic Acid-Based Therapy Approaches for Huntington's Disease

Directory of Open Access Journals (Sweden)

Tatyana Vagner

2012-01-01

Full Text Available Huntington's disease (HD is caused by a dominant mutation that results in an unstable expansion of a CAG repeat in the huntingtin gene leading to a toxic gain of function in huntingtin protein which causes massive neurodegeneration mainly in the striatum and clinical symptoms associated with the disease. Since the mutation has multiple effects in the cell and the precise mechanism of the disease remains to be elucidated, gene therapy approaches have been developed that intervene in different aspects of the condition. These approaches include increasing expression of growth factors, decreasing levels of mutant huntingtin, and restoring cell metabolism and transcriptional balance. The aim of this paper is to outline the nucleic acid-based therapeutic strategies that have been tested to date.
Rapid genotyping using pyrene-perylene locked nucleic acid complexes

DEFF Research Database (Denmark)

Kumar, Santhosh T.; Myznikova, Anna; Samokhina, Evgeniya

2013-01-01

We have developed an assay for single strand DNA and RNA detection which is based on novel pyrene-perylene FRET pairs attached to short LNA/DNA probes. The assay is based on ratiometric emission upon binding of target DNA/RNA by three combinations of fluorescent LNA/DNA reporter strands. Specific...... geometry of the pyrene fluorophore attached to the 2'-amino group of 2'-amino-LNA in position 4 allows for the first time to efficiently utilize dipole-dipole orientation parameter for sensing of single-nucleotide polymorphisms (SNPs) in nucleic acid targets by FRET. Using novel probes, SNP detection......-perylene FRET pairs, e.g., in imaging and clinical diagnostics....
On the role of mercury in the non-covalent stabilisation of consecutive U-HgII-U metal-mediated nucleic acid base pairs: metallophilic attraction enters the world of nucleic acids

Czech Academy of Sciences Publication Activity Database

Benda, Ladislav; Straka, Michal; Yoshiyuki, T.; Sychrovský, Vladimír

2011-01-01

Roč. 13, č. 1 (2011), s. 100-103 ISSN 1463-9076 R&D Projects: GA ČR GA203/09/2037; GA ČR GAP205/10/0228 Grant - others:European Reintegration Grant(XE) 230955 Institutional research plan: CEZ:AV0Z40550506 Keywords : metal lophilic attraction * nucleic acid * mercury Subject RIV: CF - Physical ; Theoretical Chemistry Impact factor: 3.573, year: 2011
Sensitive detection of nucleic acids by PNA hybridization directed co-localization of fluorescent beads

DEFF Research Database (Denmark)

Shiraishi, Takehiko; Deborggraeve, Stijn; Büscher, Philippe

2011-01-01

)avidin-coated fluorescent beads, differing in size and color [green beads (1 µm) and red beads (5.9 µm)], thereby allowing distinct detection of each PNA probe by conventional fluorescence microscopy. These two PNA beads showed easily detectable co-localization when simultaneously hybridizing to a target nucleic acid...
Catalytic phosphonation of high performance polymers and POSS. Novel components for polymer blend and nanocomposite fuel cell membranes

Energy Technology Data Exchange (ETDEWEB)

Bock, T.R.

2006-10-15

Aim of this thesis was the preparation and evaluation of phosphonated high performance (HP) polyelectrolytes and polyhedral oligomeric silsesquioxanes (POSS) for polyelectrolyte membrane fuel cell (PEMFC) application. Brominated derivatives of the commercial high performance (HP) polymers poly(ethersulfone) (PES), poly(etheretherketone) (PEEK), poly(phenylsulfone) (PPSu), poly(sulfone) (PSU) and of octaphenyl-POSS of own production were phosphonated by Ni-catalysed Arbuzov reaction. Phosphonated PSU was cast into pure and blend films with sulfonated PEEK (s-PEEK) to investigate H+-conductivity, water uptake and film morphology. Blend films' properties were referenced to films containing unmodified blend partners. Solution-compounding of phosphonated octaphenyl-POSS and s-PEEK was used to produce novel nanocomposite films. An in-situ zirconisation method was assessed as convenient strategy for novel ionically crosslinked membranes of enhanced swelling resistance. Dibromo isocyanuric acid (DBI) and N-bromo succinimide (NBS) as brominating agents allowed polymer analogous preparation of the novel brominated PES and PEEK with precise reaction control. A random distribution of functional groups, i.e. polyelectrolytes' microstructural homogeneity was revealed as decisive factor concerning solubility of phosphonated PSU. Brominated phT8 was prepared with Br2 by a high temperature approach in tetrachloroethane (TCE). Brominated polymers were phosphonated by Ni-catalysis in non-coordinating high temperature solvents, such as diphenylether, benzophenone and diphenylsulfone without notable solvent influence. The lack of solvent - catalyst complexes and high reaction temperatures of 180-200 C led to halogen-free phosphonates with unprecedented high functionalities. Polymer analogous application of P(OSiMe3)3 offered a novel direct access to easily cleavable disilyl ester derivatives. These were obtained from PEEK and PSU in near quantitative yields at NiCl2-loads as
Ebselen inhibits hepatitis C virus NS3 helicase binding to nucleic acid and prevents viral replication.

Science.gov (United States)

Mukherjee, Sourav; Weiner, Warren S; Schroeder, Chad E; Simpson, Denise S; Hanson, Alicia M; Sweeney, Noreena L; Marvin, Rachel K; Ndjomou, Jean; Kolli, Rajesh; Isailovic, Dragan; Schoenen, Frank J; Frick, David N

2014-10-17

The hepatitis C virus (HCV) nonstructural protein 3 (NS3) is both a protease, which cleaves viral and host proteins, and a helicase that separates nucleic acid strands, using ATP hydrolysis to fuel the reaction. Many antiviral drugs, and compounds in clinical trials, target the NS3 protease, but few helicase inhibitors that function as antivirals have been reported. This study focuses on the analysis of the mechanism by which ebselen (2-phenyl-1,2-benzisoselenazol-3-one), a compound previously shown to be a HCV antiviral agent, inhibits the NS3 helicase. Ebselen inhibited the abilities of NS3 to unwind nucleic acids, to bind nucleic acids, and to hydrolyze ATP, and about 1 μM ebselen was sufficient to inhibit each of these activities by 50%. However, ebselen had no effect on the activity of the NS3 protease, even at 100 times higher ebselen concentrations. At concentrations below 10 μM, the ability of ebselen to inhibit HCV helicase was reversible, but prolonged incubation of HCV helicase with higher ebselen concentrations led to irreversible inhibition and the formation of covalent adducts between ebselen and all 14 cysteines present in HCV helicase. Ebselen analogues with sulfur replacing the selenium were just as potent HCV helicase inhibitors as ebselen, but the length of the linker between the phenyl and benzisoselenazol rings was critical. Modifications of the phenyl ring also affected compound potency over 30-fold, and ebselen was a far more potent helicase inhibitor than other, structurally unrelated, thiol-modifying agents. Ebselen analogues were also more effective antiviral agents, and they were less toxic to hepatocytes than ebselen. Although the above structure-activity relationship studies suggest that ebselen targets a specific site on NS3, we were unable to confirm binding to either the NS3 ATP binding site or nucleic acid binding cleft by examining the effects of ebselen on NS3 proteins lacking key cysteines.

Rapid one-step selection method for generating nucleic acid aptamers: development of a DNA aptamer against α-bungarotoxin.

Directory of Open Access Journals (Sweden)

Lasse H Lauridsen

Full Text Available BACKGROUND: Nucleic acids based therapeutic approaches have gained significant interest in recent years towards the development of therapeutics against many diseases. Recently, research on aptamers led to the marketing of Macugen®, an inhibitor of vascular endothelial growth factor (VEGF for the treatment of age related macular degeneration (AMD. Aptamer technology may prove useful as a therapeutic alternative against an array of human maladies. Considering the increased interest in aptamer technology globally that rival antibody mediated therapeutic approaches, a simplified selection, possibly in one-step, technique is required for developing aptamers in limited time period. PRINCIPAL FINDINGS: Herein, we present a simple one-step selection of DNA aptamers against α-bungarotoxin. A toxin immobilized glass coverslip was subjected to nucleic acid pool binding and extensive washing followed by PCR enrichment of the selected aptamers. One round of selection successfully identified a DNA aptamer sequence with a binding affinity of 7.58 µM. CONCLUSION: We have demonstrated a one-step method for rapid production of nucleic acid aptamers. Although the reported binding affinity is in the low micromolar range, we believe that this could be further improved by using larger targets, increasing the stringency of selection and also by combining a capillary electrophoresis separation prior to the one-step selection. Furthermore, the method presented here is a user-friendly, cheap and an easy way of deriving an aptamer unlike the time consuming conventional SELEX-based approach. The most important application of this method is that chemically-modified nucleic acid libraries can also be used for aptamer selection as it requires only one enzymatic step. This method could equally be suitable for developing RNA aptamers.
Sustainable Catalytic Process for Synthesis of Triethyl Citrate Plasticizer over Phosphonated USY Zeolite

Directory of Open Access Journals (Sweden)

Kakasaheb Y. Nandiwaleand

2016-10-01

Full Text Available Fruits wastage is harmful to health and environment concerning spreading diseases and soil pollution, respectively. To avoid this issue, use of citrus fruit waste for the production of citric acid (CA is one of viable mean to obtain value added chemicals. Moreover, synthesis of triethyl citrate (TEC, a non-toxic plasticizer by esterification of CA with ethanol over heterogeneous catalyst would be renewable and sustainable catalytic process. In this context, parent Ultrastable Y (USY and different percentage phosphonated USY (P-USY zeolites were used for the synthesis of TEC in a closed batch reactor, for the first time. The synthesized catalysts were characterized by N2-adsorption desorption isotherm, powder X-ray diffraction (XRD and NH3 temperature programmed desorption (TPD. Effect of reaction conditions, such as the molar ratio of ethanol to CA (5:1 - 20:1, the catalyst to CA ratio (0.05 - 0.25 and reaction temperature (363-403 K, were studied in view to maximizing CA conversion and TEC yield. Phosphonated USY catalysts were found to be superior in activity (CA conversion and TEC yield than parent USY, which is attributed to the increased in total acidity with phosphonation. Among the studied catalysts, the P2USY (2% phosphorous loaded on USY was found to be an optimum catalyst with 99% CA conversion and 82% TEC yield, which is higher than the reported values. This study opens new avenues of research demonstrating principles of green chemistry such as easy separable and reusable catalyst, non-toxic product, bio-renewable synthetic route, milder operating parameters and waste minimization. Copyright © 2016 BCREC GROUP. All rights reserved Received: 12nd October 2015; Revised: 22nd December 2015; Accepted: 29th January 2016 How to Cite: Nandiwale, K.Y., Bokade, V.V. (2016. Sustainable Catalytic Process for Synthesis of Triethyl Citrate Plasticizer over Phosphonated USY Zeolite. Bulletin of Chemical Reaction Engineering & Catalysis, 11 (3: 292
Synthesis, hybridization characteristics, and fluorescence properties of oligonucleotides modified with nucleobase-functionalized locked nucleic acid adenosine and cytidine monomers.

Science.gov (United States)

Kaura, Mamta; Kumar, Pawan; Hrdlicka, Patrick J

2014-07-03

Conformationally restricted nucleotides such as locked nucleic acid (LNA) are very popular as affinity-, specificity-, and stability-enhancing modifications in oligonucleotide chemistry to produce probes for nucleic acid targeting applications in molecular biology, biotechnology, and medicinal chemistry. Considerable efforts have been devoted in recent years to optimize the biophysical properties of LNA through additional modification of the sugar skeleton. We recently introduced C5-functionalization of LNA uridines as an alternative and synthetically more straightforward approach to improve the biophysical properties of LNA. In the present work, we set out to test the generality of this concept by studying the characteristics of oligonucleotides modified with four different C5-functionalized LNA cytidine and C8-functionalized LNA adenosine monomers. The results strongly suggest that C5-functionalization of LNA pyrimidines is indeed a viable approach for improving the binding affinity, target specificity, and/or enzymatic stability of LNA-modified ONs, whereas C8-functionalization of LNA adenosines is detrimental to binding affinity and specificity. These insights will impact the future design of conformationally restricted nucleotides for nucleic acid targeting applications.
Autoradiographic studies on nucleic acid synthesis of human gastric cancer cells, 1. Relationship between nucleic acid synthesis of cancer cells and clinicopathological findings

Energy Technology Data Exchange (ETDEWEB)

Inoue, K [Kobe Univ. (Japan). School of Medicine

1982-03-01

The rate of nucleic acid synthesis of human gastric cancer cells was studied autoradiographically and was compared with clinicopathological findings. 1) /sup 3/H-thymidine labeling index (TLI, mean 22.4%, n = 21) ranged from 6.2% to 39.5%. Mitotic index (mean 1.96%) ranged from 1.18% to 3.48%. 2) Average TLIs in the cancerous lesions with serosal invasion, in microscopical stages III and IV, in scirrhous type and in cancer cells locating in pm- and ss-layers showed lower values compared with the counterparts. 3) /sup 3/H-uridine labeling index (mean 92.7%) ranged from 75.0% to 99.8%.
Amino Acid and Peptide Immobilization on Oxidized Nanocellulose: Spectroscopic Characterization

Science.gov (United States)

Barazzouk, Saïd; Daneault, Claude

2012-01-01

In this work, oxidized nanocellulose (ONC) was synthesized and chemically coupled with amino acids and peptides using a two step coupling method at room temperature. First, ONC was activated by N-ethyl-N’-(3-dimethylaminopropyl) carbodiimide hydrochloride, forming a stable active ester in the presence of N-hydroxysuccinimide. Second, the active ester was reacted with the amino group of the amino acid or peptide, forming an amide bond between ONC and the grafted molecule. Using this method, the intermolecular interaction of amino acids and peptides was avoided and uniform coupling of these molecules on ONC was achieved. The coupling reaction was very fast in mild conditions and without alteration of the polysaccharide. The coupling products (ONC-amino acids and ONC-peptides) were characterized by transmission electron microscopy and by the absorption, emission, Fourier transform infrared spectroscopy (FTIR) and X-ray photoelectron spectroscopy (XPS) spectroscopic techniques. PMID:28348303
Amino Acid and Peptide Immobilization on Oxidized Nanocellulose: Spectroscopic Characterization

Directory of Open Access Journals (Sweden)

Claude Daneault

2012-06-01

Full Text Available In this work, oxidized nanocellulose (ONC was synthesized and chemically coupled with amino acids and peptides using a two step coupling method at room temperature. First, ONC was activated by N-ethyl-N’-(3-dimethylaminopropyl carbodiimide hydrochloride, forming a stable active ester in the presence of N-hydroxysuccinimide. Second, the active ester was reacted with the amino group of the amino acid or peptide, forming an amide bond between ONC and the grafted molecule. Using this method, the intermolecular interaction of amino acids and peptides was avoided and uniform coupling of these molecules on ONC was achieved. The coupling reaction was very fast in mild conditions and without alteration of the polysaccharide. The coupling products (ONC-amino acids and ONC-peptides were characterized by transmission electron microscopy and by the absorption, emission, Fourier transform infrared spectroscopy (FTIR and X-ray photoelectron spectroscopy (XPS spectroscopic techniques.
Increasing the Analytical Sensitivity by Oligonucleotides Modified with Para- and Ortho-Twisted Intercalating Nucleic Acids - TINA

DEFF Research Database (Denmark)

Schneider, Uffe V; Géci, Imrich; Jøhnk, Nina

2011-01-01

-TINA molecules increased the melting point (Tm) of Watson-Crick based antiparallel DNA duplexes. The increase in Tm was greatest when the intercalators were placed at the 5' and 3' termini (preferable) or, if placed internally, for each half or whole helix turn. Terminally positioned TINA molecules improved......The sensitivity and specificity of clinical diagnostic assays using DNA hybridization techniques are limited by the dissociation of double-stranded DNA (dsDNA) antiparallel duplex helices. This situation can be improved by addition of DNA stabilizing molecules such as nucleic acid intercalators....... Here, we report the synthesis of a novel ortho-Twisted Intercalating Nucleic Acid (TINA) amidite utilizing the phosphoramidite approach, and examine the stabilizing effect of ortho- and para-TINA molecules in antiparallel DNA duplex formation. In a thermal stability assay, ortho- and para...
Double Displacement: an Improved Bioorthogonal Reaction Strategy for Templated Nucleic Acid Detection

OpenAIRE

Kleinbaum, Daniel J.; Miller, Gregory P.; Kool, Eric T.

2010-01-01

Quenched autoligation probes have been employed previously in a target-templated nonenzymatic ligation strategy for detecting nucleic acids in cells by fluorescence. A common source of background signal in such probes is undesired reaction with water and other cellular nucleophiles. Here we describe a new class of self-ligating probes, double displacement (DD) probes, that rely on two displacement reactions to fully unquench a nearby fluorophore. Three potential double displacement architectu...
Utility of lab-on-a-chip technology for high-throughput nucleic acid and protein analysis

DEFF Research Database (Denmark)

Hawtin, Paul; Hardern, Ian; Wittig, Rainer

2005-01-01

On-chip electrophoresis can provide size separations of nucleic acids and proteins similar to more traditional slab gel electrophoresis. Lab-on-a-chip (LoaC) systems utilize on-chip electrophoresis in conjunction with sizing calibration, sensitive detection schemes, and sophisticated data analysi...
Evaluation of cost-effective total nucleic acids extraction protocols for cultured Mycobacterium tuberculosis; a comparison by PCR amplification of genes associated with drug resistance

Directory of Open Access Journals (Sweden)

Gyamfi Oti K

2010-02-01

Full Text Available Abstract Background The emergence of drug resistant strains of Mycobacterium tuberculosis complex has made the management of tuberculosis difficult. Also, Mycobacterium species has a peculiar cell wall, made of an impermeable complex structure rich in mycolate, making the lyses of its cell difficult. In order to apply a radio-labelled-probe based detection of mutations in selected genes leading to drug resistance, we concede that the evaluation and modifications of nucleic acid extraction protocols that are less sophisticated and less prone to contamination would be useful in the management of tuberculosis in a resource-constrained setting. Findings The average amount of nucleic acids was determined for different extraction treatments. High temperature treatment only, yielded the lowest amount of nucleic acids, i.e. 15.7 ± 3.2 μg. The average amount of nucleic acids obtained with the addition of TE and triton-X100, was 133.7 ± 8.9 μg, while that obtained with the addition of TE only, and TE and SDS were 68.4 ± 22.7 μg and 70.4 ± 20.3 μg respectively. Other treatments yielded 28.8 ± 6.7 μg, 32.5 ± 2.4 μg and 36.9 ± 15.5 μg. The average amount of nucleic acids obtained with high temperature treatment in TE, and that obtained by freezing prior to high temperature treatment, successfully amplified for the genes of interest (rpoB, KatG, rrs. Conclusion We strongly recommend the use of 1× TE buffer, and freezing and heating for improved lysis of cultured M. tuberculosis, and therefore, as an effective method for the preparation of M. tuberculosis nucleic acid useful for PCR.
A Single Electrochemical Probe Used for Analysis of Multiple Nucleic Acid Sequences

Science.gov (United States)

Mills, Dawn M.; Calvo-Marzal, Percy; Pinzon, Jeffer M.; Armas, Stephanie; Kolpashchikov, Dmitry M.; Chumbimuni-Torres, Karin Y.

2017-01-01

Electrochemical hybridization sensors have been explored extensively for analysis of specific nucleic acids. However, commercialization of the platform is hindered by the need for attachment of separate oligonucleotide probes complementary to a RNA or DNA target to an electrode’s surface. Here we demonstrate that a single probe can be used to analyze several nucleic acid targets with high selectivity and low cost. The universal electrochemical four-way junction (4J)-forming (UE4J) sensor consists of a universal DNA stem-loop (USL) probe attached to the electrode’s surface and two adaptor strands (m and f) which hybridize to the USL probe and the analyte to form a 4J associate. The m adaptor strand was conjugated with a methylene blue redox marker for signal ON sensing and monitored using square wave voltammetry. We demonstrated that a single sensor can be used for detection of several different DNA/RNA sequences and can be regenerated in 30 seconds by a simple water rinse. The UE4J sensor enables a high selectivity by recognition of a single base substitution, even at room temperature. The UE4J sensor opens a venue for a re-useable universal platform that can be adopted at low cost for the analysis of DNA or RNA targets. PMID:29371782
Reaction mechanisms in the radiolysis of peptides, polypeptides and proteins II reactions at side-chain loci in model systems

International Nuclear Information System (INIS)

Garrison, W.M.

1983-11-01

The major emphasis in radiation biology at the molecular level has been on the nucleic acid component of the nucleic acid-protein complex because of its primary genetic importance. But there is increasing evidence that radiation damage to the protein component also has important biological implications. Damage to capsid protein now appears to be a major factor in the radiation inactivation of phage and other viruses. And, there is increasing evidence that radiation-chemical change in the protein component of chromation leads to changes in the stability of the repressor-operator complexes involved in gene expression. Knowledge of the radiation chemistry of protein is also of importance in other fields such as the application of radiation sterilization to foods and drugs. Recent findings that a class of compounds, the α,α'-diaminodicarboxylic acids, not normally present in food proteins, are formed in protein radiolysis is of particular significance since certain of their peptide derivatives have been showing to exhibit immunological activity. The purpose of this review is to bring together and to correlate our present knowledge of products and mechanisms in the radiolysis of peptides, polypeptides and proteins both aqueous and solid-state. In part 1 we presented a discussion of the radiation-induced reactions of the peptide main-chain in model peptide and polypeptide systems. Here in part 2 the emphasis is on the competing radiation chemistry at side-chain loci of peptide derivatives of aliphatic, aromatic-unsaturated and sulfur-containing amino acids in similar systems. Information obtained with the various experimental techniques of product analysis, competition kinetics, spin-trapping, pulse radiolysis, and ESR spectroscopy are included
Sequence-specific label-free nucleic acid biosensor for the detection of the hepatitis C virus genotype 1a using a disposable pencil graphite electrode.

Science.gov (United States)

Donmez, Soner; Arslan, Fatma; Arslan, Halit

2016-05-01

In this paper, we demonstrate a simple, sensitive, inexpensive, disposable and label-free electrochemical nucleic acid biosensor for the detection of the hepatitis C virus genotype 1a (HCV1a). The nucleic acid biosensor was designed with the amino-linked inosine-substituted 20-mer probes, which were immobilized onto a disposable pencil graphite electrode (PGE) by covalent linking. The proposed nucleic acid biosensor was linear in the range of 0.05 and 0.75 μM, exhibiting a limit of detection of 54.9 nM. The single-stranded synthetic PCR product analogs of HCV1a were also detected with satisfactory results under optimal conditions, showing the potential application of this biosensor.
The development of Sm-153 EDTMP (ethylene diamine tetramethylene phosphonic acid), a bone seeking radiotherapeutic agent

International Nuclear Information System (INIS)

Rehir Dahalan; Wan Anuar Wan Awang; Shaaban Kassim

1997-01-01

Diagnostic radiopharmaceuticals have been the workhorse of nuclear medicine since it began in the early fifties, with the exception of radioiodine, which was also a therapeutic radiopharmaceutical. Over the years, significant advancement in the design of the diagnostic radiopharmaceuticals, have enabled optimum delivery of, the gamma emitting radionuclide to the target organs, whilst minimizing the dose to the non-target organs. This technology was utilized in this work, to deliver destructive beta emitting radionuclide to target cancerous tissues with the hope of slowing down or completely ablating its growth. This work had been in the production of Sm-153 using the MINT research reactor (MINTRR) and labeling it to the ethylene diamine tetramethylene phosphonic acid (EDTMP) ligand, a bone seeking complex. The results of this work have established the optimum labeling conditions and showed the biodistribution of the Sm-153 EDTMP complex in the rat
Functionalization with C-terminal cysteine enhances transfection efficiency of cell-penetrating peptides through dimer formation

Energy Technology Data Exchange (ETDEWEB)

Amand, Helene L., E-mail: helene.amand@chalmers.se [Chalmers University of Technology, Department of Chemical and Biological Engineering/Physical Chemistry, SE-412 96 Gothenburg (Sweden); Norden, Bengt, E-mail: norden@chalmers.se [Chalmers University of Technology, Department of Chemical and Biological Engineering/Physical Chemistry, SE-412 96 Gothenburg (Sweden); Fant, Kristina, E-mail: kristina.fant@sp.se [Chalmers University of Technology, Department of Chemical and Biological Engineering/Physical Chemistry, SE-412 96 Gothenburg (Sweden)

2012-02-17

Highlights: Black-Right-Pointing-Pointer Reversible CPP dimerisation is a simple yet efficient strategy to improve delivery. Black-Right-Pointing-Pointer Dimer formation enhances peptiplex stability, resulting in increased transfection. Black-Right-Pointing-Pointer By dimerisation, the CPP EB1 even gain endosomal escape properties while lowering cytotoxicity. -- Abstract: Cell-penetrating peptides have the ability to stimulate uptake of macromolecular cargo in mammalian cells in a non-toxic manner and therefore hold promise as efficient and well tolerated gene delivery vectors. Non-covalent peptide-DNA complexes ('peptiplexes') enter cells via endocytosis, but poor peptiplex stability and endosomal entrapment are considered as main barriers to peptide-mediated delivery. We explore a simple, yet highly efficient, strategy to improve the function of peptide-based vectors, by adding one terminal cysteine residue. This allows the peptide to dimerize by disulfide bond formation, increasing its affinity for nucleic acids by the 'chelate effect' and, when the bond is reduced intracellularly, letting the complex dissociate to deliver the nucleic acid. By introducing a single C-terminal cysteine in the classical CPP penetratin and the penetratin analogs PenArg and EB1, we show that this minor modification greatly enhances the transfection capacity for plasmid DNA in HEK293T cells. We conclude that this effect is mainly due to enhanced thermodynamic stability of the peptiplexes as endosome-disruptive chloroquine is still required for transfection and the effect is more pronounced for peptides with lower inherent DNA condensation capacity. Interestingly, for EB1, addition of one cysteine makes the peptide able to mediate transfection in absence of chloroquine, indicating that dimerisation can also improve endosomal escape properties. Further, the cytotoxicity of EB1 peptiplexes is considerably reduced, possibly due to lower concentration of free peptide
Functionalization with C-terminal cysteine enhances transfection efficiency of cell-penetrating peptides through dimer formation

International Nuclear Information System (INIS)

Åmand, Helene L.; Nordén, Bengt; Fant, Kristina

2012-01-01

Highlights: ► Reversible CPP dimerisation is a simple yet efficient strategy to improve delivery. ► Dimer formation enhances peptiplex stability, resulting in increased transfection. ► By dimerisation, the CPP EB1 even gain endosomal escape properties while lowering cytotoxicity. -- Abstract: Cell-penetrating peptides have the ability to stimulate uptake of macromolecular cargo in mammalian cells in a non-toxic manner and therefore hold promise as efficient and well tolerated gene delivery vectors. Non-covalent peptide-DNA complexes (“peptiplexes”) enter cells via endocytosis, but poor peptiplex stability and endosomal entrapment are considered as main barriers to peptide-mediated delivery. We explore a simple, yet highly efficient, strategy to improve the function of peptide-based vectors, by adding one terminal cysteine residue. This allows the peptide to dimerize by disulfide bond formation, increasing its affinity for nucleic acids by the “chelate effect” and, when the bond is reduced intracellularly, letting the complex dissociate to deliver the nucleic acid. By introducing a single C-terminal cysteine in the classical CPP penetratin and the penetratin analogs PenArg and EB1, we show that this minor modification greatly enhances the transfection capacity for plasmid DNA in HEK293T cells. We conclude that this effect is mainly due to enhanced thermodynamic stability of the peptiplexes as endosome-disruptive chloroquine is still required for transfection and the effect is more pronounced for peptides with lower inherent DNA condensation capacity. Interestingly, for EB1, addition of one cysteine makes the peptide able to mediate transfection in absence of chloroquine, indicating that dimerisation can also improve endosomal escape properties. Further, the cytotoxicity of EB1 peptiplexes is considerably reduced, possibly due to lower concentration of free peptide dimer resulting from its stronger binding to DNA.
Determination of the Nucleic Acid Adducts Structure at the Nucleoside/Nucleotide Level by NMR Spectroscopy

Czech Academy of Sciences Publication Activity Database

Dračínský, Martin; Pohl, Radek

2015-01-01

Roč. 28, č. 2 (2015), s. 155-165 ISSN 0893-228X R&D Projects: GA ČR GA13-24880S Institutional support: RVO:61388963 Keywords : NMR spectroscopy * nucleic acids * nucleotides Subject RIV: CC - Organic Chemistry Impact factor: 3.025, year: 2015
Modulation of i-motif thermodynamic stability by the introduction of UNA (unlocked nucleic acid) monomers

DEFF Research Database (Denmark)

Pasternak, Anna; Wengel, Jesper

2011-01-01

The influence of acyclic RNA derivatives, UNA (unlocked nucleic acid) monomers, on i-DNA thermodynamic stability has been investigated. The 22 nt human telomeric fragment was chosen as the model sequence for stability studies. UNA monomers modulate i-motif stability in a position-depending manner...
The Influence of Lactic Acid Concentration on the Separation of Light Rare Earth Elements by Continuous Liquid-Liquid Extraction with 2-Ethylhexyl Phosphonic Acid Mono-2-ethylhexyl Ester

Science.gov (United States)

de Carvalho Gomes, Rafael; Seruff, Luciana Amaral; Scal, Maira Labanca Waineraich; Vera, Ysrael Marrero

2018-02-01

The separation of rare earth elements (REEs) using solvent extraction adding complexing agents appears to be an alternative to saponification of the extractant. We evaluated the effect of lactic acid concentration on didymium (praseodymium and neodymium) and lanthanum extraction with 2-ethylhexyl phosphonic acid mono-2-ethyl hexyl ester [HEH(EHP)] as extractant. First, we investigated in batch experiments the separation of lanthanum (La) and didymium (Pr and Nd) using McCabe-Thiele diagrams to estimate the number of extraction stages when the feed solution was or was not conditioned with lactic acid. Additionally, we conducted continuous liquid-liquid extraction experiments and evaluated the influence of lactic acid concentration on the REE extraction and separation. The tests showed that the extraction percentage of REEs and the separation factor Pr/La increased when the lactic acid concentration increased, but the didymium purity decreased. Lanthanum, praseodymium, and neodymium extraction rate were 23.0, 89.7, and 99.2 pct, respectively, with 1:1 aqueous/organic volume flow rate and feed solution doped with 0.52 mol L-1 lactic acid. The highest didymium purity reached was 92.0 pct with 0.26 mol L-1 lactic acid concentration.
Incorporation of radio-labelled nucleic acid precursors by Theileria parva in bovine blood and salivary glands of Rhipicephalus appendiculatus ticks

Energy Technology Data Exchange (ETDEWEB)

Irvin, A.D.; Boarer, C.D.H.; Kurtti, T.J.; Ocama, J.G.R. (International Lab. for Research on Animal Diseases, Nairobi (Kenya))

1981-12-01

The uptake of radio-labelled nucleic acid precursors by blood and tick salivary gland forms of Theileria parva was studied. Piroplasms took up tritiated purines, particularly hypoxanthine, but not pyrimidines. Similar uptake was recorded by T. parva, both in tick saliva and in salivary glands maintained in vitro. Intermediate parasite stages were those most readily labelled in glands; this reflected active nucleic acid synthesis associated with rapid parasite division. Radio-labelling of T. parva in tick salivary glands could be of value in procedures used for concentrating and purifying theilerial sporozoites.

Some links on this page may take you to non-federal websites. Their policies may differ from this site.