ALP gene expression in cDNA samples from bone tissue engineering using a HA/TCP/Chitosan scaffold

Science.gov (United States)

Stephanie, N.; Katarina, H.; Amir, L. R.; Gunawan, H. A.

2017-08-01

This study examined the potential use of hydroxyapatite (HA)/tricalcium phosphate (TCP)/Chitosan as a bone tissue engineering scaffold. The potential for using HA/TCP/chitosan as a scaffold was analyzed by measuring expression of the ALP osteogenic gene in cDNA from bone biopsies from four Macaque nemestrina. Experimental conditions included control (untreated), treatment with HA/TCP 70:30, HA/TCP 50:50, and HA/TCP/chitosan. cDNA samples were measured quantitively with Real-Time PCR (qPCR) and semi-quantitively by gel electrophoresis. There were no significant differences in ALP gene expression between treatment subjects after two weeks, but the HA/TCP/chitosan treatment gave the highest level of expression after four weeks. The scaffold using the HA/TCP/chitosan combination induced a higher level of expression of the osteogenic gene ALP than did scaffold without chitosan.

Horse cDNA clones encoding two MHC class I genes

Energy Technology Data Exchange (ETDEWEB)

Barbis, D.P.; Maher, J.K.; Stanek, J.; Klaunberg, B.A.; Antczak, D.F.

1994-12-31

Two full-length clones encoding MHC class I genes were isolated by screening a horse cDNA library, using a probe encoding in human HLA-A2.2Y allele. The library was made in the pcDNA1 vector (Invitrogen, San Diego, CA), using mRNA from peripheral blood lymphocytes obtained from a Thoroughbred stallion (No. 0834) homozygous for a common horse MHC haplotype (ELA-A2, -B2, -D2; Antczak et al. 1984; Donaldson et al. 1988). The clones were sequenced, using SP6 and T7 universal primers and horse-specific oligonucleotides designed to extend previously determined sequences.

Isolation and characterization of two cDNA clones encoding for glutamate dehydrogenase in Nicotiana plumbaginifolia.

Science.gov (United States)

Ficarelli, A; Tassi, F; Restivo, F M

1999-03-01

We have isolated two full length cDNA clones encoding Nicotiana plumbaginifolia NADH-glutamate dehydrogenase. Both clones share amino acid boxes of homology corresponding to conserved GDH catalytic domains and putative mitochondrial targeting sequence. One clone shows a putative EF-hand loop. The level of the two transcripts is affected differently by carbon source.

Isolation and structure of a cDNA encoding the B1 (CD20) cell-surface antigen of human B lymphocytes

International Nuclear Information System (INIS)

Tender, T.F.; Streuli, M.; Schlossman, S.F.; Saito, H.

1988-01-01

The B1 (CD20) molecule is a M/sub r/ 33,000 phosphoprotein on the surface of human B lymphocytes that may serve a central role in the homoral immune response by regulating B-cell proliferation and differentiation. In this report, a cDNA clone that encodes the B1 molecule was isolated and the amino acid sequence of B1 was determined. B-cell-specific cDNA clones were selected from a human tonsillar cDNA library by differential hybridization with labeled cDNA derived from either size-fractionated B-cell mRNA or size-fractionated T-cell mRNA. Of the 261 cDNA clones isolated, 3 cross-hybridizing cDNA clones were chosen as potential candidates for encoding B1 based on their selective hybridization to RNA from B1-positive cell lines. The longest clone, pB1-21, contained a 2.8-kilobase insert with an 891-base-pair open reading frame that encodes a protein of 33 kDa. mRNA synthesized from the pB1-21 cDNA clone in vitro was translated into a protein of the same apparent molecular weight as B1. Limited proteinase digestion of the pB1-21 translation product and B1 generated peptides of the same sizes, indicating that the pB1-21 cDNA encodes the B1 molecule. Gel blot analysis indicated that pB1-21 hybridized with two mRNA species of 2.8 and 3.4 kilobases only in B1-positive cell lines. The amino acid sequence deduced from the pB1-21 nucleotide sequence apparently lacks a signal sequence and contains three extensive hydrophobic regions. The deduced B1 amino acid sequence shows no significant homology with other known patients

Two-dimensional polyacrylamide gel analysis of Plodia interpunctella granulosis virus

International Nuclear Information System (INIS)

Russell, D.L.; Consigli, R.A.

1986-01-01

The structural polypeptides of purified Plodia interpunctella granulosis virus were analyzed by three different two-dimensional gel systems. Isoelectric focusing followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis allowed resolution of 53 acidic polypeptides in the enveloped nucleocapsid of the virus ranging in molecular weight from 97,300 to 8000. Nine of these polypeptides were shown to be glycoproteins by the technique of radiolabeled lectin blotting. Separation of the granulin in this system allowed resolution of five species, all of which have identical tryptic peptide maps. This matrix protein was demonstrated to be a phosphoglycoprotein by radiolabeled lectin blotting and acid phosphatase dephosphorylation. Nonequilibrium pH gel electrophoresis followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis allowed resolution of the major basic protein of the virus, VP12, from a more acidic protein of the same molecular weight. Tryptic peptide analysis demonstrated that these two proteins were indeed different and acid urea gels followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis allowed localization of the acidic protein to the envelope and the basic protein to the nucleocapsid of the virus. Finally, probing of the separated envelope nucleocapsid proteins in both the isoelectric focusing and nonequilibrium pH gel electrophoresis two-dimensional systems after transfer to nitrocellulose with iodinated, purified viral proteins allowed further insight into reactions which may be important in the maintenance of the virion structure

A tobacco cDNA reveals two different transcription patterns in vegetative and reproductive organs

Directory of Open Access Journals (Sweden)

I. da Silva

2002-08-01

Full Text Available In order to identify genes expressed in the pistil that may have a role in the reproduction process, we have established an expressed sequence tags project to randomly sequence clones from a Nicotiana tabacum stigma/style cDNA library. A cDNA clone (MTL-8 showing high sequence similarity to genes encoding glycine-rich RNA-binding proteins was chosen for further characterization. Based on the extensive identity of MTL-8 to the RGP-1a sequence of N. sylvestris, a primer was defined to extend the 5' sequence of MTL-8 by RT-PCR from stigma/style RNAs. The amplification product was sequenced and it was confirmed that MTL-8 corresponds to an mRNA encoding a glycine-rich RNA-binding protein. Two transcripts of different sizes and expression patterns were identified when the MTL-8 cDNA insert was used as a probe in RNA blots. The largest is 1,100 nucleotides (nt long and markedly predominant in ovaries. The smaller transcript, with 600 nt, is ubiquitous to the vegetative and reproductive organs analyzed (roots, stems, leaves, sepals, petals, stamens, stigmas/styles and ovaries. Plants submitted to stress (wounding, virus infection and ethylene treatment presented an increased level of the 600-nt transcript in leaves, especially after tobacco necrosis virus infection. In contrast, the level of the 1,100-nt transcript seems to be unaffected by the stress conditions tested. Results of Southern blot experiments have suggested that MTL-8 is present in one or two copies in the tobacco genome. Our results suggest that the shorter transcript is related to stress while the larger one is a flower predominant and nonstress-inducible messenger.

Unbiased diffusion of Brownian particles on disordered correlated potentials

International Nuclear Information System (INIS)

Salgado-Garcia, Raúl; Maldonado, Cesar

2015-01-01

In this work we study the diffusion of non-interacting overdamped particles, moving on unbiased disordered correlated potentials, subjected to Gaussian white noise. We obtain an exact expression for the diffusion coefficient which allows us to prove that the unbiased diffusion of overdamped particles on a random polymer does not depend on the correlations of the disordered potentials. This universal behavior of the unbiased diffusivity is a direct consequence of the validity of the Einstein relation and the decay of correlations of the random polymer. We test the independence on correlations of the diffusion coefficient for correlated polymers produced by two different stochastic processes, a one-step Markov chain and the expansion-modification system. Within the accuracy of our simulations, we found that the numerically obtained diffusion coefficient for these systems agree with the analytically calculated ones, confirming our predictions. (paper)

Human thyroid peroxidase: complete cDNA and protein sequence, chromosome mapping, and identification of two alternately spliced mRNAs

International Nuclear Information System (INIS)

Kimura, S.; Kotani, T.; McBride, O.W.; Umeki, K.; Hirai, K.; Nakayama, T.; Ohtaki, S.

1987-01-01

Two forms of human thyroid peroxidase cDNAs were isolated from a λgt11 cDNA library, prepared from Graves disease thyroid tissue mRNA, by use of oligonucleotides. The longest complete cDNA, designated phTPO-1, has 3048 nucleotides and an open reading frame consisting of 933 amino acids, which would encode a protein with a molecular weight of 103,026. Five potential asparagine-linked glycosylation sites are found in the deduced amino acid sequence. The second peroxidase cDNA, designated phTPO-2, is almost identical to phTPO-1 beginning 605 base pairs downstream except that it contains 1-base-pair difference and lacks 171 base pairs in the middle of the sequence. This results in a loss of 57 amino acids corresponding to a molecular weight of 6282. Interestingly, this 171-nucleotide sequence has GT and AG at its 5' and 3' boundaries, respectively, that are in good agreement with donor and acceptor splice site consensus sequences. Using specific oligonucleotide probes for the mRNAs derived from the cDNA sequences hTOP-1 and hTOP-2, the authors show that both are expressed in all thyroid tissues examined and the relative level of two mRNAs is different in each sample. The results suggest that two thyroid peroxidase proteins might be generated through alternate splicing of the same gene. By using somatic cell hybrid lines, the thyroid peroxidase gene was mapped to the short arm of human chromosome 2

One plunge or two?--hand disinfection with alcohol gel.

Science.gov (United States)

Macdonald, Duncan J M; Mckillop, Elisabeth C A; Trotter, Sylvia; Gray, Alastair J R

2006-04-01

To compare health care workers' hand surface coverage using two different volumes of alcohol gel for hand disinfection. and methods. A total of 84 members of staff in our hospital were studied. Subjects were asked to disinfect their hands with alcohol gel containing a clear fluorescent substance. Performance was assessed by using UV light to identify areas which had been missed, and the total surface area missed was calculated. A total of 42 subjects received 3.5 ml of alcohol gel, and 42 age-, sex-, and job-matched subjects received 1.75 ml of alcohol gel. Significantly less area was missed when hand disinfecting with double the volume of alcohol gel; 1.23 versus 6.35% surface area was missed (P disinfection significantly improves the efficiency of coverage of the hands with alcohol gel. This may result in lower bacterial count on the hands and may reduce the spread of nosocomial infections including that of methicillin-resistant Staphylococcus aureus.

Matching Two-dimensional Gel Electrophoresis' Spots

DEFF Research Database (Denmark)

Dos Anjos, António; AL-Tam, Faroq; Shahbazkia, Hamid Reza

2012-01-01

This paper describes an approach for matching Two-Dimensional Electrophoresis (2-DE) gels' spots, involving the use of image registration. The number of false positive matches produced by the proposed approach is small, when compared to academic and commercial state-of-the-art approaches. This ar...

Human pro. cap alpha. 1)(I) collagen: cDNA sequence for the C-propeptide domain

Energy Technology Data Exchange (ETDEWEB)

Maekelae, J K; Raassina, M; Virta, A; Vuorio, E

1988-01-11

The authors have previously constructed a cDNA clone pHCAL1, covering most of the C-terminal propeptide domain of human pro..cap alpha..1(I) collagen mRNA,by inserting a 678 bp EcoRI-XhoI fragment of cDNA into pBR322. Since the XhoI/SalI ligation prevented removal of the insert, they used the same strategy to obtain a similar clone in pUC8. RNA was isolated from fetal calvarial bones. The cDNA was digested with EcoRI and XhoI and fractionated on a 1 % agarose gel. Fragments of 650-700 bp were cloned in pUC8 at the polylinker site, which now permits easy removal of the insert. The new clone was named pHCAL1U since the RNA was isolated from another individual. The approach outlined is useful for studies on individual variation which is important to recognize when searching for disease-related mutations in type I collagen.

Cloning of a cDNA encoding a novel human nuclear phosphoprotein belonging to the WD-40 family

DEFF Research Database (Denmark)

Honoré, B; Leffers, H; Madsen, Peder

1994-01-01

We have cloned and expressed in vaccinia virus a cDNA encoding an ubiquitous 501-amino-acid (aa) phosphoprotein that corresponds to protein IEF SSP 9502 (79,400 Da, pI 4.5) in the master 2-D-gel keratinocyte protein database [Celis et al., Electrophoresis 14 (1993) 1091-1198]. The deduced aa...

Molecular cloning and expression of cDNA encoding a lumenal calcium binding glycoprotein from sarcoplasmic reticulum

International Nuclear Information System (INIS)

Leberer, E.; Charuk, J.H.M.; MacLennan, D.H.; Green, N.M.

1989-01-01

Antibody screening was used to isolate a cDNA encoding the 160-kDa glycoprotein of rabbit skeletal muscle sarcoplasmic reticulum. The cDNA is identical to that encoding the 53-kDa glycoprotein except that it contains an in-frame insertion of 1,308 nucleotides near its 5' end, apparently resulting from alternative splicing. The protein encoded by the cDNA would contain a 19-residue NH 2 -terminal signal sequence and a 453-residue COOH-terminal sequence identical to the 53-kDa glycoprotein. It would also contain a 436-amino acid insert between these sequences. This insert would be highly acidic, suggesting that it might bind Ca 2+ . The purified 160-kDa glycoprotein and the glycoprotein expressed in COS-1 cells transfected with cDNA encoding the 160-kDa glycoprotein were shown to bind 45 C 2+ in a gel overlay assay. The protein was shown to be located in the lumen of the sarcoplasmic reticulum and to be associated through Ca 2+ with the membrane. The authors propose that this lumenal Ca 2+ binding glycoprotein of the sarcoplasmic reticulum be designated sarcalumenin

Functional cloning using pFB retroviral cDNA expression libraries.

Science.gov (United States)

Felts, Katherine A; Chen, Keith; Zaharee, Kim; Sundar, Latha; Limjoco, Jamie; Miller, Anna; Vaillancourt, Peter

2002-09-01

Retroviral cDNA expression libraries allow the efficient introduction of complex cDNA libraries into virtually any mitotic cell type for screening based on gene function. The cDNA copy number per cell can be easily controlled by adjusting the multiplicity of infection, thus cell populations may be generated in which >90% of infected cells contain one to three cDNAs. We describe the isolation of two known oncogenes and one cell-surface receptor from a human Burkitt's lymphoma (Daudi) cDNA library inserted into the high-titer retroviral vector pFB.

Preparation of fluorescent-dye-labeled cDNA from RNA for microarray hybridization.

Science.gov (United States)

Ares, Manuel

2014-01-01

This protocol describes how to prepare fluorescently labeled cDNA for hybridization to microarrays. It consists of two steps: first, a mixture of anchored oligo(dT) and random hexamers is used to prime amine-modified cDNA synthesis by reverse transcriptase using a modified deoxynucleotide with a reactive amine group (aminoallyl-dUTP) and an RNA sample as a template. Second, the cDNA is purified and exchanged into bicarbonate buffer so that the amine groups in the cDNA react with the dye N-hydroxysuccinimide (NHS) esters, covalently joining the dye to the cDNA. The dye-coupled cDNA is purified again, and the amount of dye incorporated per microgram of cDNA is determined.

Two-dimensional polyacrylamide gel electrophoresis of intracellular proteins

International Nuclear Information System (INIS)

Ojima, N.; Sakamoto, T.; Yamashita, M.

1996-01-01

Since two-dimensional electrophoresis was established by O'Farrell for analysis of intracellular proteins of Escherichia coli, it has been applied to separation of proteins of animal cells and tissues, and especially to identification of stress proteins. Using this technique, proteins are separated by isoelectric focusing containing 8 m urea in the first dimension and by SDS-PAGE in the second dimension. The gels are stained with Coomassie Blue R-250 dye, followed by silver staining. In the case of radio-labeled proteins, the gels are dried and then autoradiographed. In order to identify a specific protein separated by two-dimensional electrophoresis, a technique determining the N-terminal amino acid sequence of the protein has been developed recently. After the proteins in the gel were electrotransferred to a polyvinylidene difluoride membrane, the membrane was stained for protein with Commassie Blue and a stained membrane fragment was applied to a protein sequencer. Our recent studies demonstrated that fish cells newly synthesized various proteins in response to heat shock, cold nd osmotic stresses. For example, when cellular proteins extracted from cold-treated rainbow trout cells were subjected to two-dimensional gel electrophoresis, the 70 kDa protein was found to be synthesized during the cold-treatment. N-Terminal sequence analysis showed that the cold-inducible protein was a homolog of mammalian valosin-containing protein and yeast cell division cycle gene product CDC48p. Furthermore, the sequence data were useful for preparing PCR primers and a rabbit antibody against a synthetic peptide to analyze a role for the protein in the function of trout cells and mechanisms for regulation

Two human cDNA molecules coding for the Duchenne muscular dystrophy (DMD) locus are highly homologous

Energy Technology Data Exchange (ETDEWEB)

Rosenthal, A.; Speer, A.; Billwitz, H. (Zentralinstitut fuer Molekularbiologie, Berlin-Buch (Germany Democratic Republic)); Cross, G.S.; Forrest, S.M.; Davies, K.E. (Univ. of Oxford (England))

1989-07-11

Recently the complete sequence of the human fetal cDNA coding for the Duchenne muscular dystrophy (DMD) locus was reported and a 3,685 amino acid long, rod-shaped cytoskeletal protein (dystrophin) was predicted as the protein product. Independently, the authors have isolated and sequenced different DMD cDNA molecules from human adult and fetal muscle. The complete 12.5 kb long sequence of all their cDNA clones has now been determined and they report here the nucleotide (nt) and amino acid (aa) differences between the sequences of both groups. The cDNA sequence comprises the whole coding region but lacks the first 110 nt from the 5{prime}-untranslated region and the last 1,417 nt of the 3{prime}-untranslated region. They have found 11 nt differences (approximately 99.9% homology) from which 7 occurred at the aa level.

Analysis of Two-Dimensional Electrophoresis Gel Images

DEFF Research Database (Denmark)

Pedersen, Lars

2002-01-01

This thesis describes and proposes solutions to some of the currently most important problems in pattern recognition and image analysis of two-dimensional gel electrophoresis (2DGE) images. 2DGE is the leading technique to separate individual proteins in biological samples with many biological...

Cloning, sequencing, and expression of cDNA for human β-glucuronidase

International Nuclear Information System (INIS)

Oshima, A.; Kyle, J.W.; Miller, R.D.

1987-01-01

The authors report here the cDNA sequence for human placental β-glucuronidase (β-D-glucuronoside glucuronosohydrolase, EC 3.2.1.31) and demonstrate expression of the human enzyme in transfected COS cells. They also sequenced a partial cDNA clone from human fibroblasts that contained a 153-base-pair deletion within the coding sequence and found a second type of cDNA clone from placenta that contained the same deletion. Nuclease S1 mapping studies demonstrated two types of mRNAs in human placenta that corresponded to the two types of cDNA clones isolated. The NH 2 -terminal amino acid sequence determined for human spleen β-glucuronidase agreed with that inferred from the DNA sequence of the two placental clones, beginning at amino acid 23, suggesting a cleaved signal sequence of 22 amino acids. When transfected into COS cells, plasmids containing either placental clone expressed an immunoprecipitable protein that contained N-linked oligosaccharides as evidenced by sensitivity to endoglycosidase F. However, only transfection with the clone containing the 153-base-pair segment led to expression of human β-glucuronidase activity. These studies provide the sequence for the full-length cDNA for human β-glucuronidase, demonstrate the existence of two populations of mRNA for β-glucuronidase in human placenta, only one of which specifies a catalytically active enzyme, and illustrate the importance of expression studies in verifying that a cDNA is functionally full-length

Identification of differentially expressed genes in seeds of two - AJOL

African Journals Online (AJOL)

STORAGESEVER

2009-10-19

Oct 19, 2009 ... cDNA subtraction Kit (Clontech) according to the manufacturer's protocols. To detect .... removal of vector, poor quality and polyA sequences). Sequence analysis ... suggests different activity of S-ACP-DES in the two lines at the different ... the products were resolved on 1.5% agarose gels. These are major ...

cDNA encoding a polypeptide including a hevein sequence

Energy Technology Data Exchange (ETDEWEB)

Raikhel, N.V.; Broekaert, W.F.; Namhai Chua; Kush, A.

1993-02-16

A cDNA clone (HEV1) encoding hevein was isolated via polymerase chain reaction (PCR) using mixed oligonucleotides corresponding to two regions of hevein as primers and a Hevea brasiliensis latex cDNA library as a template. HEV1 is 1,018 nucleotides long and includes an open reading frame of 204 amino acids.

Normalized cDNA libraries

Science.gov (United States)

Soares, Marcelo B.; Efstratiadis, Argiris

1997-01-01

This invention provides a method to normalize a directional cDNA library constructed in a vector that allows propagation in single-stranded circle form comprising: (a) propagating the directional cDNA library in single-stranded circles; (b) generating fragments complementary to the 3' noncoding sequence of the single-stranded circles in the library to produce partial duplexes; (c) purifying the partial duplexes; (d) melting and reassociating the purified partial duplexes to moderate Cot; and (e) purifying the unassociated single-stranded circles, thereby generating a normalized cDNA library.

Mutually unbiased bases

Indian Academy of Sciences (India)

Mutually unbiased bases play an important role in quantum cryptography [2] and in the optimal determination of the density operator of an ensemble [3,4]. A density operator ρ in N-dimensions depends on N2 1 real quantities. With the help of MUB's, any such density operator can be encoded, in an optimal way, in terms of ...

[Establishment of two-dimensional differential gel electrophoresis using cerebrospinal fluid from neurocysticercosis patients].

Science.gov (United States)

Li, Jing-Yi; Tian, Xiao-Jun; Huang, Yong; Yang, Yan-Jun; Ma, Qiao-Rong; Xue, Yan-Ping

2008-06-30

To establish the method of two-dimensional differential gel electrophoresis and obtain high resolution 2D images from cerebrospinal fluid (CSF) of patients with neurocysticercosis. CSF samples were collected from four patients diagnosed as neurocysticercosis clinically and by ELISA, computed tomography (CT) or magnetic resonance imaging (MRI), and from four healthy subjects without neurological disorders. The CSF samples were precipitated with cold acetone, then pooled by equal amount as patients and controls. The internal standard comprised equal amounts of proteins extracted from both groups. Internal standard, and proteins from the two groups were labeled prior to electrophoresis with spectrally resolvable fluorescent dyes, cyanein dye2 (Cy2), Cy3 and Cy5. Sodium dodecylsulfonate polyacrylamide gel chromatography (SDS-PAGE) and two-dimensional differential in-gel electrophoresis (2-D DIGE) of labeled samples were then run. The differential expressed proteins showed in the images of SDS-PAGE and 2-D DIGE gels scanned with 488 nm, 532 nm and 633 nm wavelength laser were analyzed by ImageQuant and DeCyde 5.0 respectively. Spot detection and quantification was performed for the differential in-gel analysis (DIA) module of DeCyder. Biological variation analysis (BVA) module of DeCyder was matched gel 1 and gel 2 images to provide data on differential protein expression levels between the two groups. The ImageQuant result displayed that the CSF protein was compatible with the dye, and the difference of protein amount was revealed by the difference of fluorescence intensity. DIA indicated that there were 896 and 894 protein dots on gel 1 and gel 2 respectively, and 90% of them were matched each other. BVA showed that there were 55 protein spots with different expressional level between neurocysticercosis and control groups. Protein spots with two-fold increase or decrease were 47 and 8 respectively in neurocysticercosis patients compared with healthy controls. The

Cloning of the cDNA for human 12-lipoxygenase

International Nuclear Information System (INIS)

Izumi, T.; Hoshiko, S.; Radmark, O.; Samuelsson, B.

1990-01-01

A full-length cDNA clone encoding 12-lipoxygenase was isolated from a human platelet cDNA library by using a cDNA for human reticulocyte 15-lipoxygenase as probe for the initial screening. The cDNA had an open reading frame encoding 662 amino acid residues with a calculated molecular weight of 75,590. Three independent clones revealed minor heterogeneities in their DNA sequences. Thus, in three positions of the deduced amino acid sequence, there is a choice between two different amino acids. The deduced sequence from the clone plT3 showed 65% identity with human reticulocyte 15-lipoxygenase and 42% identity with human leukocyte 5-lipoxygenase. The 12-lipoxygenase cDNA recognized a 3.0-kilobase mRNA species in platelets and human erythroleukemia cells (HEL cells). Phorbol 12-tetradecanoyl 13-acetate induced megakaryocytic differentiation of HEL cells and 12-lipoxygenase activity and increased mRNA for 12-lipoxygenase. The identity of the cloned 12-lipoxygenase was assured by expression in a mammalian cell line (COS cells). Human platelet 12-lipoxygenase has been difficult to purify to homogeneity. The cloning of this cDNA will increase the possibilities to elucidate the structure and function of this enzyme

cDNA sequence quality data - Budding yeast cDNA sequencing project | LSDB Archive [Life Science Database Archive metadata

Lifescience Database Archive (English)

Full Text Available List Contact us Budding yeast cDNA sequencing project cDNA sequence quality data Data detail Data name cDNA sequence quality... data DOI 10.18908/lsdba.nbdc00838-003 Description of data contents Phred's quality score. P...tion Download License Update History of This Database Site Policy | Contact Us cDNA sequence quality

An unbiased expression screen for synaptogenic proteins identifies the LRRTM protein family as synaptic organizers.

Science.gov (United States)

Linhoff, Michael W; Laurén, Juha; Cassidy, Robert M; Dobie, Frederick A; Takahashi, Hideto; Nygaard, Haakon B; Airaksinen, Matti S; Strittmatter, Stephen M; Craig, Ann Marie

2009-03-12

Delineating the molecular basis of synapse development is crucial for understanding brain function. Cocultures of neurons with transfected fibroblasts have demonstrated the synapse-promoting activity of candidate molecules. Here, we performed an unbiased expression screen for synaptogenic proteins in the coculture assay using custom-made cDNA libraries. Reisolation of NGL-3/LRRC4B and neuroligin-2 accounts for a minority of positive clones, indicating that current understanding of mammalian synaptogenic proteins is incomplete. We identify LRRTM1 as a transmembrane protein that induces presynaptic differentiation in contacting axons. All four LRRTM family members exhibit synaptogenic activity, LRRTMs localize to excitatory synapses, and artificially induced clustering of LRRTMs mediates postsynaptic differentiation. We generate LRRTM1(-/-) mice and reveal altered distribution of the vesicular glutamate transporter VGLUT1, confirming an in vivo synaptic function. These results suggest a prevalence of LRR domain proteins in trans-synaptic signaling and provide a cellular basis for the reported linkage of LRRTM1 to handedness and schizophrenia.

UNBIASED ESTIMATORS OF SPECIFIC CONNECTIVITY

Directory of Open Access Journals (Sweden)

Jean-Paul Jernot

2011-05-01

Full Text Available This paper deals with the estimation of the specific connectivity of a stationary random set in IRd. It turns out that the "natural" estimator is only asymptotically unbiased. The example of a boolean model of hypercubes illustrates the amplitude of the bias produced when the measurement field is relatively small with respect to the range of the random set. For that reason unbiased estimators are desired. Such an estimator can be found in the literature in the case where the measurement field is a right parallelotope. In this paper, this estimator is extended to apply to measurement fields of various shapes, and to possess a smaller variance. Finally an example from quantitative metallography (specific connectivity of a population of sintered bronze particles is given.

Ocular Proteomics with Emphasis on Two-Dimensional Gel Electrophoresis and Mass Spectrometry

Directory of Open Access Journals (Sweden)

Honoré Bent

2010-01-01

Full Text Available Abstract The intention of this review is to provide an overview of current methodologies employed in the rapidly developing field of ocular proteomics with emphasis on sample preparation, two-dimensional polyacrylamide gel electrophoresis (2D-PAGE and mass spectrometry (MS. Appropriate sample preparation for the diverse range of cells and tissues of the eye is essential to ensure reliable results. Current methods of protein staining for 2D-PAGE, protein labelling for two-dimensional difference gel electrophoresis, gel-based expression analysis and protein identification by MS are summarised. The uses of gel-free MS-based strategies (MuDPIT, iTRAQ, ICAT and SILAC are also discussed. Proteomic technologies promise to shed new light onto ocular disease processes that could lead to the discovery of strong novel biomarkers and therapeutic targets useful in many ophthalmic conditions.

Unbiased Sampling and Meshing of Isosurfaces

KAUST Repository

Yan, Dongming

2014-05-07

In this paper, we present a new technique to generate unbiased samples on isosurfaces. An isosurface, F(x,y,z) = c , of a function, F , is implicitly defined by trilinear interpolation of background grid points. The key idea of our approach is that of treating the isosurface within a grid cell as a graph (height) function in one of the three coordinate axis directions, restricted to where the slope is not too high, and integrating / sampling from each of these three. We use this unbiased sampling algorithm for applications in Monte Carlo integration, Poisson-disk sampling, and isosurface meshing.

Unbiased Sampling and Meshing of Isosurfaces

KAUST Repository

Yan, Dongming; Wallner, Johannes; Wonka, Peter

2014-01-01

In this paper, we present a new technique to generate unbiased samples on isosurfaces. An isosurface, F(x,y,z) = c , of a function, F , is implicitly defined by trilinear interpolation of background grid points. The key idea of our approach is that of treating the isosurface within a grid cell as a graph (height) function in one of the three coordinate axis directions, restricted to where the slope is not too high, and integrating / sampling from each of these three. We use this unbiased sampling algorithm for applications in Monte Carlo integration, Poisson-disk sampling, and isosurface meshing.

Constructing and detecting a cDNA library for mites.

Science.gov (United States)

Hu, Li; Zhao, YaE; Cheng, Juan; Yang, YuanJun; Li, Chen; Lu, ZhaoHui

2015-10-01

RNA extraction and construction of complementary DNA (cDNA) library for mites have been quite challenging due to difficulties in acquiring tiny living mites and breaking their hard chitin. The present study is to explore a better method to construct cDNA library for mites that will lay the foundation on transcriptome and molecular pathogenesis research. We selected Psoroptes cuniculi as an experimental subject and took the following steps to construct and verify cDNA library. First, we combined liquid nitrogen grinding with TRIzol for total RNA extraction. Then, switching mechanism at 5' end of the RNA transcript (SMART) technique was used to construct full-length cDNA library. To evaluate the quality of cDNA library, the library titer and recombination rate were calculated. The reliability of cDNA library was detected by sequencing and analyzing positive clones and genes amplified by specific primers. The results showed that the RNA concentration was 836 ng/μl and the absorbance ratio at 260/280 nm was 1.82. The library titer was 5.31 × 10(5) plaque-forming unit (PFU)/ml and the recombination rate was 98.21%, indicating that the library was of good quality. In the 33 expressed sequence tags (ESTs) of P. cuniculi, two clones of 1656 and 1658 bp were almost identical with only three variable sites detected, which had an identity of 99.63% with that of Psoroptes ovis, indicating that the cDNA library was reliable. Further detection by specific primers demonstrated that the 553-bp Pso c II gene sequences of P. cuniculi had an identity of 98.56% with those of P. ovis, confirming that the cDNA library was not only reliable but also feasible.

[Primary culture of cat intestinal epithelial cell and construction of its cDNA library].

Science.gov (United States)

Ye, L; Gui-Hua, Z; Kun, Y; Hong-Fa, W; Ting, X; Gong-Zhen, L; Wei-Xia, Z; Yong, C

2017-04-12

Objective To establish the primary cat intestinal epithelial cells (IECs) culture methods and construct the cDNA library for the following yeast two-hybrid experiment, so as to screen the virulence interaction factors among the final host. Methods The primary cat IECs were cultured by the tissue cultivation and combined digestion with collagenase XI and dispase I separately. Then the cat IECs cultured was identified with the morphological observation and cyto-keratin detection, by using goat anti-cyto-keratin monoclonal antibodies. The mRNA of cat IECs was isolated and used as the template to synthesize the first strand cDNA by SMART™ technology, and then the double-strand cDNAs were acquired by LD-PCR, which were subsequently cloned into the plasmid PGADT7-Rec to construct yeast two-hybrid cDNA library in the yeast strain Y187 by homologous recombination. Matchmaker™ Insert Check PCR was used to detect the size distribution of cDNA fragments after the capacity calculation of the cDNA library. Results The comparison of the two cultivation methods indicated that the combined digestion of collagenase XI and dispase I was more effective than the tissue cultivation. The cat IECs system of continuous culture was established and the cat IECs with high purity were harvested for constructing the yeast two-hybrid cDNA library. The library contained 1.1×10 6 independent clones. The titer was 2.8×10 9 cfu/ml. The size of inserted fragments was among 0.5-2.0 kb. Conclusion The yeast two-hybrid cDNA library of cat IECs meets the requirements of further screen research, and this study lays the foundation of screening the Toxoplasma gondii virulence interaction factors among the cDNA libraries of its final hosts.

Construction of a T7 Human Lung Cancer cDNA Library

Directory of Open Access Journals (Sweden)

Wentao YUE

2008-10-01

Full Text Available Background and objective Currently, only a limited numbers of tumor markers for non small lung cancer (NSCLC diagnosis, new biomarker, such as serum autoantibody may improve the early detection of lung cancer. Our objective is construction human lung squamous carcinoma and adenocarcinoma T7 phage display cDNA library from the tissues of NSCLC patients. Methods mRNA was isolated from a pool of total RNA extract from NSCLC tissues obtained from 5 adenocarcinomas and 5 squamous carcinomas, and then mRNA was reverse transcribed into double stranded cDNA. After digestion, the cDNA was inserted into T7Select 10-3 vector. The phage display cDNA library was constructed by package reaction in vitro and plate proliferation. Plaque assay and PCR were used to evaluate the library.Results Two T7 phage display cDNA library were established. Plaque assay show the titer of lung squamas carcinoma library was 1.8×106 pfu, and the adenocarcinoma library was 5×106 pfu. The phage titer of the amplified library were 3.2×1010 pfu/mL and 2.5×1010 pfu/mL. PCR amplification of random plaque show insert ratio were 100% (24/24 in adenocarcinoma library and 95.8% in human lung squamas carcinoma library (23/24. Insert range from 300 bp to 1 500 bp. Conclusion Two phage display cDNA library from NSCLC were constructed.

Analysis of coupled mass transfer and sol-gel reaction in a two-phase system

NARCIS (Netherlands)

Castelijns, H.J.; Huinink, H.P.; Pel, L.; Zitha, P.L.J.

2006-01-01

The coupled mass transfer and chemical reactions of a gel-forming compound in a two-phase system were studied in detail. Tetra-methyl-ortho-silicate (TMOS) is often used as a precursor in sol-gel chemistry to produce silica gels in aqueous systems. TMOS can also be mixed with many hydrocarbons

Molecular cloning and mammalian expression of human beta 2-glycoprotein I cDNA

DEFF Research Database (Denmark)

Kristensen, Torsten; Schousboe, Inger; Boel, Espen

1991-01-01

Human β2-glycoprotein (β2gpI) cDNA was isolated from a liver cDNA library and sequenced. The cDNA encoded a 19-residue hydrophobic signal peptide followed by the mature β2gpI of 326 amino acid residues. In liver and in the hepatoma cell line HepG2 there are two mRNA species of about 1.4 and 4.3 kb......, respectively, hybridizing specifically with the β2gpI cDNA. Upon isoelectric focusing, recombinant β2gpI obtained from expression of β2gpI cDNA in baby hamster kidney cells showed the same pattern of bands as β2gpI isolated from plasma, and at least 5 polypeptides were visible...

Cloning and functional expression of a human pancreatic islet glucose-transporter cDNA

International Nuclear Information System (INIS)

Permutt, M.A.; Koranyi, L.; Keller, K.; Lacy, P.E.; Scharp, D.W.; Mueckler, M.

1989-01-01

Previous studies have suggested that pancreatic islet glucose transport is mediated by a high-K m , low-affinity facilitated transporter similar to that expressed in liver. To determine the relationship between islet and liver glucose transporters, liver-type glucose-transporter cDNA clones were isolated from a human liver cDNA library. The liver-type glucose-transporter cDNA clone hybridized to mRNA transcripts of the same size in human liver and pancreatic islet RNA. A cDNA library was prepared from purified human pancreatic islet tissue and screened with human liver-type glucose-transporter cDNA. The authors isolated two overlapping cDNA clones encompassing 2600 base pairs, which encode a pancreatic islet protein identical in sequence to that of the putative liver-type glucose-transporter protein. Xenopus oocytes injected with synthetic mRNA transcribed from a full-length cDNA construct exhibited increased uptake of 2-deoxyglucose, confirming the functional identity of the clone. These cDNA clones can now be used to study regulation of expression of the gene and to assess the role of inherited defects in this gene as a candidate for inherited susceptibility to non-insulin-dependent diabetes mellitus

Brain cDNA clone for human cholinesterase

International Nuclear Information System (INIS)

McTiernan, C.; Adkins, S.; Chatonnet, A.; Vaughan, T.A.; Bartels, C.F.; Kott, M.; Rosenberry, T.L.; La Du, B.N.; Lockridge, O.

1987-01-01

A cDNA library from human basal ganglia was screened with oligonucleotide probes corresponding to portions of the amino acid sequence of human serum cholinesterase. Five overlapping clones, representing 2.4 kilobases, were isolated. The sequenced cDNA contained 207 base pairs of coding sequence 5' to the amino terminus of the mature protein in which there were four ATG translation start sites in the same reading frame as the protein. Only the ATG coding for Met-(-28) lay within a favorable consensus sequence for functional initiators. There were 1722 base pairs of coding sequence corresponding to the protein found circulating in human serum. The amino acid sequence deduced from the cDNA exactly matched the 574 amino acid sequence of human serum cholinesterase, as previously determined by Edman degradation. Therefore, our clones represented cholinesterase rather than acetylcholinesterase. It was concluded that the amino acid sequences of cholinesterase from two different tissues, human brain and human serum, were identical. Hybridization of genomic DNA blots suggested that a single gene, or very few genes coded for cholinesterase

Sequence of a cloned cDNA encoding human ribosomal protein S11

Energy Technology Data Exchange (ETDEWEB)

Lott, J B; Mackie, G A

1988-02-11

The authors have isolated a cloned cDNA that encodes human ribosomal protein (rp) S11 by screening a human fibroblast cDNA library with a labelled 204 bp DNA fragment encompassing residues 212-416 of pRS11, a rat rp Sll cDNA clone. The human rp S11 cloned cDNA consists of 15 residues of the 5' leader, the entire coding sequence and all 51 residues of the 3' untranslated region. The predicted amino acid sequence of 158 residues is identical to rat rpS11. The nucleotide sequence in the coding region differs, however, from that in rat in the first position in two codons and in the third position in 44 codons.

Expression analysis of a ''Cucurbita'' cDNA encoding endonuclease

International Nuclear Information System (INIS)

Szopa, J.

1995-01-01

The nuclear matrices of plant cell nuclei display intrinsic nuclease activity which consists in nicking supercoiled DNA. A cDNA encoding a 32 kDa endonuclease has been cloned and sequenced. The nucleotide and deduced amino-acid sequences show high homology to known 14-3-3-protein sequences from other sources. The amino-acid sequence shows agreement with consensus sequences for potential phosphorylation by protein kinase A and C and for calcium, lipid and membrane-binding sites. The nucleotide-binding site is also present within the conserved part of the sequence. By Northern blot analysis, the differential expression of the corresponding mRNA was detected; it was the strongest in sink tissues. The endonuclease activity found on DNA-polyacrylamide gel electrophoresis coincided with mRNA content and was the highest in tuber. (author). 22 refs, 6 figs

Ultralow-density SiO2 aerogels prepared by a two-step sol-gel process

International Nuclear Information System (INIS)

Wang Jue; Li Qing; Shen Jun; Zhou Bin; Chen Lingyan; Jiang; Weiyang

1996-01-01

Low density SiO 2 gels are prepared by a two-step sol-gel process from TEOS. The influence of various solution ratios on the gelation process is investigated. The comparative characterization of gels using different solvent, such as ethanol, acetone and methyl cyanide, is also given. The ultralow-density SiO 2 aerogels with density less than 10 kg/m 3 are prepared by CO 2 supercritical drying technique. The structure difference between SiO 2 aerogels prepared by conventional single-step process and the two-step process is also presented

Quantum process reconstruction based on mutually unbiased basis

International Nuclear Information System (INIS)

Fernandez-Perez, A.; Saavedra, C.; Klimov, A. B.

2011-01-01

We study a quantum process reconstruction based on the use of mutually unbiased projectors (MUB projectors) as input states for a D-dimensional quantum system, with D being a power of a prime number. This approach connects the results of quantum-state tomography using mutually unbiased bases with the coefficients of a quantum process, expanded in terms of MUB projectors. We also study the performance of the reconstruction scheme against random errors when measuring probabilities at the MUB projectors.

Two-dimensional gel electrophoresis analysis of different parts of ...

African Journals Online (AJOL)

Two-dimensional gel electrophoresis analysis of different parts of Panax quinquefolius L. root. ... From these results it was concluded that proteomic analysis method was an effective way to identify the different parts of quinquefolius L. root. These findings may contribute to further understanding of the physiological ...

Extracting information from two-dimensional electrophoresis gels by partial least squares regression

DEFF Research Database (Denmark)

Jessen, Flemming; Lametsch, R.; Bendixen, E.

2002-01-01

of all proteins/spots in the gels. In the present study it is demonstrated how information can be extracted by multivariate data analysis. The strategy is based on partial least squares regression followed by variable selection to find proteins that individually or in combination with other proteins vary......Two-dimensional gel electrophoresis (2-DE) produces large amounts of data and extraction of relevant information from these data demands a cautious and time consuming process of spot pattern matching between gels. The classical approach of data analysis is to detect protein markers that appear...... or disappear depending on the experimental conditions. Such biomarkers are found by comparing the relative volumes of individual spots in the individual gels. Multivariate statistical analysis and modelling of 2-DE data for comparison and classification is an alternative approach utilising the combination...

cDNA encoding a polypeptide including a hevein sequence

Energy Technology Data Exchange (ETDEWEB)

Raikhel, Natasha V. (Okemos, MI); Broekaert, Willem F. (Dilbeek, BE); Chua, Nam-Hai (Scarsdale, NY); Kush, Anil (New York, NY)

1993-02-16

A cDNA clone (HEV1) encoding hevein was isolated via polymerase chain reaction (PCR) using mixed oligonucleotides corresponding to two regions of hevein as primers and a Hevea brasiliensis latex cDNA library as a template. HEV1 is 1018 nucleotides long and includes an open reading frame of 204 amino acids. The deduced amino acid sequence contains a pu GOVERNMENT RIGHTS This application was funded under Department of Energy Contract DE-AC02-76ER01338. The U.S. Government has certain rights under this application and any patent issuing thereon.

The function analysis of full-length cDNA sequence from IRM-2 mouse cDNA library

International Nuclear Information System (INIS)

Wang Qin; Liu Xiaoqiu; Xu Chang; Du Liqing; Sun Zhijuan; Wang Yan; Liu Qiang; Song Li; Li Jin; Fan Feiyue

2013-01-01

Objective: To identify the function of full-length cDNA sequence from IRM-2 mouse cDNA library. Methods: Full-length cDNA products were amplified by PCR from IRM-2 mouse cDNA library according to twenty-one pieces of expressed sequence tag. The expression of full-length cDNAs were detected after mouse embryonic fibroblasts were exposed to 6.5 Gy γ-ray radiation. And the effect on the growth of radiosensitivity cells AT5B1VA transfected with full-length cDNAs was investigated. Results: The expression of No.4, 5 and 2 full-length cDNAs from IRM-2 mouse were higher than that of parental ICR and 615 mouse after mouse embryonic fibroblasts irradiated with γ-ray radiation. And the survival rate of AT5B1VA cells transfected with No.4, 5 and 2 full-length cDNAs was high. Conclusion: No.4, 5 and 2 full-length cDNAs of IRM-2 mouse are of high radioresistance. (authors)

[cDNA library construction from panicle meristem of finger millet].

Science.gov (United States)

Radchuk, V; Pirko, Ia V; Isaenkov, S V; Emets, A I; Blium, Ia B

2014-01-01

The protocol for production of full-size cDNA using SuperScript Full-Length cDNA Library Construction Kit II (Invitrogen) was tested and high quality cDNA library from meristematic tissue of finger millet panicle (Eleusine coracana (L.) Gaertn) was created. The titer of obtained cDNA library comprised 3.01 x 10(5) CFU/ml in avarage. In average the length of cDNA insertion consisted about 1070 base pairs, the effectivity of cDNA fragment insertions--99.5%. The selective sequencing of cDNA clones from created library was performed. The sequences of cDNA clones were identified with usage of BLAST-search. The results of cDNA library analysis and selective sequencing represents prove good functionality and full length character of inserted cDNA clones. Obtained cDNA library from meristematic tissue of finger millet panicle represents good and valuable source for isolation and identification of key genes regulating metabolism and meristematic development and for mining of new molecular markers to conduct out high quality genetic investigations and molecular breeding as well.

Molecular cloning of lupin leghemoglobin cDNA

DEFF Research Database (Denmark)

Konieczny, A; Jensen, E O; Marcker, K A

1987-01-01

Poly(A)+ RNA isolated from root nodules of yellow lupin (Lupinus luteus, var. Ventus) has been used as a template for the construction of a cDNA library. The ds cDNA was synthesized and inserted into the Hind III site of plasmid pBR 322 using synthetic Hind III linkers. Clones containing sequences...... specific for nodules were selected by differential colony hybridization using 32P-labeled cDNA synthesized either from nodule poly(A)+ RNA or from poly(A)+ RNA of uninfected root as probes. Among the recombinant plasmids, the cDNA gene for leghemoglobin was identified. The protein structure derived from...... its nucleotide sequence was consistent with known amino acid sequence of lupin Lb II. The cloned lupin Lb cDNA hybridized to poly(A)+ RNA from nodules only, which is in accordance with the general concept, that leghemoglobin is expressed exclusively in nodules. Udgivelsesdato: 1987-null...

RICD: A rice indica cDNA database resource for rice functional genomics

Directory of Open Access Journals (Sweden)

Zhang Qifa

2008-11-01

Full Text Available Abstract Background The Oryza sativa L. indica subspecies is the most widely cultivated rice. During the last few years, we have collected over 20,000 putative full-length cDNAs and over 40,000 ESTs isolated from various cDNA libraries of two indica varieties Guangluai 4 and Minghui 63. A database of the rice indica cDNAs was therefore built to provide a comprehensive web data source for searching and retrieving the indica cDNA clones. Results Rice Indica cDNA Database (RICD is an online MySQL-PHP driven database with a user-friendly web interface. It allows investigators to query the cDNA clones by keyword, genome position, nucleotide or protein sequence, and putative function. It also provides a series of information, including sequences, protein domain annotations, similarity search results, SNPs and InDels information, and hyperlinks to gene annotation in both The Rice Annotation Project Database (RAP-DB and The TIGR Rice Genome Annotation Resource, expression atlas in RiceGE and variation report in Gramene of each cDNA. Conclusion The online rice indica cDNA database provides cDNA resource with comprehensive information to researchers for functional analysis of indica subspecies and for comparative genomics. The RICD database is available through our website http://www.ncgr.ac.cn/ricd.

Fluctuations and symmetries in two-dimensional active gels.

Science.gov (United States)

Sarkar, N; Basu, A

2011-04-01

Motivated by the unique physical properties of biological active matter, e.g., cytoskeletal dynamics in eukaryotic cells, we set up effective two-dimensional (2d) coarse-grained hydrodynamic equations for the dynamics of thin active gels with polar or nematic symmetries. We use the well-known three-dimensional (3d) descriptions (K. Kruse et al., Eur. Phys. J. E 16, 5 (2005); A. Basu et al., Eur. Phys. J. E 27, 149 (2008)) for thin active-gel samples confined between parallel plates with appropriate boundary conditions to derive the effective 2d constitutive relations between appropriate thermodynamic fluxes and generalised forces for small deviations from equilibrium. We consider three distinct cases, characterised by spatial symmetries and boundary conditions, and show how such considerations dictate the structure of the constitutive relations. We use these to study the linear instabilities, calculate the correlation functions and the diffusion constant of a small tagged particle, and elucidate their dependences on the activity or nonequilibrium drive.

Direct metagenomic detection of viral pathogens in nasal and fecal specimens using an unbiased high-throughput sequencing approach.

Directory of Open Access Journals (Sweden)

Shota Nakamura

Full Text Available With the severe acute respiratory syndrome epidemic of 2003 and renewed attention on avian influenza viral pandemics, new surveillance systems are needed for the earlier detection of emerging infectious diseases. We applied a "next-generation" parallel sequencing platform for viral detection in nasopharyngeal and fecal samples collected during seasonal influenza virus (Flu infections and norovirus outbreaks from 2005 to 2007 in Osaka, Japan. Random RT-PCR was performed to amplify RNA extracted from 0.1-0.25 ml of nasopharyngeal aspirates (N = 3 and fecal specimens (N = 5, and more than 10 microg of cDNA was synthesized. Unbiased high-throughput sequencing of these 8 samples yielded 15,298-32,335 (average 24,738 reads in a single 7.5 h run. In nasopharyngeal samples, although whole genome analysis was not available because the majority (>90% of reads were host genome-derived, 20-460 Flu-reads were detected, which was sufficient for subtype identification. In fecal samples, bacteria and host cells were removed by centrifugation, resulting in gain of 484-15,260 reads of norovirus sequence (78-98% of the whole genome was covered, except for one specimen that was under-detectable by RT-PCR. These results suggest that our unbiased high-throughput sequencing approach is useful for directly detecting pathogenic viruses without advance genetic information. Although its cost and technological availability make it unlikely that this system will very soon be the diagnostic standard worldwide, this system could be useful for the earlier discovery of novel emerging viruses and bioterrorism, which are difficult to detect with conventional procedures.

RFLP for Duchenne muscular dystrophy cDNA clone 44-1

Energy Technology Data Exchange (ETDEWEB)

Laing, N G; Siddique, T; Bartlett, R J; Yamaoka, L H; Chen, J C; Walker, A P; Hung, W Y; Roses, A D [Duke Univ. Medical Center, Durham, NC (USA)

1988-07-25

Clone 44-1 is one of six cDNA clones which comprise the cDNA for the Duchenne muscular dystrophy gene. It is a 0.9kb fragment in the EcoR1 site of Bluescript. Taq1 (TlCGA) identifies two alleles with bands at 6.8 and 5.7kb, as well as four constant bands at 4.8, 3.9, 3.5 and 2.5kb. Its frequency was studied in 62 unrelated individuals. Mendelian inheritance was demonstrated in one three generation and three two generation informative families, 26 individuals. There were no problems on RFLP analysis under normal stringency conditions.

Triangulation based inclusion probabilities: a design-unbiased sampling approach

OpenAIRE

Fehrmann, Lutz; Gregoire, Timothy; Kleinn, Christoph

2011-01-01

A probabilistic sampling approach for design-unbiased estimation of area-related quantitative characteristics of spatially dispersed population units is proposed. The developed field protocol includes a fixed number of 3 units per sampling location and is based on partial triangulations over their natural neighbors to derive the individual inclusion probabilities. The performance of the proposed design is tested in comparison to fixed area sample plots in a simulation with two forest stands. ...

Sequence of a cDNA encoding turtle high mobility group 1 protein.

Science.gov (United States)

Zheng, Jifang; Hu, Bi; Wu, Duansheng

2005-07-01

In order to understand sequence information about turtle HMG1 gene, a cDNA encoding HMG1 protein of the Chinese soft-shell turtle (Pelodiscus sinensis) was amplified by RT-PCR from kidney total RNA, and was cloned, sequenced and analyzed. The results revealed that the open reading frame (ORF) of turtle HMG1 cDNA is 606 bp long. The ORF codifies 202 amino acid residues, from which two DNA-binding domains and one polyacidic region are derived. The DNA-binding domains share higher amino acid identity with homologues sequences of chicken (96.5%) and mammalian (74%) than homologues sequence of rainbow trout (67%). The polyacidic region shows 84.6% amino acid homology with the equivalent region of chicken HMG1 cDNA. Turtle HMG1 protein contains 3 Cys residues located at completely conserved positions. Conservation in sequence and structure suggests that the functions of turtle HMG1 cDNA may be highly conserved during evolution. To our knowledge, this is the first report of HMG1 cDNA sequence in any reptilian.

Rationalization of reduced penetration of drugs through ceramide gel phase membrane.

Science.gov (United States)

Paloncýová, Markéta; DeVane, Russell H; Murch, Bruce P; Berka, Karel; Otyepka, Michal

2014-11-25

Since computing resources have advanced enough to allow routine molecular simulation studies of drug molecules interacting with biologically relevant membranes, a considerable amount of work has been carried out with fluid phospholipid systems. However, there is very little work in the literature on drug interactions with gel phase lipids. This poses a significant limitation for understanding permeation through the stratum corneum where the primary pathway is expected to be through a highly ordered lipid matrix. To address this point, we analyzed the interactions of p-aminobenzoic acid (PABA) and its ethyl (benzocaine) and butyl (butamben) esters with two membrane bilayers, which differ in their fluidity at ambient conditions. We considered a dioleoylphosphatidylcholine (DOPC) bilayer in a fluid state and a ceramide 2 (CER2, ceramide NS) bilayer in a gel phase. We carried out unbiased (100 ns long) and biased z-constraint molecular dynamics simulations and calculated the free energy profiles of all molecules along the bilayer normal. The free energy profiles converged significantly slower for the gel phase. While the compounds have comparable affinities for both membranes, they exhibit penetration barriers almost 3 times higher in the gel phase CER2 bilayer. This elevated barrier and slower diffusion in the CER2 bilayer, which are caused by the high ordering of CER2 lipid chains, explain the low permeability of the gel phase membranes. We also compared the free energy profiles from MD simulations with those obtained from COSMOmic. This method provided the same trends in behavior for the guest molecules in both bilayers; however, the penetration barriers calculated by COSMOmic did not differ between membranes. In conclusion, we show how membrane fluid properties affect the interaction of drug-like molecules with membranes.

Second-strand cDNA synthesis: classical method

International Nuclear Information System (INIS)

Gubler, U.

1987-01-01

The classical scheme for the synthesis of double-stranded cDNA as it was reported in 1976 is described. Reverse transcription of mRNA with oligo(dT) as the primer generates first strands with a small loop at the 3' end of the cDNA (the end that corresponds to the 5' end of the mRNA). Subsequent removal of the mRNA by alkaline hydrolysis leaves single-stranded cDNA molecules again with a small 3' loop. This loop can be used by either reverse transcriptase or Klenow fragment of DNA polymerase I as a primer for second-strand synthesis. The resulting products are double-stranded cDNA molecules that are covalently closed at the end corresponding to the 5' end of the original mRNA. Subsequent cleavage of the short piece of single-stranded cDNA within the loop with the single-strand-specific S 1 nuclease generate open double-stranded molecules that can be used for molecular cloning in plasmids or in phage. Useful variations of this scheme have been described

LEDGF/p75 Deficiency Increases Deletions at the HIV-1 cDNA Ends.

Science.gov (United States)

Bueno, Murilo T D; Reyes, Daniel; Llano, Manuel

2017-09-15

Processing of unintegrated linear HIV-1 cDNA by the host DNA repair system results in its degradation and/or circularization. As a consequence, deficient viral cDNA integration generally leads to an increase in the levels of HIV-1 cDNA circles containing one or two long terminal repeats (LTRs). Intriguingly, impaired HIV-1 integration in LEDGF/p75-deficient cells does not result in a correspondent increase in viral cDNA circles. We postulate that increased degradation of unintegrated linear viral cDNA in cells lacking the lens epithelium-derived growth factor (LEDGF/p75) account for this inconsistency. To evaluate this hypothesis, we characterized the nucleotide sequence spanning 2-LTR junctions isolated from LEDGF/p75-deficient and control cells. LEDGF/p75 deficiency resulted in a significant increase in the frequency of 2-LTRs harboring large deletions. Of note, these deletions were dependent on the 3' processing activity of integrase and were not originated by aberrant reverse transcription. Our findings suggest a novel role of LEDGF/p75 in protecting the unintegrated 3' processed linear HIV-1 cDNA from exonucleolytic degradation.

Construction of Infectious cDNA Clone of a Chrysanthemum stunt viroid Korean Isolate

Directory of Open Access Journals (Sweden)

Ju-Yeon Yoon

2014-03-01

Full Text Available Chrysanthemum stunt viroid (CSVd, a noncoding infectious RNA molecule, causes seriously economic losses of chrysanthemum for 3 or 4 years after its first infection. Monomeric cDNA clones of CSVd isolate SK1 (CSVd-SK1 were constructed in the plasmids pGEM-T easy vector and pUC19 vector. Linear positive-sense transcripts synthesized in vitro from the full-length monomeric cDNA clones of CSVd-SK1 could infect systemically tomato seedlings and chrysanthemum plants, suggesting that the linear CSVd RNA transcribed from the cDNA clones could be replicated as efficiently as circular CSVd in host species. However, direct inoculation of plasmid cDNA clones containing full-length monomeric cDNA of CSVd-SK1 failed to infect tomato and chrysanthemum and linear negative-sense transcripts from the plasmid DNAs were not infectious in the two plant species. The cDNA sequences of progeny viroid in systemically infected tomato and chrysanthemum showed a few substitutions at a specific nucleotide position, but there were no deletions and insertions in the sequences of the CSVd progeny from tomato and chrysanthemum plants.

Preparation of a differentially expressed, full-length cDNA expression library by RecA-mediated triple-strand formation with subtractively enriched cDNA fragments

NARCIS (Netherlands)

Hakvoort, T. B.; Spijkers, J. A.; Vermeulen, J. L.; Lamers, W. H.

1996-01-01

We have developed a fast and general method to obtain an enriched, full-length cDNA expression library with subtractively enriched cDNA fragments. The procedure relies on RecA-mediated triple-helix formation of single-stranded cDNA fragments with a double-stranded cDNA plasmid library. The complexes

Structural and optical studies of nano-structure silica gel doped with different rare earth elements, prepared by two different sol -gel techniques

International Nuclear Information System (INIS)

Battisha, I.K.; El Beyally, A.; Seliman, S.I.; El Nahrawi, A.S.

2005-01-01

Structural and optical characteristics of pure silica gel (silica-xerogel, SiO 2 ) and doped with different concentrations ranging from 1 up to 6% of some rare earth (REEs) ions such as, praseodymium Pr +3 ,and Europium Eu +3 , Erbium Er +3 and Holmium Ho +3 , ions, in the form of thin film and monolith materials were prepared by sol - gel technique, Using tetra-ethoxysilane as precursor materials, which are of particular interest for sol-gel integrated optics applications. Some structural and optical features of sol-gel derived monolith and thin films are analyzed and compared, namely the structure of nano-particle monolith and thin film silica-gel samples, based on X-ray diffraction (XRD). The types of structural information obtainable are compared in detail. It is show that the XRD spectra of a-cristobalite are obtained for the two type materials and even by doping with the four REEs ions. Optical measurements of monolith and thin films were also studied and compared, the normal transmission and specular reflection were measured. The refractive index were calculated and discussed

Avoiding acidic region streaking in two-dimensional gel electrophoresis: case study with two bacterial whole cell protein extracts.

Science.gov (United States)

Roy, Arnab; Varshney, Umesh; Pal, Debnath

2014-09-01

Acidic region streaking (ARS) is one of the lacunae in two-dimensional gel electrophoresis (2DE) of bacterial proteome. This streaking is primarily caused by nucleic acid (NuA) contamination and poses major problem in the downstream processes like image analysis and protein identification. Although cleanup and nuclease digestion are practiced as remedial options, these strategies may incur loss in protein recovery and perform incomplete removal of NuA. As a result, ARS has remained a common observation across publications, including the recent ones. In this work, we demonstrate how ultrasound wave can be used to shear NuA in plain ice-cooled water, facilitating the elimination of ARS in the 2DE gels without the need for any additional sample cleanup tasks. In combination with a suitable buffer recipe, IEF program and frequent paper-wick changing approach, we are able to reproducibly demonstrate the production of clean 2DE gels with improved protein recovery and negligible or no ARS. We illustrate our procedure using whole cell protein extracts from two diverse organisms, Escherichia coli and Mycobacterium smegmatis. Our designed protocols are straightforward and expected to provide good 2DE gels without ARS, with comparable times and significantly lower cost.

Sequence of human protamine 2 cDNA

Energy Technology Data Exchange (ETDEWEB)

Domenjoud, L; Fronia, C; Uhde, F; Engel, W [Universitaet Goettingen (West Germany)

1988-08-11

The authors report the cloning and sequencing of a cDNA clone for human protamine 2 (hp2), isolated from a human testis cDNA library cloned in the vector {lambda}-gt11. A 66mer oligonucleotide, that corresponds to an amino acid sequence which is highly conserved between hp2 and mouse protamine 2 (mp2) served as hybridization probe. The homology between the amino acid sequence deduced from our cDNA and the published amino acid sequence for hp2 is 100%.

Sources of variability among replicate samples separated by two-dimensional gel electrophoresis.

Science.gov (United States)

Bland, Alison M; Janech, Michael G; Almeida, Jonas S; Arthur, John M

2010-04-01

Two-dimensional gel electrophoresis (2DE) offers high-resolution separation for intact proteins. However, variability in the appearance of spots can limit the ability to identify true differences between conditions. Variability can occur at a number of levels. Individual samples can differ because of biological variability. Technical variability can occur during protein extraction, processing, or storage. Another potential source of variability occurs during analysis of the gels and is not a result of any of the causes of variability named above. We performed a study designed to focus only on the variability caused by analysis. We separated three aliquots of rat left ventricle and analyzed differences in protein abundance on the replicate 2D gels. As the samples loaded on each gel were identical, differences in protein abundance are caused by variability in separation or interpretation of the gels. Protein spots were compared across gels by quantile values to determine differences. Fourteen percent of spots had a maximum difference in intensity of 0.4 quantile values or more between replicates. We then looked individually at the spots to determine the cause of differences between the measured intensities. Reasons for differences were: failure to identify a spot (59%), differences in spot boundaries (13%), difference in the peak height (6%), and a combination of these factors (21). This study demonstrates that spot identification and characterization make major contributions to variability seen with 2DE. Methods to highlight why measured protein spot abundance is different could reduce these errors.

[Construction of forward and reverse subtracted cDNA libraries between muscle tissue of Meishan and Landrace pigs].

Science.gov (United States)

Xu, De-Quan; Zhang, Yi-Bing; Xiong, Yuan-Zhu; Gui, Jian-Fang; Jiang, Si-Wen; Su, Yu-Hong

2003-07-01

Using suppression subtractive hybridization (SSH) technique, forward and reverse subtracted cDNA libraries were constructed between Longissimus muscles from Meishan and Landrace pigs. A housekeeping gene, G3PDH, was used to estimate the efficiency of subtractive cDNA. In two cDNA libraries, G3PDH was subtracted very efficiently at appropriate 2(10) and 2(5) folds, respectively, indicating that some differentially expressed genes were also enriched at the same folds and the two subtractive cDNA libraries were very successful. A total of 709 and 673 positive clones were isolated from forward and reverse subtracted cDNA libraries, respectively. Analysis of PCR showed that most of all plasmids in the clones contained 150-750 bp inserts. The construction of subtractive cDNA libraries between muscle tissue from different pig breeds laid solid foundations for isolating and identifying the genes determining muscle growth and meat quality, which will be important to understand the mechanism of muscle growth, determination of meat quality and practice of molecular breeding.

Display of a Maize cDNA library on baculovirus infected insect cells

Directory of Open Access Journals (Sweden)

Jones Ian M

2008-08-01

Full Text Available Abstract Background Maize is a good model system for cereal crop genetics and development because of its rich genetic heritage and well-characterized morphology. The sequencing of its genome is well advanced, and new technologies for efficient proteomic analysis are needed. Baculovirus expression systems have been used for the last twenty years to express in insect cells a wide variety of eukaryotic proteins that require complex folding or extensive posttranslational modification. More recently, baculovirus display technologies based on the expression of foreign sequences on the surface of Autographa californica (AcMNPV have been developed. We investigated the potential of a display methodology for a cDNA library of maize young seedlings. Results We constructed a full-length cDNA library of young maize etiolated seedlings in the transfer vector pAcTMVSVG. The library contained a total of 2.5 × 105 independent clones. Expression of two known maize proteins, calreticulin and auxin binding protein (ABP1, was shown by western blot analysis of protein extracts from insect cells infected with the cDNA library. Display of the two proteins in infected insect cells was shown by selective biopanning using magnetic cell sorting and demonstrated proof of concept that the baculovirus maize cDNA display library could be used to identify and isolate proteins. Conclusion The maize cDNA library constructed in this study relies on the novel technology of baculovirus display and is unique in currently published cDNA libraries. Produced to demonstrate proof of principle, it opens the way for the development of a eukaryotic in vivo display tool which would be ideally suited for rapid screening of the maize proteome for binding partners, such as proteins involved in hormone regulation or defence.

CDNA encoding a polypeptide including a hevein sequence

Science.gov (United States)

Raikhel, Natasha V.; Broekaert, Willem F.; Chua, Nam-Hai; Kush, Anil

1995-03-21

A cDNA clone (HEV1) encoding hevein was isolated via polymerase chain reaction (PCR) using mixed oligonucleotides corresponding to two regions of hevein as primers and a Hevea brasiliensis latex cDNA library as a template. HEV1 is 1018 nucleotides long and includes an open reading frame of 204 amino acids. The deduced amino acid sequence contains a putative signal sequence of 17 amino acid residues followed by a 187 amino acid polypeptide. The amino-terminal region (43 amino acids) is identical to hevein and shows homology to several chitin-binding proteins and to the amino-termini of wound-induced genes in potato and poplar. The carboxyl-terminal portion of the polypeptide (144 amino acids) is 74-79% homologous to the carboxyl-terminal region of wound-inducible genes of potato. Wounding, as well as application of the plant hormones abscisic acid and ethylene, resulted in accumulation of hevein transcripts in leaves, stems and latex, but not in roots, as shown by using the cDNA as a probe. A fusion protein was produced in E. coli from the protein of the present invention and maltose binding protein produced by the E. coli.

Method for construction of normalized cDNA libraries

Science.gov (United States)

Soares, Marcelo B.; Efstratiadis, Argiris

1998-01-01

This invention provides a method to normalize a directional cDNA library constructed in a vector that allows propagation in single-stranded circle form comprising: (a) propagating the directional cDNA library in single-stranded circles; (b) generating fragments complementary to the 3' noncoding sequence of the single-stranded circles in the library to produce partial duplexes; (c) purifying the partial duplexes; (d) melting and reassociating the purified partial duplexes to appropriate Cot; and (e) purifying the unassociated single-stranded circles, thereby generating a normalized cDNA library. This invention also provides normalized cDNA libraries generated by the above-described method and uses of the generated libraries.

Biased and unbiased perceptual decision-making on vocal emotions.

Science.gov (United States)

Dricu, Mihai; Ceravolo, Leonardo; Grandjean, Didier; Frühholz, Sascha

2017-11-24

Perceptual decision-making on emotions involves gathering sensory information about the affective state of another person and forming a decision on the likelihood of a particular state. These perceptual decisions can be of varying complexity as determined by different contexts. We used functional magnetic resonance imaging and a region of interest approach to investigate the brain activation and functional connectivity behind two forms of perceptual decision-making. More complex unbiased decisions on affective voices recruited an extended bilateral network consisting of the posterior inferior frontal cortex, the orbitofrontal cortex, the amygdala, and voice-sensitive areas in the auditory cortex. Less complex biased decisions on affective voices distinctly recruited the right mid inferior frontal cortex, pointing to a functional distinction in this region following decisional requirements. Furthermore, task-induced neural connectivity revealed stronger connections between these frontal, auditory, and limbic regions during unbiased relative to biased decision-making on affective voices. Together, the data shows that different types of perceptual decision-making on auditory emotions have distinct patterns of activations and functional coupling that follow the decisional strategies and cognitive mechanisms involved during these perceptual decisions.

Influence of gel/LED-laser application on cervical microleakage of two barrier materials used for endodontically treated teeth whitening

Science.gov (United States)

Marchesan, Melissa Andréia; Barros, Felipe; Porto, Saulo; Zaitter, Suellen; Brugnera, Aldo, Jr.; Sousa-Neto, Manoel D.

2007-02-01

This study evaluated ex vivo the influence of the number of gel/LED-laser applications/activations on cervical microleakage of two different barrier materials used for protection during whitening of endodontically treated teeth. Eighty-four canines were instrumented and obturated with epoxy resin sealer. The seal was removed 2 mm beyond the cemento-enamel junction for barrier placement and the teeth were divided into two groups of 40 teeth each: G1, zinc phosphate cement; G2, glass ionomer cement. The two groups were subdivided into 4 subgroups (n=10 each): I) no gel or LED-laser application; II) one gel application and two LED-laser activations; III) two gel applications and four LED-laser activations; IV) three gel applications and six LED-laser activations. The teeth were immersed in India ink for 7 days, decalcified and cleared. Cervical microleakage was quantified with a measurement microscope. Statistical analysis showed that zinc phosphate caused significantly lower microleakage than glass ionomer cement (presented microleakage in all subgroups). However, after two (p<0.01) and three (p<0.001) applications of gel, there was statistially significant microleakage in zinc phosphate barriers. Based on the present results, it can be concluded that cervical barriers with zinc phosphate cement show less cervical microleakage and that two or more applications/activations of gel/LED-laser significantly increase microleakage.

A Robust Identification of the Protein Standard Bands in Two-Dimensional Electrophoresis Gel Images

Directory of Open Access Journals (Sweden)

Serackis Artūras

2017-12-01

Full Text Available The aim of the investigation presented in this paper was to develop a software-based assistant for the protein analysis workflow. The prior characterization of the unknown protein in two-dimensional electrophoresis gel images is performed according to the molecular weight and isoelectric point of each protein spot estimated from the gel image before further sequence analysis by mass spectrometry. The paper presents a method for automatic and robust identification of the protein standard band in a two-dimensional gel image. In addition, the method introduces the identification of the positions of the markers, prepared by using pre-selected proteins with known molecular mass. The robustness of the method was achieved by using special validation rules in the proposed original algorithms. In addition, a self-organizing map-based decision support algorithm is proposed, which takes Gabor coefficients as image features and searches for the differences in preselected vertical image bars. The experimental investigation proved the good performance of the new algorithms included into the proposed method. The detection of the protein standard markers works without modification of algorithm parameters on two-dimensional gel images obtained by using different staining and destaining procedures, which results in different average levels of intensity in the images.

Styl RFLP recognized by a human IRBP cDNA localized to chromosome 10

Energy Technology Data Exchange (ETDEWEB)

Chin, K S; Mathew, C G.P.; Fong, S L; Bridges, C D; Ponder, B A.J.

1988-02-25

A 2184 bp cDNA (H.4 IRBP) encoding human interstitial retinol-biding protein isolated from a human retina cDNA library in lambdagt10 by screening with a bovine IRBP cDNA probe. Styl identifies a 2-allele polymorphism with bands at 2.3 kb (Cl) and 1.95 kb (C2) and invariant bands at 1.1, 1.0 and 0.8kb. Codominant segregation was observed in two informative families. The RFLP was mapped to chromosome 10 using somatic cell hybrids. In situ hybridization suggests regional assignments near p11.2 -q11.2 with a secondary site of hybridization at q24-25.

cDNA - ASTRA | LSDB Archive [Life Science Database Archive metadata

Lifescience Database Archive (English)

Full Text Available ontents List of cDNA in locus Data file File name: astra_cdna.zip File URL: ftp://ftp.biosciencedbc.jp/archive/astra/LATEST/astra_cdn...a.zip File size: 3.3 MB Simple search URL http://togodb.biosciencedbc.jp/togodb/view/astra_cdna...n, Department of Molecular Genetics, National Institute of Agrobiological Sciences (Kikuchi et al., 2003; ftp://cdna

Isolation of full-length putative rat lysophospholipase cDNA using improved methods for mRNA isolation and cDNA cloning

International Nuclear Information System (INIS)

Han, J.H.; Stratowa, C.; Rutter, W.J.

1987-01-01

The authors have cloned a full-length putative rat pancreatic lysophospholipase cDNA by an improved mRNA isolation method and cDNA cloning strategy using [ 32 P]-labelled nucleotides. These new methods allow the construction of a cDNA library from the adult rat pancreas in which the majority of recombinant clones contained complete sequences for the corresponding mRNAs. A previously recognized but unidentified long and relatively rare cDNA clone containing the entire sequence from the cap site at the 5' end to the poly(A) tail at the 3' end of the mRNA was isolated by single-step screening of the library. The size, amino acid composition, and the activity of the protein expressed in heterologous cells strongly suggest this mRNA codes for lysophospholipase

Rapid in vitro labeling procedures for two-dimensional gel fingerprinting

International Nuclear Information System (INIS)

Lee, Y.F.; Fowlks, E.R.

1982-01-01

Improvements of existing in vitro procedures for labeling RNA radioactively, and modifications of the two-dimensional polyacrylamide gel electrophoresis system for making RNA fingerprints are described. These improvements are (a) inactivation of phosphatase with nitric acid at pH 2.0 eliminated the phenol-cholorform extraction step during 5'-end labeling with polynucleotide kinase and [γ- 32 P]ATP; (b) ZnSO 4 inactivation of R Nase T 1 results in a highly efficient procedure for 3'-end labeling with T4 ligase and [5'- 32 P]pCp; and (c) a rapid 4-min procedure for variable quantity range of 125 I and RNA results in a qualitative and quantitative sample for high-molecular weight RNA fingerprinting. Thus, these in vitro procedures become rapid and reproducible when combined with two-dimensional gel electrophoresis which eliminates simultaneously labeled impurities. Each labeling procedure is compared, using tobacco mosaic virus, Brome mosaic virus, and polio RNA. A series of Ap-rich oligonucleotides was discovered in the inner genome of Brome mosaic Virus RNA-3

Some Structural Observations of Self-Assembling, Fibrillar Gels Composed of Two-Directional Bolaform Arborols

Energy Technology Data Exchange (ETDEWEB)

Sun, J.

2005-01-12

Arborols are dumbbell shaped molecules (bolaform amphiphiles) in which a hydrophobic spacer separates two hydrophilic end groups. They are a valuable model for naturally occurring fibers, such as actin or amyloid. Applications to materials science can be envisioned. On cooling from warm aqueous or methanolic solutions, arborols spontaneously assemble into long fibers. When the solutions are above a certain concentration that depends on the hydrophilic/hydrophobic balance, this leads to thermally reversible gels stabilized by a mechanism that is poorly understood. With the help of wide angle X-ray scattering, details of the arborol fiber and gel structure were obtained on wet gels. The characteristic dimensions of the fibers vary in a sensible fashion with the molecular specifics. Solvent character appears to affect the average domain length of arborols stacked into fibers. Fluorescently labeled arborols were prepared. The label does not prevent incorporation into the fibrillar structure, rendering fibril bundles visible in wet gels. Bundles are visible in concentrated gels, but not in less concentrated sols. These results are consistent with observations of dried arborols using atomic force microscopy and with previously published freeze-fracture electron microscopy and small angle X-ray scattering experiments on dried gels.

[Preparation of the cDNA microarray on the differential expressed cDNA of senescence-accelerated mouse's hippocampus].

Science.gov (United States)

Cheng, Xiao-Rui; Zhou, Wen-Xia; Zhang, Yong-Xiang

2006-05-01

Alzheimer' s disease (AD) is the most common form of dementia in the elderly. AD is an invariably fatal neurodegenerative disorder with no effective treatment. Senescence-accelerated mouse prone 8 (SAMP8) is a model for studying age-related cognitive impairments and also is a good model to study brain aging and one of mouse model of AD. The technique of cDNA microarray can monitor the expression levels of thousands of genes simultaneously and can be used to study AD with the character of multi-mechanism, multi-targets and multi-pathway. In order to disclose the mechanism of AD and find the drug targets of AD, cDNA microarray containing 3136 cDNAs amplified from the suppression subtracted cDNA library of hippocampus of SAMP8 and SAMR1 was prepared with 16 blocks and 14 x 14 pins, the housekeeping gene beta-actin and G3PDH as inner conference. The background of this microarray was low and unanimous, and dots divided evenly. The conditions of hybridization and washing were optimized during the hybridization of probe and target molecule. After the data of hybridization analysis, the differential expressed cDNAs were sequenced and analyzed by the bioinformatics, and some of genes were quantified by the real time RT-PCR and the reliability of this cDNA microarray were validated. This cDNA microarray may be the good means to select the differential expressed genes and disclose the molecular mechanism of SAMP8's brain aging and AD.

Electromyography analysis of natural mastication behavior using varying mouthful quantities of two types of gels.

Science.gov (United States)

Kohyama, Kaoru; Gao, Zhihong; Ishihara, Sayaka; Funami, Takahiro; Nishinari, Katsuyoshi

2016-07-01

The objectives of this study were to examine the effects of mouthful quantities and mechanical properties of gels on natural mastication behaviors using electromyography (EMG). Two types of hydrocolloid gels (A and K) with similar fracture loads but different moduli and fracture strains were served to eleven normal women in 3-, 6-, 12-, and 24-g masses in a randomized order. EMG activities from both masseter muscles were recorded during natural mastication. Because of the similar fracture loads, the numbers of chews, total muscle activities, and entire oral processing times were similar for similar masses of both gel types. Prior to the first swallow, the more elastic K gel with a higher fracture strain required higher muscle activities than the brittle A gel, which had higher modulus. Majority of subjects had preferred sides of chewing, but all subjects with or without preferred sides used both masseters during the consumption of gels. Similar effects of masses and types of gels were observed in EMG activities of both sides of masseters. Contributions of the dominant side of chewing were diminished with increasing masses of gels, and the mass dependency on ratio of the dominant side was more pronounced with K gel. More repetitions of smaller masses required greater muscle activities and longer periods for the consumption of 24-g gel portions. Reduction in the masses with an increased number of repetitions necessitated slower eating and more mastication to consume the gel portions. These observations suggest that chewing using both sides is more effective and unconsciously reduces mastication times during the consumption of gels. Copyright © 2016 Elsevier Inc. All rights reserved.

Unbiased water and methanol maser surveys of NGC 1333

Energy Technology Data Exchange (ETDEWEB)

Lyo, A-Ran; Kim, Jongsoo; Byun, Do-Young; Lee, Ho-Gyu, E-mail: arl@kasi.re.kr [Korea Astronomy and Space Science Institute, 776, Daedeokdae-ro Yuseong-gu, Daejeon 305-348 (Korea, Republic of)

2014-11-01

We present the results of unbiased 22 GHz H{sub 2}O water and 44 GHz class I CH{sub 3}OH methanol maser surveys in the central 7' × 10' area of NGC 1333 and two additional mapping observations of a 22 GHz water maser in a ∼3' × 3' area of the IRAS4A region. In the 22 GHz water maser survey of NGC 1333 with a sensitivity of σ ∼ 0.3 Jy, we confirmed the detection of masers toward H{sub 2}O(B) in the region of HH 7-11 and IRAS4B. We also detected new water masers located ∼20'' away in the western direction of IRAS4B or ∼25'' away in the southern direction of IRAS4A. We could not, however, find young stellar objects or molecular outflows associated with them. They showed two different velocity components of ∼0 and ∼16 km s{sup –1}, which are blue- and redshifted relative to the adopted systemic velocity of ∼7 km s{sup –1} for NGC 1333. They also showed time variabilities in both intensity and velocity from multi-epoch observations and an anti-correlation between the intensities of the blue- and redshifted velocity components. We suggest that the unidentified power source of these masers might be found in the earliest evolutionary stage of star formation, before the onset of molecular outflows. Finding this kind of water maser is only possible through an unbiased blind survey. In the 44 GHz methanol maser survey with a sensitivity of σ ∼ 0.5 Jy, we confirmed masers toward IRAS4A2 and the eastern shock region of IRAS2A. Both sources are also detected in 95 and 132 GHz methanol maser lines. In addition, we had new detections of methanol masers at 95 and 132 GHz toward IRAS4B. In terms of the isotropic luminosity, we detected methanol maser sources brighter than ∼5 × 10{sup 25} erg s{sup –1} from our unbiased survey.

Procedure for normalization of cDNA libraries

Science.gov (United States)

Bonaldo, Maria DeFatima; Soares, Marcelo Bento

1997-01-01

This invention provides a method to normalize a cDNA library constructed in a vector capable of being converted to single-stranded circles and capable of producing complementary nucleic acid molecules to the single-stranded circles comprising: (a) converting the cDNA library in single-stranded circles; (b) generating complementary nucleic acid molecules to the single-stranded circles; (c) hybridizing the single-stranded circles converted in step (a) with complementary nucleic acid molecules of step (b) to produce partial duplexes to an appropriate Cot; (e) separating the unhybridized single-stranded circles from the hybridized single-stranded circles, thereby generating a normalized cDNA library.

Automated and unbiased classification of chemical profiles from fungi using high performance liquid chromatography

DEFF Research Database (Denmark)

Hansen, Michael Edberg; Andersen, Birgitte; Smedsgaard, Jørn

2005-01-01

In this paper we present a method for unbiased/unsupervised classification and identification of closely related fungi, using chemical analysis of secondary metabolite profiles created by HPLC with UV diode array detection. For two chromatographic data matrices a vector of locally aligned full sp...

Mutually Unbiased Maximally Entangled Bases for the Bipartite System Cd⊗ C^{dk}

Science.gov (United States)

Nan, Hua; Tao, Yuan-Hong; Wang, Tian-Jiao; Zhang, Jun

2016-10-01

The construction of maximally entangled bases for the bipartite system Cd⊗ Cd is discussed firstly, and some mutually unbiased bases with maximally entangled bases are given, where 2≤ d≤5. Moreover, we study a systematic way of constructing mutually unbiased maximally entangled bases for the bipartite system Cd⊗ C^{dk}.

Gel versus capillary electrophoresis genotyping for categorizing treatment outcomes in two anti-malarial trials in Uganda

Directory of Open Access Journals (Sweden)

Hubbard Alan E

2010-01-01

Full Text Available Abstract Background Molecular genotyping is performed in anti-malarial trials to determine whether recurrent parasitaemia after therapy represents a recrudescence (treatment failure or new infection. The use of capillary instead of agarose gel electrophoresis for genotyping offers technical advantages, but it is unclear whether capillary electrophoresis will result in improved classification of anti-malarial treatment outcomes. Methods Samples were genotyped using both gel and capillary electrophoresis from randomized trials of artemether-lumefantrine (AL vs. dihydroartemisinin-piperaquine (DP performed in two areas of Uganda: Kanungu, where transmission is moderate, and Apac, where transmission is very high. Both gel and capillary methods evaluated polymorphic regions of the merozoite surface protein 1 and 2 and glutamine rich protein genes. Results Capillary electrophoresis detected more alleles and provided higher discriminatory power than agarose gel electrophoresis at both study sites. There was only moderate agreement between classification of outcomes with the two methods in Kanungu (kappa = 0.66 and poor agreement in Apac (kappa = 0.24. Overall efficacy results were similar when using gel vs. capillary methods in Kanungu (42-day risk of treatment failure for AL: 6.9% vs. 5.5%, p = 0.4; DP 2.4% vs. 2.9%, p = 0.5. However, the measured risk of recrudescence was significantly higher when using gel vs. capillary electrophoresis in Apac (risk of treatment failure for AL: 17.0% vs. 10.7%, p = 0.02; DP: 8.5% vs. 3.4%, p = 0.03. Risk differences between AL and DP were not significantly different whether gel or capillary methods were used. Conclusions Genotyping with gel electrophoresis overestimates the risk of recrudescence in anti-malarial trials performed in areas of high transmission intensity. Capillary electrophoresis provides more accurate outcomes for such trials and should be performed when possible. In areas of moderate transmission

Gel versus capillary electrophoresis genotyping for categorizing treatment outcomes in two anti-malarial trials in Uganda.

Science.gov (United States)

Gupta, Vinay; Dorsey, Grant; Hubbard, Alan E; Rosenthal, Philip J; Greenhouse, Bryan

2010-01-15

Molecular genotyping is performed in anti-malarial trials to determine whether recurrent parasitaemia after therapy represents a recrudescence (treatment failure) or new infection. The use of capillary instead of agarose gel electrophoresis for genotyping offers technical advantages, but it is unclear whether capillary electrophoresis will result in improved classification of anti-malarial treatment outcomes. Samples were genotyped using both gel and capillary electrophoresis from randomized trials of artemether-lumefantrine (AL) vs. dihydroartemisinin-piperaquine (DP) performed in two areas of Uganda: Kanungu, where transmission is moderate, and Apac, where transmission is very high. Both gel and capillary methods evaluated polymorphic regions of the merozoite surface protein 1 and 2 and glutamine rich protein genes. Capillary electrophoresis detected more alleles and provided higher discriminatory power than agarose gel electrophoresis at both study sites. There was only moderate agreement between classification of outcomes with the two methods in Kanungu (kappa = 0.66) and poor agreement in Apac (kappa = 0.24). Overall efficacy results were similar when using gel vs. capillary methods in Kanungu (42-day risk of treatment failure for AL: 6.9% vs. 5.5%, p = 0.4; DP 2.4% vs. 2.9%, p = 0.5). However, the measured risk of recrudescence was significantly higher when using gel vs. capillary electrophoresis in Apac (risk of treatment failure for AL: 17.0% vs. 10.7%, p = 0.02; DP: 8.5% vs. 3.4%, p = 0.03). Risk differences between AL and DP were not significantly different whether gel or capillary methods were used. Genotyping with gel electrophoresis overestimates the risk of recrudescence in anti-malarial trials performed in areas of high transmission intensity. Capillary electrophoresis provides more accurate outcomes for such trials and should be performed when possible. In areas of moderate transmission, gel electrophoresis appears adequate to estimate comparative

Higher-dimensional orbital-angular-momentum-based quantum key distribution with mutually unbiased bases

CSIR Research Space (South Africa)

Mafu, M

2013-09-01

Full Text Available We present an experimental study of higher-dimensional quantum key distribution protocols based on mutually unbiased bases, implemented by means of photons carrying orbital angular momentum. We perform (d + 1) mutually unbiased measurements in a...

cDNA encoding a polypeptide including a hevein sequence

Energy Technology Data Exchange (ETDEWEB)

Raikhel, N.V.; Broekaert, W.F.; Chua, N.H.; Kush, A.

2000-07-04

A cDNA clone (HEV1) encoding hevein was isolated via polymerase chain reaction (PCR) using mixed oligonucleotides corresponding to two regions of hevein as primers and a Hevea brasiliensis latex cDNA library as a template. HEV1 is 1018 nucleotides long and includes an open reading frame of 204 amino acids. The deduced amino acid sequence contains a putative signal sequence of 17 amino acid residues followed by a 187 amino acid polypeptide. The amino-terminal region (43 amino acids) is identical to hevein and shows homology to several chitin-binding proteins and to the amino-termini of wound-induced genes in potato and poplar. The carboxyl-terminal portion of the polypeptide (144 amino acids) is 74--79% homologous to the carboxyl-terminal region of wound-inducible genes of potato. Wounding, as well as application of the plant hormones abscisic acid and ethylene, resulted in accumulation of hevein transcripts in leaves, stems and latex, but not in roots, as shown by using the cDNA as a probe. A fusion protein was produced in E. coli from the protein of the present invention and maltose binding protein produced by the E. coli.

cDNA encoding a polypeptide including a hevein sequence

Energy Technology Data Exchange (ETDEWEB)

Raikhel, N.V.; Broekaert, W.F.; Chua, N.H.; Kush, A.

1999-05-04

A cDNA clone (HEV1) encoding hevein was isolated via polymerase chain reaction (PCR) using mixed oligonucleotides corresponding to two regions of hevein as primers and a Hevea brasiliensis latex cDNA library as a template. HEV1 is 1018 nucleotides long and includes an open reading frame of 204 amino acids. The deduced amino acid sequence contains a putative signal sequence of 17 amino acid residues followed by a 187 amino acid polypeptide. The amino-terminal region (43 amino acids) is identical to hevein and shows homology to several chitin-binding proteins and to the amino-termini of wound-induced genes in potato and poplar. The carboxyl-terminal portion of the polypeptide (144 amino acids) is 74--79% homologous to the carboxyl-terminal region of wound-inducible genes of potato. Wounding, as well as application of the plant hormones abscisic acid and ethylene, resulted in accumulation of hevein transcripts in leaves, stems and latex, but not in roots, as shown by using the cDNA as a probe. A fusion protein was produced in E. coli from the protein of the present invention and maltose binding protein produced by the E. coli. 12 figs.

cDNA encoding a polypeptide including a hevein sequence

Energy Technology Data Exchange (ETDEWEB)

Raikhel, Natasha V. (Okemos, MI); Broekaert, Willem F. (Dilbeek, BE); Chua, Nam-Hai (Scarsdale, NY); Kush, Anil (New York, NY)

1999-05-04

A cDNA clone (HEV1) encoding hevein was isolated via polymerase chain reaction (PCR) using mixed oligonucleotides corresponding to two regions of hevein as primers and a Hevea brasiliensis latex cDNA library as a template. HEV1 is 1018 nucleotides long and includes an open reading frame of 204 amino acids. The deduced amino acid sequence contains a putative signal sequence of 17 amino acid residues followed by a 187 amino acid polypeptide. The amino-terminal region (43 amino acids) is identical to hevein and shows homology to several chitin-binding proteins and to the amino-termini of wound-induced genes in potato and poplar. The carboxyl-terminal portion of the polypeptide (144 amino acids) is 74-79% homologous to the carboxyl-terminal region of wound-inducible genes of potato. Wounding, as well as application of the plant hormones abscisic acid and ethylene, resulted in accumulation of hevein transcripts in leaves, stems and latex, but not in roots, as shown by using the cDNA as a probe. A fusion protein was produced in E. coli from the protein of the present invention and maltose binding protein produced by the E. coli.

cDNA encoding a polypeptide including a hevein sequence

Energy Technology Data Exchange (ETDEWEB)

Raikhel, N.V.; Broekaert, W.F.; Chua, N.H.; Kush, A.

1995-03-21

A cDNA clone (HEV1) encoding hevein was isolated via polymerase chain reaction (PCR) using mixed oligonucleotides corresponding to two regions of hevein as primers and a Hevea brasiliensis latex cDNA library as a template. HEV1 is 1,018 nucleotides long and includes an open reading frame of 204 amino acids. The deduced amino acid sequence contains a putative signal sequence of 17 amino acid residues followed by a 187 amino acid polypeptide. The amino-terminal region (43 amino acids) is identical to hevein and shows homology to several chitin-binding proteins and to the amino-termini of wound-induced genes in potato and poplar. The carboxyl-terminal portion of the polypeptide (144 amino acids) is 74--79% homologous to the carboxyl-terminal region of wound-inducible genes of potato. Wounding, as well as application of the plant hormones abscisic acid and ethylene, resulted in accumulation of hevein transcripts in leaves, stems and latex, but not in roots, as shown by using the cDNA as a probe. A fusion protein was produced in E. coli from the protein of the present invention and maltose binding protein produced by the E. coli. 11 figures.

Lectin cDNA and transgenic plants derived therefrom

Science.gov (United States)

Raikhel, Natasha V.

2000-10-03

Transgenic plants containing cDNA encoding Gramineae lectin are described. The plants preferably contain cDNA coding for barley lectin and store the lectin in the leaves. The transgenic plants, particularly the leaves exhibit insecticidal and fungicidal properties.

cDNA table - RPD | LSDB Archive [Life Science Database Archive metadata

Lifescience Database Archive (English)

Full Text Available of data contents Results of homology search to cDNA clones in the KOME. Data file File name: rpd_cdna.zip F...ile URL: ftp://ftp.biosciencedbc.jp/archive/rpd/LATEST/rpd_cdna.zip File size: 15 KB Simple search URL http:...//togodb.biosciencedbc.jp/togodb/view/rpd_cdna#en Data acquisition method - Data

Cloning and sequence analysis of cDNA coding for rat nucleolar protein C23

International Nuclear Information System (INIS)

Ghaffari, S.H.; Olson, M.O.J.

1986-01-01

Using synthetic oligonucleotides as primers and probes, the authors have isolated and sequenced cDNA clones encoding protein C23, a putative nucleolus organizer protein. Poly(A + ) RNA was isolated from rat Novikoff hepatoma cells and enriched in C23 mRNA by sucrose density gradient ultracentrifugation. Two deoxyoligonuleotides, a 48- and a 27-mer, were synthesized on the basis of amino acid sequence from the C-terminal half of protein C23 and cDNA sequence data from CHO cell protein. The 48-mer was used a primer for synthesis of cDNA which was then inserted into plasmid pUC9. Transformed bacterial colonies were screened by hybridization with 32 P labeled 27-mer. Two clones among 5000 gave a strong positive signal. Plasmid DNAs from these clones were purified and characterized by blotting and nucleotide sequence analysis. The length of C23 mRNA was estimated to be 3200 bases in a northern blot analysis. The sequence of a 267 b.p. insert shows high homology with the CHO cDNA with only 9 nucleotide differences and an identical amino acid sequence. These studies indicate that this region of the protein is highly conserved

A new unbiased stochastic derivative estimator for discontinuous sample performances with structural parameters

NARCIS (Netherlands)

Peng, Yijie; Fu, Michael C.; Hu, Jian Qiang; Heidergott, Bernd

In this paper, we propose a new unbiased stochastic derivative estimator in a framework that can handle discontinuous sample performances with structural parameters. This work extends the three most popular unbiased stochastic derivative estimators: (1) infinitesimal perturbation analysis (IPA), (2)

[Construction of fetal mesenchymal stem cell cDNA subtractive library].

Science.gov (United States)

Yang, Li; Wang, Dong-Mei; Li, Liang; Bai, Ci-Xian; Cao, Hua; Li, Ting-Yu; Pei, Xue-Tao

2002-04-01

To identify differentially expressed genes between fetal mesenchymal stem cell (MSC) and adult MSC, especially specified genes expressed in fetal MSC, a cDNA subtractive library of fetal MSC was constructed using suppression subtractive hybridization (SSH) technique. At first, total RNA was isolated from fetal and adult MSC. Using SMART PCR synthesis method, single-strand and double-strand cDNAs were synthesized. After Rsa I digestion, fetal MSC cDNAs were divided into two groups and ligated to adaptor 1 and adaptor 2 respectively. Results showed that the amplified library contains 890 clones. Analysis of 890 clones with PCR demonstrated that 768 clones were positive. The positive rate is 86.3%. The size of inserted fragments in these positive clones was between 0.2 - 1 kb, with an average of 400 - 600 bp. SSH is a convenient and effective method for screening differentially expressed genes. The constructed cDNA subtractive library of fetal MSC cDNA lays solid foundation for screening and cloning new and specific function related genes of fetal MSC.

Molecular cloning of growth hormone encoding cDNA of Indian

Indian Academy of Sciences (India)

A modified rapid amplification of cDNA ends (RACE) strategy has been developed for cloning highly conserved cDNA sequences. Using this modified method, the growth hormone (GH) encoding cDNA sequences of Labeo rohita, Cirrhina mrigala and Catla catla have been cloned, characterized and overexpressed in ...

Two-dimensional gel electrophoresis of selenized yeast and autoradiography of 75Se-containing proteins

International Nuclear Information System (INIS)

Chery, C.C.; Dumont, E.; Cornelis, R.; Moens, L.

2001-01-01

Two-dimensional high-resolution gel electrophoresis (2DE) has been applied to the fractionation of 75 Se-containing proteins in yeast, grown in 75 Se-containing medium, and autoradiography was used for detection of the 75 Se-containing proteins. Gel filtration and ultrafiltration were used to check whether the selenium side-chains were stable in the presence of the chemicals used for lysis and 2DE. The mass distribution of the selenium-containing proteins was estimated by use of gel filtration and the results were compared with the distribution obtained by 2DE. A 2DE map of selenium-containing proteins in yeast is presented, and compared with a total protein map of yeast. (orig.)

Analysis of cDNA libraries from developing seeds of guar (Cyamopsis tetragonoloba (L. Taub

Directory of Open Access Journals (Sweden)

Dixon Richard A

2007-11-01

Full Text Available Abstract Background Guar, Cyamopsis tetragonoloba (L. Taub, is a member of the Leguminosae (Fabaceae family and is economically the most important of the four species in the genus. The endosperm of guar seed is a rich source of mucilage or gum, which forms a viscous gel in cold water, and is used as an emulsifier, thickener and stabilizer in a wide range of foods and industrial applications. Guar gum is a galactomannan, consisting of a linear (1→4-β-linked D-mannan backbone with single-unit, (1→6-linked, α-D-galactopyranosyl side chains. To better understand regulation of guar seed development and galactomannan metabolism we created cDNA libraries and a resulting EST dataset from different developmental stages of guar seeds. Results A database of 16,476 guar seed ESTs was constructed, with 8,163 and 8,313 ESTs derived from cDNA libraries I and II, respectively. Library I was constructed from seeds at an early developmental stage (15–25 days after flowering, DAF, and library II from seeds at 30–40 DAF. Quite different sets of genes were represented in these two libraries. Approximately 27% of the clones were not similar to known sequences, suggesting that these ESTs represent novel genes or may represent non-coding RNA. The high flux of energy into carbohydrate and storage protein synthesis in guar seeds was reflected by a high representation of genes annotated as involved in signal transduction, carbohydrate metabolism, chaperone and proteolytic processes, and translation and ribosome structure. Guar unigenes involved in galactomannan metabolism were identified. Among the seed storage proteins, the most abundant contig represented a conglutin accounting for 3.7% of the total ESTs from both libraries. Conclusion The present EST collection and its annotation provide a resource for understanding guar seed biology and galactomannan metabolism.

Comparison and Application of Two types of Filling Gel to Prevent Spontaneous Combustion at the Region where Top-Coal Caves above Entry

Directory of Open Access Journals (Sweden)

Wang Yuhuai

2016-01-01

Full Text Available Two types of gel were developed, by taking fly ash and foaming cement as aggregate, which is usually used as filling material at the region where top-coal caves above coal entry in the Jinggezhuang coal mine, and adding high molecular polymer and bio-gel as additive. Sweating rates of the two types of gel under various matching ratio and temperature were tested. And then sweating ratio and water retention ratio of the two gels were calculated, based on which, the optimized matching ratios, were determined. Viscosity indexes of the two-type gel under different ratios were tested. The optimized filling ratios of the two types of gel were determined according to the two indexes, water retention rate and the viscosity. The filling experiments were implemented and evaluated in site, the Jinggezhuang coal mine. The results show that the fly ash gel has a good achievement on preventing spontaneous combustion at the Region where Top-Coal Caves above entries. It is promising, economically and environmental friendly, and valuable in popularization in coal mines.

Multivariate data analysis of two-dimensional gel electrophoresis protein patterns from few samples

DEFF Research Database (Denmark)

Jensen, Kristina Nedenskov; Jessen, Flemming; Jørgensen, Bo

2008-01-01

One application of 2D gel electrophoresis is to reveal differences in protein pattern between two or more groups of individuals, attributable to their group membership. Multivariate data analytical methods are useful in pinpointing the spots relevant for discrimination by focusing not only...... on single spot differences, but on the covariance structure between proteins. However, their outcome is dependent on data scaling, and they may fail in producing valid multivariate models due to the much higher number of "irrelevant" spots present in the gels. The case where only few gels are available...... and where the aim is to find as many as possible of the group-dependent proteins seems particularly difficult to handle. The present paper investigates such a case regarding the effect of scaling and of prefiltering by univariate nonparametric statistics on the selection of spots. Besides, a modified...

Cloning and analysis of the mouse Fanconi anemia group a cDNA and an overlapping penta zinc finger cDNA

NARCIS (Netherlands)

Wong, JCY; Alon, N; Norga, K; Kruyt, FAE; Youssoufian, H; Buchwald, M

2000-01-01

Despite the cloning of four disease-associated genes for Fanconi anemia (FA), the molecular pathogenesis of FA remains largely unknown. To study FA complementation group A using the mouse as a mode I system, we cloned and characterized the mouse homolog of the human FANCA cDNA, The mouse cDNA

Purification, reactivity with IgE and cDNA cloning of parvalbumin as the major allergen of mackerels.

Science.gov (United States)

Hamada, Y; Tanaka, H; Ishizaki, S; Ishida, M; Nagashima, Y; Shiomi, K

2003-08-01

Three species of mackerels (Scomber japonicus, S. australasicus and S. scombrus) are widely consumed and considered to be most frequently involved in incidents of IgE-mediated fish allergy in Japan. In this study, parvalbumin, a possible candidate for the major allergen, was purified from the white muscle of three species of mackerels by gel filtration on Sephadex G-75 and reverse-phase HPLC on TSKgel ODS-120T. All the purified preparations from three species gave a single band of about 11 kDa and were clearly identified as parvalbumins by analyses of their partial amino acid sequences. In ELISA experiments, four of five sera from fish-allergic patients reacted to all the purified parvalbumins, demonstrating that parvalbumin is the major allergen in common with the mackerels. Antigenic cross-reactivity among the mackerel parvalbumins was also established by ELISA inhibition experiments. A cDNA library was constructed from the white muscle of S. japonicus and the cDNA encoding parvalbumin was cloned. The amino acid sequence translated from the nucleotide sequence revealed that the S. japonicus parvalbumin is composed of 108 residues, being a member of beta-type parvalbumins.

Electrophoresis of DNA in agarose gels, polyacrylamide gels and in free solution

Science.gov (United States)

Stellwagen, Nancy C.

2009-01-01

This review describes the electrophoresis of curved and normal DNA molecules in agarose gels, polyacrylamide gels and in free solution. These studies were undertaken to clarify why curved DNA molecules migrate anomalously slowly in polyacrylamide gels but not in agarose gels. Two milestone papers are cited, in which Ferguson plots were used to estimate the effective pore size of agarose and polyacrylamide gels. Subsequent studies on the effect of the electric field on agarose and polyacrylamide gel matrices, DNA interactions with the two gel matrices, and the effect of curvature on the free solution mobility of DNA are also described. The combined results suggest that the anomalously slow mobilities observed for curved DNA molecules in polyacrylamide gels are due primarily to preferential interactions of curved DNAs with the polyacrylamide gel matrix; the restrictive pore size of the matrix is of lesser importance. In free solution, DNA mobilities increase with increasing molecular mass until leveling off at a plateau value of (3.17 ± 0.01) × 10-4 cm2/Vs in 40 mM Tris-acetate-EDTA buffer at 20°C. Curved DNA molecules migrate anomalously slowly in free solution as well as in polyacrylamide gels, explaining why the Ferguson plots of curved and normal DNAs containing the same number of base pairs extrapolate to different mobilities at zero gel concentration. PMID:19517510

cDNA library information - Dicty_cDB | LSDB Archive [Life Science Database Archive metadata

Lifescience Database Archive (English)

Full Text Available List Contact us Dicty_cDB cDNA library information Data detail Data name cDNA library information DOI 10.189...s Data item Description cDNA library name Names of cDNA libraries (AF, AH, CF, CH, FC, FC-IC, FCL, SF, SH, S...(C) 5) sexually fusion-competent KAX3 cells (Gamete phase) (F) cDNA library construction method How to construct cDNA library...dir) 2) Full-length cDNA libraries (oligocapped method)(fl) 3) Gamete-specific subtraction library (sub) cDNA library... construction protocol Link to the webpage describing the protocol for generating cDNA library Size

An alternative method for cDNA cloning from surrogate eukaryotic cells transfected with the corresponding genomic DNA.

Science.gov (United States)

Hu, Lin-Yong; Cui, Chen-Chen; Song, Yu-Jie; Wang, Xiang-Guo; Jin, Ya-Ping; Wang, Ai-Hua; Zhang, Yong

2012-07-01

cDNA is widely used in gene function elucidation and/or transgenics research but often suitable tissues or cells from which to isolate mRNA for reverse transcription are unavailable. Here, an alternative method for cDNA cloning is described and tested by cloning the cDNA of human LALBA (human alpha-lactalbumin) from genomic DNA. First, genomic DNA containing all of the coding exons was cloned from human peripheral blood and inserted into a eukaryotic expression vector. Next, by delivering the plasmids into either 293T or fibroblast cells, surrogate cells were constructed. Finally, the total RNA was extracted from the surrogate cells and cDNA was obtained by RT-PCR. The human LALBA cDNA that was obtained was compared with the corresponding mRNA published in GenBank. The comparison showed that the two sequences were identical. The novel method for cDNA cloning from surrogate eukaryotic cells described here uses well-established techniques that are feasible and simple to use. We anticipate that this alternative method will have widespread applications.

New Approach for Segmentation and Quantification of Two-Dimensional Gel Electrophoresis Images

DEFF Research Database (Denmark)

Anjo, Antonio dos; Laurell Blom Møller, Anders; Ersbøll, Bjarne Kjær

2011-01-01

Motivation: Detection of protein spots in two-dimensional gel electrophoresis images (2-DE) is a very complex task and current approaches addressing this problem still suffer from significant shortcomings. When quantifying a spot, most of the current software applications include a lot of backgro...

cDNA library Table - KAIKOcDNA | LSDB Archive [Life Science Database Archive metadata

Lifescience Database Archive (English)

Full Text Available c00951-005 Description of data contents List of Bombyx mori cDNA libraries. Data file File name: kaiko_cdna_...library.zip File URL: ftp://ftp.biosciencedbc.jp/archive/kaiko-cdna/LATEST/kaiko_cdna_library.zip File size:... 4.8 KB Simple search URL http://togodb.biosciencedbc.jp/togodb/view/kaiko_cdna_l

Neutron spin-echo studies on dynamic and static fluctuations in two types of poly(vinyl alcohol) gels

International Nuclear Information System (INIS)

Kanaya, T.; Takahashi, N.; Nishida, K.; Seto, H.; Nagao, M.; Takeda, T.

2005-01-01

We report neutron spin-echo measurements on two types of poly(vinyl alcohol) (PVA) gels. The first is PVA gel in a mixture of dimethyl sulfoxide (DMSO) and water with volume ratio 60/40, and the second is PVA gel in an aqueous borax solution. The observed normalized intermediate scattering functions I(Q,t)/I(Q,0) are very different between them. The former I(Q,t)/I(Q,0) shows a nondecaying component in addition to a fast decay, but the latter does not have the nondecaying one. This clearly indicates that the fluctuations in the former PVA gel consist of static and dynamic fluctuations whereas the latter PVA gel does include only the dynamic fluctuations. The dynamic fluctuations of the former and latter gels have been analyzed in terms of a restricted motion in the network and Zimm motion, respectively, and the origins of these motions will be discussed

Transcutaneous Drainage of Gel-Like Substance after Application of Hydrogel Dural Sealant: Report of Two Cases.

Science.gov (United States)

Siman, Homayoun; Techy, Fernando

2016-02-01

Study Design Case report. Objective Incidental durotomy (IDT) is a common complication of spinal surgery. The use of collagen matrix graft along with hydrogel dural sealant is a common method of IDT repair. With this method, there have been several reported cases of detrimental dural sealant expansion in the literature. One case study reported an expansion rate greater than 300%; many report neurologic damage. This article reports the clinical course of two patients who developed postoperative transcutaneous drainage of a gel-like substance after the use of a dural sealant, which is a previously unreported complication. Methods The clinical course and treatment outcome of two patients is presented. Results Both patients experienced postoperative transcutaneous drainage of a gel-like substance at the surgical site. Case one began draining this substance on postoperative day 14. This patient required no further intervention, and the drainage ended after 3 mL of a gel-like substance was expressed from his incision while in the clinic. Case two began draining the gel on postoperative day 16. This patient underwent two washout procedures and resolution of the drainage. No infection was ever detected. Conclusions To our knowledge, our patients are the first reported cases of transcutaneous drainage of expanded dural sealant. It is important to take into consideration the unexpected expansion of a dural sealant when using it for the repair of IDT.

Construction and characterization of cDNA library for IRM-2 mice

International Nuclear Information System (INIS)

Wang Qin; Li Jin; Song Li; Liu Qiang; Yue Jingyin; Mu Chuanjie; Tang Weisheng; Fan Feiyue

2010-01-01

Objective: To screen and isolate the radioresistance related genes of IRM-2 mice. Methods: cDNA library of IRM-2 mice was constructed by SMART technique. Total RNA was isolated from spleens of IRM-2 male mice. The first-strand cDNA was synthesized by using PowerScript reverse transcriptase, and double-strand cDNA was synthesized and amplified by long PCR. The PCR products were purified, digested with restriction enzyme Sfi I. The ds-cDNA fragment less than 500 bp was fractionated and ligated to the Sfi I-digested pDNR-LIB vector. The ligation mixture was transformed into E. coil DH5 α by electroporation transformation to generate the unamplified cDNA library. The quality of cDNA library was identified by PCR technique. 130 clones from cDNA library were sequenced and compared with GenBank database. Results: The cDNA library contained 2.25 x 10 6 independent clones with an average insert size of 1.2 kb. The ratio of recombination and full-length was 95% and 55%, respectively. 21 pieces of EST sequences from cDNA library were not the same as the known mice genes and registered into GenBank EST database, with registered number DW474856-DW474876. Conclusions: cDNA library of IRM-2 mice has been constructed successfully. 21 pieces of EST implies that radioresistance correlative genes may be in IRM-2 mice, which will lay a foundation for isolating and identifying radioresistance related genes in further study. (authors)

cDNA encoding a polypeptide including a hev ein sequence

Energy Technology Data Exchange (ETDEWEB)

Raikhel, Natasha V. (Okemos, MI); Broekaert, Willem F. (Dilbeek, BE); Chua, Nam-Hai (Scarsdale, NY); Kush, Anil (New York, NY)

2000-07-04

A cDNA clone (HEV1) encoding hevein was isolated via polymerase chain reaction (PCR) using mixed oligonucleotides corresponding to two regions of hevein as primers and a Hevea brasiliensis latex cDNA library as a template. HEV1 is 1018 nucleotides long and includes an open reading frame of 204 amino acids. The deduced amino acid sequence contains a putative signal sequence of 17 amino acid residues followed by a 187 amino acid polypeptide. The amino-terminal region (43 amino acids) is identical to hevein and shows homology to several chitin-binding proteins and to the amino-termini of wound-induced genes in potato and poplar. The carboxyl-terminal portion of the polypeptide (144 amino acids) is 74-79% homologous to the carboxyl-terminal region of wound-inducible genes of potato. Wounding, as well as application of the plant hormones abscisic acid and ethylene, resulted in accumulation of hevein transcripts in leaves, stems and latex, but not in roots, as shown by using the cDNA as a probe. A fusion protein was produced in E. coli from the protein of the present invention and maltose binding protein produced by the E. coli.

Isolation and characterization of human cDNA clones encoding the α and the α' subunits of casein kinase II

International Nuclear Information System (INIS)

Lozeman, F.J.; Litchfield, D.W.; Piening, C.; Takio, Koji; Walsh, K.A.; Krebs, E.G.

1990-01-01

Casein kinase II is a widely distributed protein serine/threonine kinase. The holoenzyme appears to be a tetramer, containing two α or α' subunits (or one of each) and two β subunits. Complementary DNA clones encoding the subunits of casein kinase II were isolated from a human T-cell λgt 10 library using cDNA clones isolated from Drosophila melanogasten. One of the human cDNA clones (hT4.1) was 2.2 kb long, including a coding region of 1176 bp preceded by 156 bp (5' untranslated region) and followed by 871 bp (3' untranslated region). The hT4.1 close was nearly identical in size and sequence with a cDNA clone from HepG2 human hepatoma cultured cells. Another of the human T-cell cDNA clones (hT9.1) was 1.8 kb long, containing a coding region of 1053 bp preceded by 171 by (5' untranslated region) and followed by 550 bp (3' untranslated region). Amino acid sequences deduced from these two cDNA clones were about 85% identical. Most of the difference between the two encoded polypeptides was in the carboxy-terminal region, but heterogeneity was distributed throughout the molecules. Partial amino acid sequence was determined in a mixture of α and α' subunits from bovine lung casein kinase II. The bovine sequences aligned with the 2 human cDNA-encoded polypeptides with only 2 discrepancies out of 535 amino acid positions. This confirmed that the two human T-cell cDNA clones encoded the α and α' subunits of casein kinase II. These studies show that there are two distinct catalytic subunits for casein II (α and α') and that the sequence of these subunits is largely conserved between the bovine and the human

Unbiased stereologic techniques for practical use in diagnostic histopathology

DEFF Research Database (Denmark)

Sørensen, Flemming Brandt

1995-01-01

by introducing quantitative techniques in the histopathologic discipline of malignancy grading. Unbiased stereologic methods, especially based on measurements of nuclear three-dimensional mean size, have during the last decade proved their value in this regard. In this survey, the methods are reviewed regarding......Grading of malignancy by the examination of morphologic and cytologic details in histologic sections from malignant neoplasms is based exclusively on qualitative features, associated with significant subjectivity, and thus rather poor reproducibility. The traditional way of malignancy grading may...... of solid tumors. This new, unbiased attitude to malignancy grading is associated with excellent virtues, which ultimately may help the clinician in the choice of optimal treatment of the individual patient suffering from cancer. Stereologic methods are not solely applicable to the field of malignancy...

Hierarchical thinking in network biology: the unbiased modularization of biochemical networks.

Science.gov (United States)

Papin, Jason A; Reed, Jennifer L; Palsson, Bernhard O

2004-12-01

As reconstructed biochemical reaction networks continue to grow in size and scope, there is a growing need to describe the functional modules within them. Such modules facilitate the study of biological processes by deconstructing complex biological networks into conceptually simple entities. The definition of network modules is often based on intuitive reasoning. As an alternative, methods are being developed for defining biochemical network modules in an unbiased fashion. These unbiased network modules are mathematically derived from the structure of the whole network under consideration.

Progress in two-dimensional polyacrylamide gel electrophoresis and application in radiation research

International Nuclear Information System (INIS)

Wang Zhidong; Chen Xiaohua

2003-01-01

Two-dimensional polyacrylamide gel electrophoresis is the key separation technique in proteomics research, which is designed by protein character: molecular weight and PI. Some progress has been made in disease mechanism detection, tumor indicator research and drug development. This technique also has some potential application in radiation research

Cloning and cDNA sequence of the dihydrolipoamide dehydrogenase component of human α-ketoacid dehydrogenase complexes

International Nuclear Information System (INIS)

Pons, G.; Raefsky-Estrin, C.; Carothers, D.J.; Pepin, R.A.; Javed, A.A.; Jesse, B.W.; Ganapathi, M.K.; Samols, D.; Patel, M.S.

1988-01-01

cDNA clones comprising the entire coding region for human dihydrolipoamide dehydrogenase have been isolated from a human liver cDNA library. The cDNA sequence of the largest clone consisted of 2082 base pairs and contained a 1527-base open reading frame that encodes a precursor dihydrolipoamide dehydrogenase of 509 amino acid residues. The first 35-amino acid residues of the open reading frame probably correspond to a typical mitochondrial import leader sequence. The predicted amino acid sequence of the mature protein, starting at the residue number 36 of the open reading frame, is almost identical (>98% homology) with the known partial amino acid sequence of the pig heart dihydrolipoamide dehydrogenase. The cDNA clone also contains a 3' untranslated region of 505 bases with an unusual polyadenylylation signal (TATAAA) and a short poly(A) track. By blot-hybridization analysis with the cDNA as probe, two mRNAs, 2.2 and 2.4 kilobases in size, have been detected in human tissues and fibroblasts, whereas only one mRNA (2.4 kilobases) was detected in rat tissues

Characterization of the porcine carboxypeptidase E cDNA

DEFF Research Database (Denmark)

Hreidarsdôttir, G.E.; Cirera, Susanna; Fredholm, Merete

2007-01-01

the sequence of the cDNA for the porcine CPE gene including all the coding region and the 3'-UTR region was generated. Comparisons with bovine, human, mouse, and rat CPE cDNA sequences showed that the coding regions of the gene are highly conserved both at the nucleotide and at the amino acid level. A very low...

Fiscal 2000 report on result of the full-length cDNA structure analysis; 2000 nendo kanzen cho cDNA kozo kaiseki seika hokokusho

Energy Technology Data Exchange (ETDEWEB)

NONE

2001-03-01

This paper explains the results of research on full-length cDNA structure analysis for the period from April, 2000 to March, 2001. The outline of human genome sequence was published in June, 2000. In Japan, human gene analysis was such that, as the basic technology of the bio industry, a millennium project was decided in the budget of fiscal 2000. The full-length cDNA structure analysis is the core of the project. The libraries of cDNA were prepared using full-length and more than 4-5kbp-long cDNAs by oligo-capping method. It began from determining partial sequence data at end cDNA, and then, with new clones selected therefrom, full-length human cDNA sequence data were determined. The partial sequence data determined by fiscal 2000 were 1,035,000 clones while the full-length sequence data were 12,144 clones. The sequence data obtained were analyzed by homology search and translated into amino acid coding sequences, with predictions conducted on protein functions. A clustering method was examined that selects new clones from partial sequences. Database was constructed on gene expression profiles and disease-related gene sequence data. (NEDO)

[Study on the method of two dimensional polycrylamide gel electrophoresis on rat condylar chondrocyte].

Science.gov (United States)

Wu, Tuo-jiang; Li, Huang; Ma, Qiao-lin; Wang, Wen-mei

2010-08-01

To investigate the protein profile by two dimensional polycrylamide gel electrophoresis on the rat condylar chondrocyte in vitro. The third-passage chondrocytes were harvested from the mandibular condyles of 2-day-old rats in this study. The protein profile of the rat mandibular condylar chondrocytes was examined by two dimensional polycrylamide gel electrophoresis (2-DE-PAGE). The 2-DE gel maps on different pH gradients were obtained. The result of modified coomassi blue-sliver staining and sliver staining was compared using Pdquest 7.1 image analysis software. The results showed that the good protein profile of the condylar chondrocytes was obtained by standard Bio-Rad manual. The protein was mainly in the field from pH4 to pH7. The 1203±86 protein points were examined on 2-DE gel map by modified coomassi blue-sliver staining, and 1769±97 protein points was examined by sliver staining. The silver staining map showed more distinctly but higher background than modified coomassi blue-sliver staining. The protein profile of the condylar chondrocytes enriches the proteomic database and gives evidence to further proteomic research. The 2-DE map obtained by modified coomassi blue-sliver staining is more suitable for MALDI-TOF mass identification. Supported by National Natural Science Foundation of China (Grant No. C30700963), China Postdoctoral Science Foundation(Grant No.20090461088), Jiangsu Provincial Postdoctoral Science Foundation (Grant No.0802003C) and Nanjing City's Science and Technology Foundation (Grant No.200905011).

cDNA sequencing improves the detection of P53 missense mutations in colorectal cancer

International Nuclear Information System (INIS)

Szybka, Malgorzata; Kordek, Radzislaw; Zakrzewska, Magdalena; Rieske, Piotr; Pasz-Walczak, Grazyna; Kulczycka-Wojdala, Dominika; Zawlik, Izabela; Stawski, Robert; Jesionek-Kupnicka, Dorota; Liberski, Pawel P

2009-01-01

Recently published data showed discrepancies beteween P53 cDNA and DNA sequencing in glioblastomas. We hypothesised that similar discrepancies may be observed in other human cancers. To this end, we analyzed 23 colorectal cancers for P53 mutations and gene expression using both DNA and cDNA sequencing, real-time PCR and immunohistochemistry. We found P53 gene mutations in 16 cases (15 missense and 1 nonsense). Two of the 15 cases with missense mutations showed alterations based only on cDNA, and not DNA sequencing. Moreover, in 6 of the 15 cases with a cDNA mutation those mutations were difficult to detect in the DNA sequencing, so the results of DNA analysis alone could be misinterpreted if the cDNA sequencing results had not also been available. In all those 15 cases, we observed a higher ratio of the mutated to the wild type template by cDNA analysis, but not by the DNA analysis. Interestingly, a similar overexpression of P53 mRNA was present in samples with and without P53 mutations. In terms of colorectal cancer, those discrepancies might be explained under three conditions: 1, overexpression of mutated P53 mRNA in cancer cells as compared with normal cells; 2, a higher content of cells without P53 mutation (normal cells and cells showing K-RAS and/or APC but not P53 mutation) in samples presenting P53 mutation; 3, heterozygous or hemizygous mutations of P53 gene. Additionally, for heterozygous mutations unknown mechanism(s) causing selective overproduction of mutated allele should also be considered. Our data offer new clues for studying discrepancy in P53 cDNA and DNA sequencing analysis

Phenol emulsion-enhanced DNA-driven subtractive cDNA cloning: isolation of low-abundance monkey cortex-specific mRNAs

International Nuclear Information System (INIS)

Travis, G.H.; Sutcliffe, J.G.

1988-01-01

To isolate cDNA clones of low-abundance mRNAs expressed in monkey cerebral cortex but absent from cerebellum, the authors developed an improved subtractive cDNA cloning procedure that requires only modest quantities of mRNA. Plasmid DNA from a monkey cerebellum cDNA library was hybridized in large excess to radiolabeled monkey cortex cDNA in a phenol emulsion-enhanced reaction. The unhybridized cortex cDNA was isolated by chromatography on hydroxyapatite and used to probe colonies from a monkey cortex cDNA library. Of 60,000 colonies screened, 163 clones were isolated and confirmed by colony hybridization or RNA blotting to represent mRNAs, ranging from 0.001% to 0.1% abundance, specific to or highly enriched in cerebral cortex relative to cerebellum. Clones of one medium-abundance mRNA were recovered almost quantitatively. Two of the lower-abundance mRNAs were expressed at levels reduced by a factor of 10 in Alzheimer disease relative to normal human cortex. One of these was identified as the monkey preprosomatostatin I mRNA

Morphology-tunable and photoresponsive properties in a self-assembled two-component gel system.

Science.gov (United States)

Zhou, Yifeng; Xu, Miao; Yi, Tao; Xiao, Shuzhang; Zhou, Zhiguo; Li, Fuyou; Huang, Chunhui

2007-01-02

Photoresponsive C3-symmetrical trisurea self-assembling building blocks containing three azobenzene groups (LC10 and LC4) at the rim were designed and synthesized. By introducing a trisamide gelator (G18), which can self-aggregate through hydrogen bonds of acylamino moieties to form a fibrous network, the mixture of LC10 (or LC4) and G18 forms an organogel with coral-like supramolecular structure from 1,4-dioxane. The cooperation of hydrogen bonding and the hydrophobic diversity between these components are the main contributions to the specific superstructure. The two-component gel exhibits reversible photoisomerization from trans to cis transition without breakage of the gel state.

Categorization of rheological scaling models for particle gels applied to casein gels

NARCIS (Netherlands)

Mellema, M.; Opheusden, van J.H.J.; Vliet, van T.

2002-01-01

Rennet-induced casein gels made from skim milk were studied rheologically. A scaling model or framework for describing the rheological behavior of gels is discussed and used for classification of the structure of casein gels. There are two main parameters in the model that describe the number of

Electrophoretic analysis of proteins from Mycoplasma hominis strains detected by SDS-PAGE, two-dimensional gel electrophoresis and immunoblotting

DEFF Research Database (Denmark)

Andersen, H; Birkelund, Svend; Christiansen, Gunna

1987-01-01

The proteins of 14 strains of Mycoplasma hominis were compared by SDS-PAGE in gradient gels, by two-dimensional (2D) gel electrophoresis of extracts of 35S-labelled cells and by immunoblot analysis of cell proteins. The strains examined included the M. hominis type strain PG21 and 13 others...... isolated variously from genital tract, mouth, blood, upper urinary tract and a wound. These 14 strains shared 76-99% of proteins in SDS-gradient gel analysis and 41-72% in the 2D gels. As expected, the immunoblot analysis likewise revealed the existence of an extensive common protein pattern in M. hominis...

cDNA microarray screening in food safety

International Nuclear Information System (INIS)

Roy, Sashwati; Sen, Chandan K.

2006-01-01

The cDNA microarray technology and related bioinformatics tools presents a wide range of novel application opportunities. The technology may be productively applied to address food safety. In this mini-review article, we present an update highlighting the late breaking discoveries that demonstrate the vitality of cDNA microarray technology as a tool to analyze food safety with reference to microbial pathogens and genetically modified foods. In order to bring the microarray technology to mainstream food safety, it is important to develop robust user-friendly tools that may be applied in a field setting. In addition, there needs to be a standardized process for regulatory agencies to interpret and act upon microarray-based data. The cDNA microarray approach is an emergent technology in diagnostics. Its values lie in being able to provide complimentary molecular insight when employed in addition to traditional tests for food safety, as part of a more comprehensive battery of tests

cDNA structure, genomic organization and expression patterns of ...

African Journals Online (AJOL)

Visfatin was a newly identified adipocytokine, which was involved in various physiologic and pathologic processes of organisms. The cDNA structure, genomic organization and expression patterns of silver Prussian carp visfatin were described in this report. The silver Prussian carp visfatin cDNA cloned from the liver was ...

Construction of C35 gene bait recombinants and T47D cell cDNA library.

Science.gov (United States)

Yin, Kun; Xu, Chao; Zhao, Gui-Hua; Liu, Ye; Xiao, Ting; Zhu, Song; Yan, Ge

2017-11-20

C35 is a novel tumor biomarker associated with metastasis progression. To investigate the interaction factors of C35 in its high expressed breast cancer cell lines, we constructed bait recombinant plasmids of C35 gene and T47D cell cDNA library for yeast two-hybrid screening. Full length C35 sequences were subcloned using RT-PCR from cDNA template extracted from T47D cells. Based on functional domain analysis, the full-length C35 1-348bp was also truncated into two fragments C351-153bp and C35154-348bp to avoid auto-activation. The three kinds of C35 genes were successfully amplified and inserted into pGBKT7 to construct bait recombinant plasmids pGBKT7-C351-348bp, pGBKT7-C351-153bp and pGBKT7-C35154-348bp, then transformed into Y187 yeast cells by the lithium acetate method. Auto-activation and toxicity of C35 baits were detected using nutritional deficient medium and X-α-Gal assays. The T47D cell ds cDNA was generated by SMART TM technology and the library was constructed using in vivo recombination-mediated cloning in the AH109 yeast strain using a pGADT7-Rec plasmid. The transformed Y187/pGBKT7-C351-348bp line was intensively inhibited while the truncated Y187/pGBKT7-C35 lines had no auto-activation and toxicity in yeast cells. The titer of established cDNA library was 2 × 10 7 pfu/mL with high transformation efficiency of 1.4 × 10 6 , and the insert size of ds cDNA was distributed homogeneously between 0.5-2.0 kb. Our research generated a T47D cell cDNA library with high titer, and the constructed two C35 "baits" contained a respective functional immunoreceptor tyrosine based activation motif (ITAM) and the conserved last four amino acids Cys-Ile-Leu-Val (CILV) motif, and therefore laid a foundation for screening the C35 interaction factors in a BC cell line.

Infectious Maize rayado fino virus from cloned cDNA

Science.gov (United States)

Maize rayado fino virus (MRFV) is the type member of the marafiviruses within the family Tymoviridae. A cDNA clone from which infectious RNA can be transcribed was produced from a US isolate of MRFV (MRFV-US). Infectivity of transcripts derived from cDNA clones was demonstrated by infection of mai...

Molecular cloning of a human glycophorin B cDNA: nucleotide sequence and genomic relationship to glycophorin A

International Nuclear Information System (INIS)

Siebert, P.D.; Fukuda, M.

1987-01-01

The authors describe the isolation and nucleotide sequence of a human glycophorin B cDNA. The cDNA was identified by differential hybridization of synthetic oligonucleotide probes to a human erythroleukemic cell line (K562) cDNA library constructed in phage vector λgt10. The nucleotide sequence of the glycophorin B cDNA was compared with that of a previously cloned glycophorin A cDNA. The nucleotide sequences encoding the NH 2 -terminal leader peptide and first 26 amino acids of the two proteins are nearly identical. This homologous region is followed by areas specific to either glycophorin A or B and a number of small regions of homology, which in turn are followed by a very homologous region encoding the presumed membrane-spanning portion of the proteins. They used RNA blot hybridization with both cDNA and synthetic oligonucleotide probes to prove our previous hypothesis that glycophorin B is encoded by a single 0.5- to 0.6-kb mRNA and to show that glycophorins A and B are negatively and coordinately regulated by a tumor-promoting phorbol ester, phorbol 12-myristate 13-acetate. They established the intron/exon structure of the glycophorin A and B genes by oligonucleotide mapping; the results suggest a complex evolution of the glycophorin genes

Mouse tetranectin: cDNA sequence, tissue-specific expression, and chromosomal mapping

DEFF Research Database (Denmark)

Ibaraki, K; Kozak, C A; Wewer, U M

1995-01-01

regulation, mouse tetranectin cDNA was cloned from a 16-day-old mouse embryo library. Sequence analysis revealed a 992-bp cDNA with an open reading frame of 606 bp, which is identical in length to the human tetranectin cDNA. The deduced amino acid sequence showed high homology to the human cDNA with 76......(s) of tetranectin. The sequence analysis revealed a difference in both sequence and size of the noncoding regions between mouse and human cDNAs. Northern analysis of the various tissues from mouse, rat, and cow showed the major transcript(s) to be approximately 1 kb, which is similar in size to that observed...

A rapid method for screening arrayed plasmid cDNA library by PCR

International Nuclear Information System (INIS)

Hu Yingchun; Zhang Kaitai; Wu Dechang; Li Gang; Xiang Xiaoqiong

1999-01-01

Objective: To develop a PCR-based method for rapid and effective screening of arrayed plasmid cDNA library. Methods: The plasmid cDNA library was arrayed and screened by PCR with a particular set of primers. Results: Four positive clones were obtained through about one week. Conclusion: This method can be applied to screening not only normal cDNA clones, but also cDNA clones-containing small size fragments. This method offers significant advantages over traditional screening method in terms of sensitivity, specificity and efficiency

Application of Fluorescence Two-Dimensional Difference In-Gel Electrophoresis as a Proteomic Biomarker Discovery Tool in Muscular Dystrophy Research

Science.gov (United States)

Carberry, Steven; Zweyer, Margit; Swandulla, Dieter; Ohlendieck, Kay

2013-01-01

In this article, we illustrate the application of difference in-gel electrophoresis for the proteomic analysis of dystrophic skeletal muscle. The mdx diaphragm was used as a tissue model of dystrophinopathy. Two-dimensional gel electrophoresis is a widely employed protein separation method in proteomic investigations. Although two-dimensional gels usually underestimate the cellular presence of very high molecular mass proteins, integral membrane proteins and low copy number proteins, this method is extremely powerful in the comprehensive analysis of contractile proteins, metabolic enzymes, structural proteins and molecular chaperones. This gives rise to two-dimensional gel electrophoretic separation as the method of choice for studying contractile tissues in health and disease. For comparative studies, fluorescence difference in-gel electrophoresis has been shown to provide an excellent biomarker discovery tool. Since aged diaphragm fibres from the mdx mouse model of Duchenne muscular dystrophy closely resemble the human pathology, we have carried out a mass spectrometry-based comparison of the naturally aged diaphragm versus the senescent dystrophic diaphragm. The proteomic comparison of wild type versus mdx diaphragm resulted in the identification of 84 altered protein species. Novel molecular insights into dystrophic changes suggest increased cellular stress, impaired calcium buffering, cytostructural alterations and disturbances of mitochondrial metabolism in dystrophin-deficient muscle tissue. PMID:24833232

cDNA sequence of human transforming gene hst and identification of the coding sequence required for transforming activity

International Nuclear Information System (INIS)

Taira, M.; Yoshida, T.; Miyagawa, K.; Sakamoto, H.; Terada, M.; Sugimura, T.

1987-01-01

The hst gene was originally identified as a transforming gene in DNAs from human stomach cancers and from a noncancerous portion of stomach mucosa by DNA-mediated transfection assay using NIH3T3 cells. cDNA clones of hst were isolated from the cDNA library constructed from poly(A) + RNA of a secondary transformant induced by the DNA from a stomach cancer. The sequence analysis of the hst cDNA revealed the presence of two open reading frames. When this cDNA was inserted into an expression vector containing the simian virus 40 promoter, it efficiently induced the transformation of NIH3T3 cells upon transfection. It was found that one of the reading frames, which coded for 206 amino acids, was responsible for the transforming activity

Cloning and analysis of the mouse Fanconi anemia group A cDNA and an overlapping penta zinc finger cDNA.

Science.gov (United States)

Wong, J C; Alon, N; Norga, K; Kruyt, F A; Youssoufian, H; Buchwald, M

2000-08-01

Despite the cloning of four disease-associated genes for Fanconi anemia (FA), the molecular pathogenesis of FA remains largely unknown. To study FA complementation group A using the mouse as a model system, we cloned and characterized the mouse homolog of the human FANCA cDNA. The mouse cDNA (Fanca) encodes a 161-kDa protein that shares 65% amino acid sequence identity with human FANCA. Fanca is located at the distal region of mouse chromosome 8 and has a ubiquitous pattern of expression in embryonic and adult tissues. Expression of the mouse cDNA in human FA-A cells restores the cellular drug sensitivity to normal levels. Thus, the expression pattern, protein structure, chromosomal location, and function of FANCA are conserved in the mouse. We also isolated a novel zinc finger protein, Zfp276, which has five C(2)H(2) domains. Interestingly, Zfp276 is situated in the Fanca locus, and the 3'UTR of its cDNA overlaps with the last four exons of Fanca in a tail-to-tail manner. Zfp276 is expressed in the same tissues as Fanca, but does not complement the mitomycin C (MMC)-sensitive phenotype of FA-A cells. The overlapping genomic organization between Zfp276 and Fanca may have relevance to the disease phenotype of FA. Copyright 2000 Academic Press.

Applications of gel dosimetry

International Nuclear Information System (INIS)

Ibbott, Geoffrey S

2004-01-01

Gel dosimetry has been examined as a clinical dosimeter since the 1950s. During the last two decades, however, a rapid increase in the number of investigators has been seen, and the body of knowledge regarding gel dosimetry has expanded considerably. Gel dosimetry is still considered a research project, and the introduction of this tool into clinical use is proceeding slowly. This paper will review the characteristics of gel dosimetry that make it desirable for clinical use, the postulated and demonstrated applications of gel dosimetry, and some complications, set-backs, and failures that have contributed to the slow introduction into routine clinical use

About mutually unbiased bases in even and odd prime power dimensions

Science.gov (United States)

Durt, Thomas

2005-06-01

Mutually unbiased bases generalize the X, Y and Z qubit bases. They possess numerous applications in quantum information science. It is well known that in prime power dimensions N = pm (with p prime and m a positive integer), there exists a maximal set of N + 1 mutually unbiased bases. In the present paper, we derive an explicit expression for those bases, in terms of the (operations of the) associated finite field (Galois division ring) of N elements. This expression is shown to be equivalent to the expressions previously obtained by Ivanovic (1981 J. Phys. A: Math. Gen. 14 3241) in odd prime dimensions, and Wootters and Fields (1989 Ann. Phys. 191 363) in odd prime power dimensions. In even prime power dimensions, we derive a new explicit expression for the mutually unbiased bases. The new ingredients of our approach are, basically, the following: we provide a simple expression of the generalized Pauli group in terms of the additive characters of the field, and we derive an exact groupal composition law between the elements of the commuting subsets of the generalized Pauli group, renormalized by a well-chosen phase-factor.

About mutually unbiased bases in even and odd prime power dimensions

International Nuclear Information System (INIS)

Durt, Thomas

2005-01-01

Mutually unbiased bases generalize the X, Y and Z qubit bases. They possess numerous applications in quantum information science. It is well known that in prime power dimensions N = p m (with p prime and m a positive integer), there exists a maximal set of N + 1 mutually unbiased bases. In the present paper, we derive an explicit expression for those bases, in terms of the (operations of the) associated finite field (Galois division ring) of N elements. This expression is shown to be equivalent to the expressions previously obtained by Ivanovic (1981 J. Phys. A: Math. Gen. 14 3241) in odd prime dimensions, and Wootters and Fields (1989 Ann. Phys. 191 363) in odd prime power dimensions. In even prime power dimensions, we derive a new explicit expression for the mutually unbiased bases. The new ingredients of our approach are, basically, the following: we provide a simple expression of the generalized Pauli group in terms of the additive characters of the field, and we derive an exact groupal composition law between the elements of the commuting subsets of the generalized Pauli group, renormalized by a well-chosen phase-factor

Ridge regression estimator: combining unbiased and ordinary ridge regression methods of estimation

Directory of Open Access Journals (Sweden)

Sharad Damodar Gore

2009-10-01

Full Text Available Statistical literature has several methods for coping with multicollinearity. This paper introduces a new shrinkage estimator, called modified unbiased ridge (MUR. This estimator is obtained from unbiased ridge regression (URR in the same way that ordinary ridge regression (ORR is obtained from ordinary least squares (OLS. Properties of MUR are derived. Results on its matrix mean squared error (MMSE are obtained. MUR is compared with ORR and URR in terms of MMSE. These results are illustrated with an example based on data generated by Hoerl and Kennard (1975.

Cloning and characterization of the human colipase cDNA

International Nuclear Information System (INIS)

Lowe, M.E.; Rosenblum, J.L.; McEwen, P.; Strauss, A.W.

1990-01-01

Pancreatic lipase hydrolyzes dietary triglycerides to monoglycerides and fatty acids. In the presence of bile salts, the activity of pancreatic lipase is markedly decreased. The activity can be restored by the addition of colipase, a low molecular weight protein secreted by the pancreas. The action of pancreatic lipase in the gut lumen is dependent upon its interaction with colipase. As a first step in elucidating the molecular events governing the interaction of lipase and colipase with each other and with fatty acids, a cDNA encoding human colipase was isolated from a λgt11 cDNA library with a rabbit polyclonal anti-human colipase antibody. The full-length 525 bp cDNA contained an open reading frame encoding 112 amino acids, including a 17 amino acid signal peptide. The predicted sequence contains 100% of the published protein sequence for human colipase determined by chemical methods, but predicts the presence of five additional NH 2 -terminal amino acids and four additional COOH-terminal amino acids. Comparison of the predicted protein sequence with the known sequences of colipase from other species reveals regions of extensive identity. The authors report, for the first time, a cDNA for colipase. The cDNA predicts a human procolipase an suggests that there may also be processing at the COOH-terminus. The regions of identity with colipase from other species will aid in defining the interaction with lipase and lipids through site-specific mutagenesis

Characterization of cDNA for human tripeptidyl peptidase II: The N-terminal part of the enzyme is similar to subtilisin

International Nuclear Information System (INIS)

Tomkinson, B.; Jonsson, A-K

1991-01-01

Tripeptidyl peptidase II is a high molecular weight serine exopeptidase, which has been purified from rat liver and human erythrocytes. Four clones, representing 4453 bp, or 90% of the mRNA of the human enzyme, have been isolated from two different cDNA libraries. One clone, designated A2, was obtained after screening a human B-lymphocyte cDNA library with a degenerated oligonucleotide mixture. The B-lymphocyte cDNA library, obtained from human fibroblasts, were rescreened with a 147 bp fragment from the 5' part of the A2 clone, whereby three different overlapping cDNA clones could be isolated. The deduced amino acid sequence, 1196 amino acid residues, corresponding to the longest open rading frame of the assembled nucleotide sequence, was compared to sequences of current databases. This revealed a 56% similarity between the bacterial enzyme subtilisin and the N-terminal part of tripeptidyl peptidase II. The enzyme was found to be represented by two different mRNAs of 4.2 and 5.0 kilobases, respectively, which probably result from the utilziation of two different polyadenylation sites. Futhermore, cDNA corresponding to both the N-terminal and C-terminal part of tripeptidyl peptidase II hybridized with genomic DNA from mouse, horse, calf, and hen, even under fairly high stringency conditions, indicating that tripeptidyl peptidase II is highly conserved

Optimizing human synovial fluid preparation for two-dimensional gel electrophoresis.

Science.gov (United States)

Chen, Carl Pc; Hsu, Chih-Chin; Yeh, Wen-Lin; Lin, Hsiu-Chu; Hsieh, Sen-Yung; Lin, Shih-Cherng; Chen, Tai-Tzung; Chen, Max Jl; Tang, Simon Ft

2011-10-11

Proteome analysis is frequently applied in identifying the proteins or biomarkers in knee synovial fluids (SF) that are associated with osteoarthritis and other arthritic disorders. The 2-dimensional gel electrophoresis (2-DE) is the technique of choice in these studies. Disease biomarkers usually appear in low concentrations and may be masked by high abundant proteins. Therefore, the main aim of this study was to find the most suitable sample preparation method that can optimize the expression of proteins on 2-DE gels that can be used to develop a reference proteome picture for non-osteoarthritic knee synovial fluid samples. Proteome pictures obtained from osteoarthritic knee synovial fluids can then be compared with the reference proteome pictures obtained in this study to assist us in identifying the disease biomarkers more correctly. The proteomic tool of 2-DE with immobilized pH gradients was applied in this study. A total of 12 2-DE gel images were constructed from SF samples that were free of osteoarthritis. In these samples, 3 were not treated with any sample preparation methods, 3 were treated with acetone, 3 were treated with 2-DE Clean-Up Kit, and 3 were treated with the combination of acetone and 2-D Clean-Up Kit prior to 2-DE analysis. Gel images were analyzed using the PDQuest Basic 8.0.1 Analytical software. Protein spots that were of interest were excised from the gels and sent for identification by mass spectrometry. Total SF total protein concentration was calculated to be 21.98 ± 0.86 mg/mL. The untreated SF samples were detected to have 456 ± 33 protein spots on 2-DE gel images. Acetone treated SF samples were detected to have 320 ± 28 protein spots, 2-D Clean-Up Kit treated SF samples were detected to have 413 ± 31 protein spots, and the combined treatment method of acetone and 2-D Clean-Up Kit was detected to have 278 ± 26 protein spots 2-DE gel images. SF samples treated with 2-D Clean-Up Kit revealed clearer presentation of the isoforms

Optimizing Human Synovial Fluid Preparation for Two-Dimensional Gel Electrophoresis

Directory of Open Access Journals (Sweden)

Chen Max JL

2011-10-01

Full Text Available Abstract Background Proteome analysis is frequently applied in identifying the proteins or biomarkers in knee synovial fluids (SF that are associated with osteoarthritis and other arthritic disorders. The 2-dimensional gel electrophoresis (2-DE is the technique of choice in these studies. Disease biomarkers usually appear in low concentrations and may be masked by high abundant proteins. Therefore, the main aim of this study was to find the most suitable sample preparation method that can optimize the expression of proteins on 2-DE gels that can be used to develop a reference proteome picture for non-osteoarthritic knee synovial fluid samples. Proteome pictures obtained from osteoarthritic knee synovial fluids can then be compared with the reference proteome pictures obtained in this study to assist us in identifying the disease biomarkers more correctly. Results The proteomic tool of 2-DE with immobilized pH gradients was applied in this study. A total of 12 2-DE gel images were constructed from SF samples that were free of osteoarthritis. In these samples, 3 were not treated with any sample preparation methods, 3 were treated with acetone, 3 were treated with 2-DE Clean-Up Kit, and 3 were treated with the combination of acetone and 2-D Clean-Up Kit prior to 2-DE analysis. Gel images were analyzed using the PDQuest Basic 8.0.1 Analytical software. Protein spots that were of interest were excised from the gels and sent for identification by mass spectrometry. Total SF total protein concentration was calculated to be 21.98 ± 0.86 mg/mL. The untreated SF samples were detected to have 456 ± 33 protein spots on 2-DE gel images. Acetone treated SF samples were detected to have 320 ± 28 protein spots, 2-D Clean-Up Kit treated SF samples were detected to have 413 ± 31 protein spots, and the combined treatment method of acetone and 2-D Clean-Up Kit was detected to have 278 ± 26 protein spots 2-DE gel images. SF samples treated with 2-D Clean-Up Kit

Mapping and identification of interferon gamma-regulated HeLa cell proteins separated by immobilized pH gradient two-dimensional gel electrophoresis

DEFF Research Database (Denmark)

Shaw, A.; Larsen, M.; Roepstorff, P.

1999-01-01

magnitude of IFN-gamma responsive genes has been reported previously. Our goal is to identify and map IFN-gamma-regulated HeLa cell proteins to the two-dimensional polyacrylamide gel electrophoresis with the immobilized pH gradient (IPG) two-dimensional polyacrylamide gel electrophoresis (2-D PAGE) system...

c-DNA of HIV-1 detection on spot of Buffy-Coat of leukocytes (DBCS

Directory of Open Access Journals (Sweden)

Marco Rossi de Gasperis

2010-03-01

Full Text Available Introduction:The elective way for the diagnosis of HIV-1-infection in the window period and in children under the age of 16-18 months is to search virus integrated in leukocytes. Aim of the study was to assess the sensitivity and specificity of extraction from Buffy-Dried Coat Spot (DBCS in leukocyte to detect c-DNA with nested-PCR in HIV-1-infected individuals compared to Dried Blood Spot (DBS both extracted by automated instrument EZ1 (QIAGEN, Hilden, Germany. Both DBCS and both DBS were compared with those tests from whole blood by conventional DNA-extraction Methods: Five ml of whole blood from 50 HIV-infected individuals were collected. 40 μl of each sample were spotted on “FTA ELUTE Micro Card” (Whatman, Inc., Clifton, NJ, 200 μl were extracted according to the manual procedure (QIAGEN “QIAamp DNA minikit and the remaining sample was incubated at 37 °C for 120 minutes. Plasma was centrifuged at 1000 rcf/1g for 10 minutes at room temperature. Forty μl of the obtained buffy-coat was spotted. Both DBCS and both DBS were dried at room temperature for 24 hours.Two of 5 punch from each spot were extracted with TISSUE DNA kit (Biorobot EZ1 DSP “Qiagen” and eluted in 50 μl of buffer.The recovery of genomic DNA was measured amplifying the ß-globin gene by Real-Time “SybrGreen I”.The DNA was amplified for the “pol” gene of HIV-1 by nested PCR and revealed in “SYBR-green I”. Eight HIV-antibody-negative samples were used as internal control. Results:The experimental protocol adopted for the DBCS showed high sensitivity and specificity.The extracted DNA from DBS and DBCS was characterized by excellent quality and without any inhibitory agents. The amount of proviral DNA extracted from DBCS is similar to that obtained by conventional extraction, while the DBS test was significantly less sensitive. Conclusion:These preliminary data suggest that the amount of c-DNA obtained with DBS technique is often not enough for the

Cloning and expression of human deoxycytidine kinase cDNA

International Nuclear Information System (INIS)

Chottiner, E.G.; Shewach, D.S.; Datta, N.S.; Ashcraft, E.; Gribbin, D.; Ginsburg, D.; Fox, I.H.; Mitchell, B.S.

1991-01-01

Deoxycytidine (dCyd) kinase is required for the phosphorylation of several deoxyribonucleosides and certain nucleoside analogs widely employed as antiviral and chemotherapeutic agents. Detailed analysis of this enzyme has been limited, however, by its low abundance and instability. Using oligonucleotides based on primary amino acid sequence derived from purified dCyd kinase, the authors have screened T-lymphoblast cDNA libraries and identified a cDNA sequence that encodes a 30.5-kDa protein corresponding to the subunit molecular mass of the purified protein. Expression of the cDNA in Escherichia coli results in a 40-fold increase in dCyd kinase activity over control levels. Northern blot analysis reveals a single 2.8-kilobase mRNA expressed in T lymphoblasts at 5- to 10-fold higher levels than in B lymphoblasts, and decreased dCyd kinase mRNA levels are present in T-lymphoblast cell lines resistant to arabinofuranosylcytosine and dideoxycytidine. These findings document that this cDNA encodes the T-lymphoblast dCyd kinase responsible for the phosphorylation of dAdo and dGuo as well as dCyd and arabinofuranosylcytosine

Correlation of acidic and basic carrier ampholyte and immobilized pH gradient two-dimensional gel electrophoresis patterns based on mass spectrometric protein identification

DEFF Research Database (Denmark)

Nawrocki, A; Larsen, Martin Røssel; Podtelejnikov, A V

1998-01-01

Separation of proteins on either carrier ampholyte-based or immobilized pH gradient-based two-dimensional (2-D) gels gives rise to electrophoretic patterns that are difficult to compare visually. In this paper we have used matrix-assisted laser desorption/ionization mass spectrometry (MALDI......-MS) to determine the identities of 335 protein spots in these two 2-D gel systems, including a substantial number of basic proteins which had never been identified before. Proteins that were identified in both gel systems allowed us to cross-reference the gel patterns. Vector analysis of these cross...

Polymer gel dosimeters with enhanced sensitivity for use in x-ray CT polymer gel dosimetry

International Nuclear Information System (INIS)

Jirasek, A; Hilts, M; McAuley, K B

2010-01-01

A primary limitation of current x-ray CT polymer gel dosimetry is the low contrast, and hence poor dose resolution, of dose images produced by the system. The low contrast is largely due to the low-dose sensitivity of current formulations of polymer gel for x-ray CT imaging. This study reports on the investigation of new dosimeter formulations with improved dose sensitivity for x-ray CT polymer gel dosimetry. We incorporate an isopropanol co-solvent into an N-isopropylacrylamide-based gel formulation in order to increase the total monomer/crosslinker concentration (%T) within the formulation. It is shown that gels of high %T exhibit enhanced dose sensitivity and dose resolutions over traditional formulations. The gels are shown to be temporally stable and reproducible. A single formulation (16%T) is used to demonstrate the capabilities of the x-ray CT polymer gel dosimetry system in measuring known dose distributions. A 1 L gel volume is exposed to three separate irradiations: a single-field percent depth dose, a two-field 'cross' and a three-field 'test case'. The first two irradiations are used to generate a dose calibration curve by which images are calibrated. The calibrated images are compared with treatment planning predictions and it is shown that the x-ray CT polymer gel dosimetry system is capable of capturing spatial and dose information accurately. The proposed new gel formulation is shown to be sensitive, stable and to improve the dose resolution over current formulations so as to provide a feasible gel for clinical applications of x-ray CT polymer gel dosimetry.

Losing the rose tinted glasses: neural substrates of unbiased belief updating in depression

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Neil eGarrett

2014-08-01

Full Text Available Recent evidence suggests that a state of good mental health is associated with biased processing of information that supports a positively skewed view of the future. Depression, on the other hand, is associated with unbiased processing of such information. Here, we use brain imaging in conjunction with a belief update task administered to clinically depressed patients and healthy controls to characterize brain activity that supports unbiased belief updating in clinically depressed individuals. Our results reveal that unbiased belief updating in depression is mediated by strong neural coding of estimation errors in response to both good news (in left inferior frontal gyrus and bilateral superior frontal gyrus and bad news (in right inferior parietal lobule and right inferior frontal gyrus regarding the future. In contrast, intact mental health was linked to a relatively attenuated neural coding of bad news about the future. These findings identify a neural substrate mediating the breakdown of biased updating in Major Depression Disorder, which may be essential for mental health.

Proteome analysis of barley seeds: Identification of major proteins from two-dimensional gels (pl 4-7)

DEFF Research Database (Denmark)

Østergaard, O.; Finnie, Christine; Laugesen, S.

2004-01-01

inhibitors), and proteins related to desiccation and oxidative stress. Sixty-four of the identifications were made using expressed sequence tags (ESTs). Numerous spots in the 2-D gel pattern changed during germination (micromalting) and an intensely stained area which contained large amounts of the serpin......Germination of monocotyledonous plants involves activation and de novo synthesis of enzymes that degrade cell walls and starch and mobilize stored endosperm reserves for embryo growth. Two-dimensional (2-D) gel electrophoresis and mass spectrometry were applied to identify major water...

Cloning and characterization of transferrin cDNA and rapid detection of transferrin gene polymorphism in rainbow trout (Oncorhynchus mykiss).

Science.gov (United States)

Tange, N; Jong-Young, L; Mikawa, N; Hirono, I; Aoki, T

1997-12-01

A cDNA clone of rainbow trout (Oncorhynchus mykiss) transferrin was obtained from a liver cDNA library. The 2537-bp cDNA sequence contained an open reading frame encoding 691 amino acids and the 5' and 3' noncoding regions. The amino acid sequences at the iron-binding sites and the two N-linked glycosylation sites, and the cysteine residues were consistent with known, conserved vertebrate transferrin cDNA sequences. Single N-linked glycosylation sites existed on the N- and C-lobe. The deduced amino acid sequence of the rainbow trout transferrin cDNA had 92.9% identities with transferrin of coho salmon (Oncorhynchus kisutch); 85%, Atlantic salmon (Salmo salar); 67.3%, medaka (Oryzias latipes); 61.3% Atlantic cod (Gadus morhua); and 59.7%, Japanese flounder (Paralichthys olivaceus). The long and accurate polymerase chain reaction (LA-PCR) was used to amplify approximately 6.5 kb of the transferrin gene from rainbow trout genomic DNA. Restriction fragment length polymorphisms (RFLPs) of the LA-PCR products revealed three digestion patterns in 22 samples.

Polymer gel dosimeters with enhanced sensitivity for use in x-ray CT polymer gel dosimetry

Energy Technology Data Exchange (ETDEWEB)

Jirasek, A [Department of Physics and Astronomy, University of Victoria, Victoria BC V8W 3P6 (Canada); Hilts, M [Medical Physics, BC Cancer Agency-Vancouver Island Centre, Victoria BC V6R 2B6 (Canada); McAuley, K B, E-mail: jirasek@uvic.c [Department of Chemical Engineering, Queens University, Kingston, ON K7 L 3N6 (Canada)

2010-09-21

A primary limitation of current x-ray CT polymer gel dosimetry is the low contrast, and hence poor dose resolution, of dose images produced by the system. The low contrast is largely due to the low-dose sensitivity of current formulations of polymer gel for x-ray CT imaging. This study reports on the investigation of new dosimeter formulations with improved dose sensitivity for x-ray CT polymer gel dosimetry. We incorporate an isopropanol co-solvent into an N-isopropylacrylamide-based gel formulation in order to increase the total monomer/crosslinker concentration (%T) within the formulation. It is shown that gels of high %T exhibit enhanced dose sensitivity and dose resolutions over traditional formulations. The gels are shown to be temporally stable and reproducible. A single formulation (16%T) is used to demonstrate the capabilities of the x-ray CT polymer gel dosimetry system in measuring known dose distributions. A 1 L gel volume is exposed to three separate irradiations: a single-field percent depth dose, a two-field 'cross' and a three-field 'test case'. The first two irradiations are used to generate a dose calibration curve by which images are calibrated. The calibrated images are compared with treatment planning predictions and it is shown that the x-ray CT polymer gel dosimetry system is capable of capturing spatial and dose information accurately. The proposed new gel formulation is shown to be sensitive, stable and to improve the dose resolution over current formulations so as to provide a feasible gel for clinical applications of x-ray CT polymer gel dosimetry.

A rapid and efficient two-step gel electrophoresis method for the purification of major rye grass pollen allergens.

Science.gov (United States)

Levy, D; Davies, J; O'Hehir, R; Suphioglu, C

2001-06-01

Purified proteins are mandatory for molecular, immunological and cellular studies. However, purification of proteins from complex mixtures requires specialised chromatography methods (i.e., gel filtration, ion exchange, etc.) using fast protein liquid chromatography (FPLC) or high-performance liquid chromatography (HPLC) systems. Such systems are expensive and certain proteins require two or more different steps for sufficient purity and generally result in low recovery. The aim of this study was to develop a rapid, inexpensive and efficient gel-electrophoresis-based protein purification method using basic and readily available laboratory equipment. We have used crude rye grass pollen extract to purify the major allergens Lol p 1 and Lol p 5 as the model protein candidates. Total proteins were resolved on large primary gel and Coomassie Brilliant Blue (CBB)-stained Lol p 1/5 allergens were excised and purified on a secondary "mini"-gel. Purified proteins were extracted from unstained separating gels and subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblot analyses. Silver-stained SDS-PAGE gels resolved pure proteins (i.e., 875 microg of Lol p 1 recovered from a 8 mg crude starting material) while immunoblot analysis confirmed immunological reactivity of the purified proteins. Such a purification method is rapid, inexpensive, and efficient in generating proteins of sufficient purity for use in monoclonal antibody (mAb) production, protein sequencing and general molecular, immunological, and cellular studies.

5'-end sequences of budding yeast full-length cDNA clones and quality scores - Budding yeast cDNA sequencing project | LSDB Archive [Life Science Database Archive metadata

Lifescience Database Archive (English)

Full Text Available east_seq_qual.zip File URL: ftp://ftp.biosciencedbc.jp/archive/yeast_cdna/LATEST/...yeast_seq_qual.zip File size: 59.9MB Simple search URL http://togodb.biosciencedbc.jp/togodb/view/budding_yeast_cdna

Cdna cloning and expression analyses of the isoflavone reductase-like gene of dendrobium officinale

International Nuclear Information System (INIS)

Qian, X.; Xu, S.Z.

2015-01-01

The full length of the isoflavone reductase-like gene (IRL) cDNA of Dendrobium officinale was cloned by using reverse transcription (RT) PCR combined with cDNA library, the IRL function was identified by Bioinformatics and prokaryotic expression analyses, and the IRL expression levels in the organs and tissues of D. officinale plants with different ages were determined by using real-time quantitative PCR (RT-qPCR). The results indicated that the full length of the cDNA of D. officinale IRL, DoIRL, was 1238 bp (accession no. KJ661023). Its open reading frame (ORF) was 930 bp which encoded 309 amino acids with a predicted molecular mass of 34 kDa, the 5 untranslated region (UTR) was 61 bp and the 3 UTR containing a poly (A) tail was 247 bp. The deduced amino acid sequence of DoIRL, DoIRL, was forecast to contain a NAD(P)H-binding motif (GGTGYIG) in the N-terminal region, two conserved N-glycosylation sites, a conserved nitrogen metabolite repression regulator (NmrA) domain and a phenylcoumaran benzylic ether reductase (PCBER) domain, to hold the nearest phylogenetic relationship with the PCBER of Striga asiatica, and to share both 73% identity with the isoflavone reductases-like (IRLs) of Cucumis sativus and Striga asiatica. In Escherichia coli 'BL21' cells, the DoIRL cDNA expression produced a protein band holding the predicted molecular mass of 34 kDa. DoIRL expressed in all organs and tissues of D. officinale plants with different ages at comparatively low levels, and the expression level in the leaves of the two-year-old plants was the highest. (author)

Binding proteins for the regulatory subunit (RII-B) of brain cAMP-dependent protein kinase II: isolation and initial characterization of cDNA clones

International Nuclear Information System (INIS)

Bregman, D.B.; Hu, E.; Rubin, C.S.

1987-01-01

In mammalian brain several proteins bind RII-B with high affinity. An example is P75, which co-purifies with RII-B and also complexes Ca 2+ -calmodulin. Thus, RII-B binding proteins (RBPs) might play a role in integrating the Ca 2+ and cAMP signalling pathways in the CNS. In order to study the structure and function of these polypeptides they have isolated cloned cDNAs for RBPs by screening brain λgt11 expression libraries using a functional assay: the binding of 32 P-labeled RII to fusion proteins produced by recombinants expressing RII binding domains. Inserts from rat brain recombinant clones λ7B and λ10B both hybridize to a brain mRNA of 7000 nucleotides. Northern gel analyses indicate that the putative RBP mRNA is also expressed in lung, but not in several other tissues. The λ7B insert was subcloned into the expression plasmid pINIA. A 50 kDa high affinity RII-B binding polypeptide accumulated in E. coli transformed with pINIA-7B. Two RBP cDNAs (λ77, λ100A) have been retrieved from a bovine λgt 11 library using a monoclonal antibody directed against P75 and the binding assay respectively. On Southern blots the insert from λ100A hybridizes to the cDNA insert from clones λ77, suggesting that λ 77 cDNA might contain sequences coding for both an RII binding domain and a P75 epitope. The bovine λ100A insert also hybridizes with the rat λ7B clone indicating that an RII binding domain is conserved in the two species

Cloning of cDNA encoding steroid 11β-hydroxylase (P450c11)

International Nuclear Information System (INIS)

Chua, S.C.; Szabo, P.; Vitek, A.; Grzeschik, K.H.; John, M.; White, P.C.

1987-01-01

The authors have isolated bovine and human adrenal cDNA clones encoding the adrenal cytochrome P-450 specific for 11β-hydroxylation (P450c11). A bovine adrenal cDNA library constructed in the bacteriophage λ vector gt10 was probed with a previously isolated cDNA clone corresponding to part of the 3' untranslated region of the 4.2-kilobase (kb) mRNA encoding P450c11. Several clones with 3.2-kb cDNA inserts were isolated. Sequence analysis showed that they overlapped the original probe by 300 base pairs (bp). Combined cDNA and RNA sequence data demonstrated a continuous open reading frame of 1509 bases. P450c11 is predicted to contain 479 amino acid residues in the mature protein in addition to a 24-residue amino-terminal mitochondrial signal sequence. A bovine clone was used to isolate a homologous clone with a 3.5-kb insert from a human adrenal cDNA library. A region of 1100 bp was 81% homologous to 769 bp of the coding sequence of the bovine cDNA except for a 400-bp segment presumed to be an unprocessed intron. Hybridization of the human cDNA to DNA from a panel of human-rodent somatic cell hybrid lines and in situ hybridization to metaphase spreads of human chromosomes localized the gene to the middle of the long arm of chromosome 8. These data should be useful in developing reagents for heterozygote detection and prenatal diagnosis of 11β-hydroxylase deficiency, the second most frequent cause of congenital adrenal hyperplasia

Avoiding acidic region streaking in two-dimensional gel ...

Indian Academy of Sciences (India)

Supplementary figure 6. 2DE gel images ... Number of acidic streaks. Fedyunin et al. 2012. 4.02. 6. Zuo et al. 2000. 2.54. 9. Valenete et ... CE, 3rd 2009 Proteasomal protein degradation in ... Nandakumar MP, Shen J, Raman B and Marten MR.

Transport Phenomena in Gel

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Masayuki Tokita

2016-05-01

Full Text Available Gel becomes an important class of soft materials since it can be seen in a wide variety of the chemical and the biological systems. The unique properties of gel arise from the structure, namely, the three-dimensional polymer network that is swollen by a huge amount of solvent. Despite the small volume fraction of the polymer network, which is usually only a few percent or less, gel shows the typical properties that belong to solids such as the elasticity. Gel is, therefore, regarded as a dilute solid because its elasticity is much smaller than that of typical solids. Because of the diluted structure, small molecules can pass along the open space of the polymer network. In addition to the viscous resistance of gel fluid, however, the substance experiences resistance due to the polymer network of gel during the transport process. It is, therefore, of importance to study the diffusion of the small molecules in gel as well as the flow of gel fluid itself through the polymer network of gel. It may be natural to assume that the effects of the resistance due to the polymer network of gel depends strongly on the network structure. Therefore, detailed study on the transport processes in and through gel may open a new insight into the relationship between the structure and the transport properties of gel. The two typical transport processes in and through gel, that is, the diffusion of small molecules due to the thermal fluctuations and the flow of gel fluid that is caused by the mechanical pressure gradient will be reviewed.

Full-length cDNA sequences from Rhesus monkey placenta tissue: analysis and utility for comparative mapping

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Lee Sang-Rae

2010-07-01

Full Text Available Abstract Background Rhesus monkeys (Macaca mulatta are widely-used as experimental animals in biomedical research and are closely related to other laboratory macaques, such as cynomolgus monkeys (Macaca fascicularis, and to humans, sharing a last common ancestor from about 25 million years ago. Although rhesus monkeys have been studied extensively under field and laboratory conditions, research has been limited by the lack of genetic resources. The present study generated placenta full-length cDNA libraries, characterized the resulting expressed sequence tags, and described their utility for comparative mapping with human RefSeq mRNA transcripts. Results From rhesus monkey placenta full-length cDNA libraries, 2000 full-length cDNA sequences were determined and 1835 rhesus placenta cDNA sequences longer than 100 bp were collected. These sequences were annotated based on homology to human genes. Homology search against human RefSeq mRNAs revealed that our collection included the sequences of 1462 putative rhesus monkey genes. Moreover, we identified 207 genes containing exon alterations in the coding region and the untranslated region of rhesus monkey transcripts, despite the highly conserved structure of the coding regions. Approximately 10% (187 of all full-length cDNA sequences did not represent any public human RefSeq mRNAs. Intriguingly, two rhesus monkey specific exons derived from the transposable elements of AluYRa2 (SINE family and MER11B (LTR family were also identified. Conclusion The 1835 rhesus monkey placenta full-length cDNA sequences described here could expand genomic resources and information of rhesus monkeys. This increased genomic information will greatly contribute to the development of evolutionary biology and biomedical research.

Identification of Cysteine Proteases and Screening of Cysteine Protease Inhibitors in Biological Samples by a Two-Dimensional Gel System of Zymography and Reverse Zymography

OpenAIRE

Saitoh, Eiichi; Yamamoto, Shinya; Okamoto, Eishiro; Hayakawa, Yoshimi; Hoshino, Takashi; Sato, Ritsuko; Isemura, Satoko; Ohtsubo, Sadami; Taniguchi, Masayuki

2007-01-01

We have developed a two-dimensional (2D-) gel system of zymography and reverse zymography for the detection and characterization of proteases and protease inhibitors. Isoelectric focusing (IEF) agarose gels with pH gradients were employed for separation in the fi rst-dimension and sodium dodecyl sulfate (SDS)-polyacrylamide gel copolymerized with gelatin used for the second dimension. Proteases and protease inhibitors separated by IEF gel were applied on the second gel without trichloroacetic...

Polymer gel dosimetry

Energy Technology Data Exchange (ETDEWEB)

Baldock, C [Institute of Medical Physics, School of Physics, University of Sydney (Australia); De Deene, Y [Radiotherapy and Nuclear Medicine, Ghent University Hospital (Belgium); Doran, S [CRUK Clinical Magnetic Resonance Research Group, Institute of Cancer Research, Surrey (United Kingdom); Ibbott, G [Radiation Physics, UT M D Anderson Cancer Center, Houston, TX (United States); Jirasek, A [Department of Physics and Astronomy, University of Victoria, Victoria, BC (Canada); Lepage, M [Centre d' imagerie moleculaire de Sherbrooke, Departement de medecine nucleaire et de radiobiologie, Universite de Sherbrooke, Sherbrooke, QC (Canada); McAuley, K B [Department of Chemical Engineering, Queen' s University, Kingston, ON (Canada); Oldham, M [Department of Radiation Oncology, Duke University Medical Center, Durham, NC (United States); Schreiner, L J [Cancer Centre of South Eastern Ontario, Kingston, ON (Canada)], E-mail: c.baldock@physics.usyd.edu.au, E-mail: yves.dedeene@ugent.be

2010-03-07

Polymer gel dosimeters are fabricated from radiation sensitive chemicals which, upon irradiation, polymerize as a function of the absorbed radiation dose. These gel dosimeters, with the capacity to uniquely record the radiation dose distribution in three-dimensions (3D), have specific advantages when compared to one-dimensional dosimeters, such as ion chambers, and two-dimensional dosimeters, such as film. These advantages are particularly significant in dosimetry situations where steep dose gradients exist such as in intensity-modulated radiation therapy (IMRT) and stereotactic radiosurgery. Polymer gel dosimeters also have specific advantages for brachytherapy dosimetry. Potential dosimetry applications include those for low-energy x-rays, high-linear energy transfer (LET) and proton therapy, radionuclide and boron capture neutron therapy dosimetries. These 3D dosimeters are radiologically soft-tissue equivalent with properties that may be modified depending on the application. The 3D radiation dose distribution in polymer gel dosimeters may be imaged using magnetic resonance imaging (MRI), optical-computerized tomography (optical-CT), x-ray CT or ultrasound. The fundamental science underpinning polymer gel dosimetry is reviewed along with the various evaluation techniques. Clinical dosimetry applications of polymer gel dosimetry are also presented. (topical review)

Primary culture of cat intestinal epithelial cells in vitro and the cDNA library construction.

Science.gov (United States)

Zhao, Gui Hua; Liu, Ye; Cheng, Yun Tang; Zhao, Qing Song; Qiu, Xiao; Xu, Chao; Xiao, Ting; Zhu, Song; Liu, Gong Zhen; Yin, Kun

2018-06-26

Felids are the only definitive hosts of Toxoplasma gondii. To lay a foundation for screening the T. gondii-felids interaction factors, we have developed a reproducible primary culture method for cat intestinal epithelial cells (IECs). The primary IECs were isolated from a new born cat's small intestine jejunum region without food ingress, and respectively in vitro cultured by tissue cultivation and combined digestion method with collagenase XI and dispase I, then purified by trypsinization. After identification, the ds cDNA of cat IECs was synthesized for constructing pGADT7 homogenization three-frame plasmid, and transformed into the yeast Y187 for generating the cDNA library. Our results indicated that cultivation of primary cat IECs relays on combined digestion to form polarized and confluent monolayers within 3 days with typical features of normal epithelial cells. The purified cells cultured by digestion method were identified to be nature intestinal epithelial cells using immunohistochemical analysis and were able to maintain viability for at least 15 passages. The homogenizable ds cDNA, which is synthesized from the total RNA extracted from our cultured IECs, distributed among 0.5-2.0 kb, and generated satisfying three-frame cDNA library with the capacity of 1.2 × 106 and the titer of 5.2 × 107 pfu/mL. Our results established an optimal method for the culturing and passage of cat IECs model in vitro, and laid a cDNA library foundation for the subsequent interaction factors screening by yeast two-hybrid.

cDNA cloning and expression analysis of two distinct Sox8 genes in Paramisgurnus dabryanus (Cypriniformes).

Science.gov (United States)

Xia, Xiaohua; Zhao, Jie; Du, Qiyan; Chang, Zhongjie

2010-08-01

The Sox9 gene attracts a lot of attention because of its connection with gonadal development and differentiation. However, Sox8, belonging to the same subgroup SoxE, has rarely been studied. To investigate the function as well as the evolutionary origin of SOXE subgroup, we amplified the genomic DNA of Paramisgurnus dabryanu using a pair of degenerate primers. Using rapid amplification of the cDNA ends (RACE), it was discovered that P. dabryanu has two duplicates: Sox8a and Sox8b. Each has an intron of different length in the conserved HMG-box region. The overall sequence similarity of the deduced amino acid of PdSox8a and PdSox8b was 46.26%, and only two amino acids changed in the HMG-box. This is the first evidence showing that there are two distinct duplications of Sox8 genes in Cypriniformes. Southern blot analysis showed only one hybrid band, with lengths 7.4 or 9.2 kb. Both semi-quantitative RT-PCR and real-time quantitative PCR assay displayed that both PdSox8a and PdSox8b are downregulated during early embryonic development. In adult tissues, the two Sox8 genes expressed ubiquitously, and expression levels are particularly high in the gonads and brain. In gonads, both PdSox8a and PdSox8b are expressed at a higher level in the tesis than in the ovary. PdSox8a and PdSox8b may have functional overlaps and are essential for the neuronal development and differentiation of gonads.

Cloning and expression of cDNA coding for bouganin.

Science.gov (United States)

den Hartog, Marcel T; Lubelli, Chiara; Boon, Louis; Heerkens, Sijmie; Ortiz Buijsse, Antonio P; de Boer, Mark; Stirpe, Fiorenzo

2002-03-01

Bouganin is a ribosome-inactivating protein that recently was isolated from Bougainvillea spectabilis Willd. In this work, the cloning and expression of the cDNA encoding for bouganin is described. From the cDNA, the amino-acid sequence was deduced, which correlated with the primary sequence data obtained by amino-acid sequencing on the native protein. Bouganin is synthesized as a pro-peptide consisting of 305 amino acids, the first 26 of which act as a leader signal while the 29 C-terminal amino acids are cleaved during processing of the molecule. The mature protein consists of 250 amino acids. Using the cDNA sequence encoding the mature protein of 250 amino acids, a recombinant protein was expressed, purified and characterized. The recombinant molecule had similar activity in a cell-free protein synthesis assay and had comparable toxicity on living cells as compared to the isolated native bouganin.

Two kinds of ketoprofen enteric gel beads (CA and CS-SA using biopolymer alginate

Directory of Open Access Journals (Sweden)

Bingchao Cheng

2018-03-01

Full Text Available To obtain expected rapid-release and sustained-release of ketoprofen gel beads, this paper adopted biopolymer alginate to prepare alginate beads and chitosan-alginate gel beads. Formulation factors were investigated and optimized by the single factor test. The release of ketoprofen from calcium alginate gel beads in pH 1.0 hydrochloric acid solution was less than 10% during 2 h, then in pH6.8 was about 95% during 45 min, which met the requirements of rapid-release preparations. However, the drug release of chitosan-alginate gel beads in pH1.0 was less than 5% during 2 h, then in pH6.8 was about 50% during 6 h and reached more than 95% during 12 h, which had a good sustained-release behavior. In addition, the release kinetics of keteprofen from the calcium alginate gel beads fitted well with the Korsmeyer–Peppas model and followed a case-II transport mechanism. However, the release of keteprofen from the chitosan-alginate gel beads exhibited a non-Fickian mechanism and based on the mixed mechanisms of diffusion and polymer relaxation from chitosan-alginate beads. In a word, alginate gel beads of ketoprofen were instant analgesic, while chitosan-alginate gel beads could control the release of ketoprofen during gastro-intestinal tract and prolong the drug's action time. Keywords: Gel beads, Enteric rapid-release, Enteric sustained-release, Ketoprofen

Experimental determination of the diffusion coefficient in two-dimensions in ferrous sulphate gels using the finite element method

International Nuclear Information System (INIS)

Baldock, C.; Harris, P.J.; Piercy, A.R.; Healy, B.

2001-01-01

A novel two-dimensional finite element method for modelling the diffusion which occurs in Fricke or ferrous sulphate type radiation dosimetry gels is presented. In most of the previous work, the diffusion coefficient has been estimated using simple one-dimensional models. This work presents a two-dimensional model which enables the diffusion coefficient to be determined in a much wider range of experimental situations. The model includes the provision for the determination of a drift parameter. To demonstrate the technique comparative diffusion measurements between ferrous sulphate radiation dosimetry gels, with and without xylenol orange chelating agent and carbohydrate additives have been undertaken. Diffusion coefficients of 9.7±0.4, 13.3±0.6 and 9.5±0.8 10-3 cm 2 per h -1 were determined for ferrous sulphate radiation dosimetry gels with and without xylenol orange and with xylenol orange and sucrose additives respectively. Copyright (2001) Australasian College of Physical Scientists and Engineers in Medicine

Construction of cDNA libraries from Pseudocercospora fijiensis Morelet infected leaves of the cultivars Calcutta 4 and Niyarma Yik

Directory of Open Access Journals (Sweden)

Milady Mendoza-Rodríguez

2004-01-01

Full Text Available Molecular studies of plant-pathogen interaction are very important for the identification of gene (s related with the pathogenic process, as well as with the plant resistance. These gene (s could be use for the genetic improvement programs in order to obtain resistant cultivars. The aim of this work was to construct complementary DNA (cDNA libraries from infected leaves with Pseudocercospora fijiensis CCIBP-Pf1 isolated of two banana cultivars (a resistant one Calcutta4 and another one susceptible Niyarma Yik. First-strand cDNA synthesis, was made beginning with one microgram of total RNA by using oligo dT primer and cDNA quality was checked by Polimerase chain reaction (PCR with cytochrome b specific primers. Second-strand cDNA synthesis was performed by using the homopolymeric tailing with dC-BamH I + dT-Not I primer combination. Four cDNA libraries of infected plants at different times of infection with the pathogen were obtained. Forty one clones of one of the libraries of Niyarma Yik were sequenced and the obtained sequences correspond with genes related to fungi. Key words: Banana-Mycosphaerella fijiensis interaction,Black Sigatoka, Musa spp.

Quantitative two-dimensional gel electrophoresis analysis of human fibroblasts transformed by ras oncogenes.

Science.gov (United States)

Miller, M J; Maher, V M; McCormick, J J

1992-11-01

Quantitative two-dimensional gel electrophoresis was used to compare the cellular protein patterns of a normal foreskin-derived human fibroblasts cell line (LG1) and three immortal derivatives of LG1. One derivative, designated MSU-1.1 VO, was selected for its ability to grow in the absence of serum and is non-tumorigenic in athymic mice. The other two strains were selected for focus-formation following transfection with either Ha-ras or N-ras oncogenes and form high grade malignant tumors. Correspondence and cluster analysis provided a nonbiased estimate of the relative similarity of the different two-dimensional patterns. These techniques separated the gel patterns into three distinct classes: LG1, MSU-1.1 VO, and the ras transformed cell strains. The MSU-1.1 VO cells were more closely related to the parental LG1 than to the ras-transformed cells. The differences between the three classes were primarily quantitative in nature: 16% of the spots demonstrated statistically significant changes (P 2) in the rate of incorporation of radioactive amino acids. The patterns from the two ras-transformed cell strains were similar, and variations in the expression of proteins that occurred between the separate experiments obscured consistent differences between the Ha-ras and N-ras transformed cells. However, while only 9 out of 758 spots were classified as different (1%), correspondence analysis could consistently separate the two ras transformants. One of these spots was five times more intense in the Ha-ras transformed cells than the N-ras.(ABSTRACT TRUNCATED AT 250 WORDS)

Statistics as Unbiased Estimators: Exploring the Teaching of Standard Deviation

Science.gov (United States)

Wasserman, Nicholas H.; Casey, Stephanie; Champion, Joe; Huey, Maryann

2017-01-01

This manuscript presents findings from a study about the knowledge for and planned teaching of standard deviation. We investigate how understanding variance as an unbiased (inferential) estimator--not just a descriptive statistic for the variation (spread) in data--is related to teachers' instruction regarding standard deviation, particularly…

Sol-gel transition accelerated by the co-assembly of two components in supramolecular hydrogels.

Science.gov (United States)

Matsumoto, Keigo; Shundo, Atsuomi; Ohno, Masashi; Saruhashi, Kowichiro; Miyachi, Nobuhide; Tsuruzoe, Nobutomo; Tanaka, Keiji

2015-10-28

N-Palmitoyl-Gly-His (PalGH) and glycerol 1-monopalmitate (GMP) in water co-assembled into fibrils with twisted ribbon structures and formed a homogeneous network, resulting in gel formation. Shaking the gel easily broke the fibril network leading to a sol in which high and low fibril density regions exist. After a period at room temperature, the higher density regions became interconnected. The spontaneous sol-gel transition did not take place for a gel made from only PalGH. Also, during the transition, the aggregation state of the co-assembly remained unchanged at a molecular level, unlike the fibril network. Thus, it can be claimed that the sol-gel transition is not associated with the assembled molecular configuration, but with the change in the fibril network. This knowledge might be useful for understanding and controlling sol-gel transition, thereby leading to the design and functionalization of hydrogels.

Internal structure analysis of particle-double network gels used in a gel organ replica

Science.gov (United States)

Abe, Mei; Arai, Masanori; Saito, Azusa; Sakai, Kazuyuki; Kawakami, Masaru; Furukawa, Hidemitsu

2016-04-01

In recent years, the fabrication of patient organ replicas using 3D printers has been attracting a great deal of attention in medical fields. However, the cost of these organ replicas is very high as it is necessary to employ very expensive 3D printers and printing materials. Here we present a new gel organ replica, of human kidney, fabricated with a conventional molding technique, using a particle-double network hydrogel (P-DN gel). The replica is transparent and has the feel of a real kidney. It is expected that gel organ replicas produced this way will be a useful tool for the education of trainee surgeons and clinical ultrasonography technologists. In addition to developing a gel organ replica, the internal structure of the P-DN gel used is also discussed. Because the P-DN gel has a complex structure comprised of two different types of network, it has not been possible to investigate them internally in detail. Gels have an inhomogeneous network structure. If it is able to get a more uniform structure, it is considered that this would lead to higher strength in the gel. In the present study we investigate the structure of P-DN gel, using the gel organ replica. We investigated the internal structure of P-DN gel using Scanning Microscopic Light Scattering (SMILS), a non-contacting and non-destructive.

Building unbiased estimators from non-Gaussian likelihoods with application to shear estimation

International Nuclear Information System (INIS)

Madhavacheril, Mathew S.; Sehgal, Neelima; McDonald, Patrick; Slosar, Anže

2015-01-01

We develop a general framework for generating estimators of a given quantity which are unbiased to a given order in the difference between the true value of the underlying quantity and the fiducial position in theory space around which we expand the likelihood. We apply this formalism to rederive the optimal quadratic estimator and show how the replacement of the second derivative matrix with the Fisher matrix is a generic way of creating an unbiased estimator (assuming choice of the fiducial model is independent of data). Next we apply the approach to estimation of shear lensing, closely following the work of Bernstein and Armstrong (2014). Our first order estimator reduces to their estimator in the limit of zero shear, but it also naturally allows for the case of non-constant shear and the easy calculation of correlation functions or power spectra using standard methods. Both our first-order estimator and Bernstein and Armstrong's estimator exhibit a bias which is quadratic in true shear. Our third-order estimator is, at least in the realm of the toy problem of Bernstein and Armstrong, unbiased to 0.1% in relative shear errors Δg/g for shears up to |g|=0.2

Cu2+-assisted two dimensional charge-mass double focusing gel electrophoresis and mass spectrometric analysis of histone variants.

Science.gov (United States)

Zhang, Wenyang; Tang, Xuemei; Ding, Mengjie; Zhong, Hongying

2014-12-10

Abundant isoforms and dynamic posttranslational modifications cause the separation and identification of histone variants to be experimentally challenging. To meet this need, we employ two-dimensional electrophoretic gel separation followed by mass spectrometric detection which takes advantage of the chelation of Cu(2+) with amino acid residues exposed on the surfaces of the histone proteins. Acid-extracted rat liver histones were first mixed with CuSO4 solution and then separated in one dimension with triton-acid-urea (TAU) gel electrophoresis and in a second dimension using sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The separations result from both the changes in charge and mass upon Cu(2+) chelation. Identities of each separated gel bands were obtained by using matrix-assisted laser desorption-ionization mass spectrometry (MALDI-MS). It was found that the migration of H3 histone isoforms of rat liver is markedly affected by the use of Cu(2+) ions. Copyright © 2014 Elsevier B.V. All rights reserved.

Unbiased free energy estimates in fast nonequilibrium transformations using Gaussian mixtures

International Nuclear Information System (INIS)

Procacci, Piero

2015-01-01

In this paper, we present an improved method for obtaining unbiased estimates of the free energy difference between two thermodynamic states using the work distribution measured in nonequilibrium driven experiments connecting these states. The method is based on the assumption that any observed work distribution is given by a mixture of Gaussian distributions, whose normal components are identical in either direction of the nonequilibrium process, with weights regulated by the Crooks theorem. Using the prototypical example for the driven unfolding/folding of deca-alanine, we show that the predicted behavior of the forward and reverse work distributions, assuming a combination of only two Gaussian components with Crooks derived weights, explains surprisingly well the striking asymmetry in the observed distributions at fast pulling speeds. The proposed methodology opens the way for a perfectly parallel implementation of Jarzynski-based free energy calculations in complex systems

3G vector-primer plasmid for constructing full-length-enriched cDNA libraries.

Science.gov (United States)

Zheng, Dong; Zhou, Yanna; Zhang, Zidong; Li, Zaiyu; Liu, Xuedong

2008-09-01

We designed a 3G vector-primer plasmid for the generation of full-length-enriched complementary DNA (cDNA) libraries. By employing the terminal transferase activity of reverse transcriptase and the modified strand replacement method, this plasmid (assembled with a polydT end and a deoxyguanosine [dG] end) combines priming full-length cDNA strand synthesis and directional cDNA cloning. As a result, the number of steps involved in cDNA library preparation is decreased while simplifying downstream gene manipulation, sequencing, and subcloning. The 3G vector-primer plasmid method yields fully represented plasmid primed libraries that are equivalent to those made by the SMART (switching mechanism at 5' end of RNA transcript) approach.

[Cloning of cDNA for RNA polymerase subunit from the fission yeast Schizosaccharomyces pombe by heterospecific complementation in Saccharomyces cerevisiae].

Science.gov (United States)

Shpakovskiĭ, G V; Lebedenko, E N; Thuriaux, P

1997-02-01

The rpb10 cDNA of the fission yeast Schizosaccharomyces pombe, encoding one of the five small subunits common to all three nuclear DNA-dependent RNA polymerases, was isolated from an expression cDNA library by two independent approaches: PCR-based screening and direct suppression by means of heterospecific complementation of a temperature-sensitive mutant defective in the corresponding gene of Saccharomyces cerevisiae. The cloned Sz. pombe cDNA encodes a protein Rpb10 of 71 amino acids with an M of 8,275 Da, sharing 51 amino acids (71% identity) with the subunit ABC10 beta of RNA polymerases I-III from S. cerevisiae. All eukaryotic members of this protein family have the same general organization featuring two highly conserved motifs (RCFT/SCGK and RYCCRRM) around an atypical zinc finger and an additional invariant HVDLIEK motif toward the C-terminal end. The last motif is only characteristics for homologs from eukaryotes. In keeping with this remarkable structural conservation, the Sz. pombe cDNA also fully complemented a S. cerevisiae deletion mutant lacking subunit ABC10 beta (null allele rpb10-delta 1::HIS3).

Thermotropic nanostructured gels with complex hierarchical structure and two gelling components for water shut-off and enhance of oil recovery

Science.gov (United States)

Altunina, L. K.; Kuvshinov, I. V.; Kuvshinov, V. A.; Kozlov, V. V.; Stasyeva, L. A.

2017-12-01

This work presents the results of laboratory and field tests of thermotropic composition MEGA with two simultaneously acting gelling components, polymer and inorganic. The composition is intended for improving oil recovery and water shut-off at oilfields developed by thermal flooding, and cyclic-steam stimulated oil production wells. The composition forms an in-situ "gel-in-gel" system with improved structural-mechanical properties, using reservoir or carrier fluid heat for gelling. The gel blocks water breakthrough into producing wells and redistribute fluid flows, thus increasing the oil recovery factor.

A putative peroxidase cDNA from turnip and analysis of the encoded protein sequence.

Science.gov (United States)

Romero-Gómez, S; Duarte-Vázquez, M A; García-Almendárez, B E; Mayorga-Martínez, L; Cervantes-Avilés, O; Regalado, C

2008-12-01

A putative peroxidase cDNA was isolated from turnip roots (Brassica napus L. var. purple top white globe) by reverse transcriptase-polymerase chain reaction (RT-PCR) and rapid amplification of cDNA ends (RACE). Total RNA extracted from mature turnip roots was used as a template for RT-PCR, using a degenerated primer designed to amplify the highly conserved distal motif of plant peroxidases. The resulting partial sequence was used to design the rest of the specific primers for 5' and 3' RACE. Two cDNA fragments were purified, sequenced, and aligned with the partial sequence from RT-PCR, and a complete overlapping sequence was obtained and labeled as BbPA (Genbank Accession No. AY423440, named as podC). The full length cDNA is 1167bp long and contains a 1077bp open reading frame (ORF) encoding a 358 deduced amino acid peroxidase polypeptide. The putative peroxidase (BnPA) showed a calculated Mr of 34kDa, and isoelectric point (pI) of 4.5, with no significant identity with other reported turnip peroxidases. Sequence alignment showed that only three peroxidases have a significant identity with BnPA namely AtP29a (84%), and AtPA2 (81%) from Arabidopsis thaliana, and HRPA2 (82%) from horseradish (Armoracia rusticana). Work is in progress to clone this gene into an adequate host to study the specific role and possible biotechnological applications of this alternative peroxidase source.

Purification of Single-Stranded cDNA Based on RNA Degradation Treatment and Adsorption Chromatography.

Science.gov (United States)

Trujillo-Esquivel, Elías; Franco, Bernardo; Flores-Martínez, Alberto; Ponce-Noyola, Patricia; Mora-Montes, Héctor M

2016-08-02

Analysis of gene expression is a common research tool to study networks controlling gene expression, the role of genes with unknown function, and environmentally induced responses of organisms. Most of the analytical tools used to analyze gene expression rely on accurate cDNA synthesis and quantification to obtain reproducible and quantifiable results. Thus far, most commercial kits for isolation and purification of cDNA target double-stranded molecules, which do not accurately represent the abundance of transcripts. In the present report, we provide a simple and fast method to purify single-stranded cDNA, exhibiting high purity and yield. This method is based on the treatment with RNase H and RNase A after cDNA synthesis, followed by separation in silica spin-columns and ethanol precipitation. In addition, our method avoids the use of DNase I to eliminate genomic DNA from RNA preparations, which improves cDNA yield. As a case report, our method proved to be useful in the purification of single-stranded cDNA from the pathogenic fungus Sporothrix schenckii.

cDNA cloning and immunological characterization of the rye grass allergen Lol p I.

Science.gov (United States)

Perez, M; Ishioka, G Y; Walker, L E; Chesnut, R W

1990-09-25

The complete amino acid sequence of two "isoallergenic" forms of Lol p I, the major rye grass (Lolium perenne) pollen allergen, was deduced from cDNA sequence analysis. cDNA clones isolated from a Lolium perenne pollen library contained an open reading frame coding for a 240-amino acid protein. Comparison of the nucleotide and deduced amino acid sequence of two of these clones revealed four changes at the amino acid level and numerous nucleotide differences. Both clones contained one possible asparagine-linked glycosylation site. Northern blot analysis shows one RNA species of 1.2 kilobases. Based on the complete amino acid sequence of Lol p I, overlapping peptides covering the entire molecule were synthesized. Utilizing these peptides we have identified a determinant within the Lol p I molecule that is recognized by human leukocyte antigen class II-restricted T cells obtained from persons allergic to rye grass pollen.

cDNA cloning and nucleotide sequence comparison of Chinese hamster metallothionein I and II mRNAs

Energy Technology Data Exchange (ETDEWEB)

Griffith, B B; Walters, R A; Enger, M D; Hildebrand, C E; Griffith, J K

1983-01-01

Polyadenylated RNA was extracted from a cadmium resistant Chinese hamster (CHO) cell line, enriched for metal-induced, abundant RNA sequences and cloned as double-stranded cDNA in the plasmid pBR322. Two cDNA clones, pCHMT1 and pCHMT2, encoding two Chinese hamster isometallothioneins were identified, and the nucleotide sequence of each insert was determined. The two Chinese hamster metallothioneins show nucleotide sequence homologies of 80% in the protein coding region and approximately 35% in both the 5' and 3' untranslated regions. Interestingly, an 8 nucleotide sequence (TGTAAATA) has been conserved in sequence and position in the 3' untranslated regions of each metallothionein mRNA sequenced thus far. Estimated nucleotide substitution rates derived from interspecies comparisons were used to calculate a metallothionein gene duplication time of 45 to 120 million years ago. 39 references, 1 figure, 1 table.

Construction of full-length cDNA library of white flower Salvia ...

African Journals Online (AJOL)

In order to screen and isolate secondary metabolite biosynthesis related gene, we construct a cDNA library of white flower Salvia miltiorrhiza bge. f.alba. High quality of total RNA was successfully isolated from roots of white flower S. miltiorrhiza using modified CTAB method. Double strand cDNA was cloned into pDNR-LIB ...

Coordinate regulation of stromelysin and collagenase genes determined with cDNA probes

International Nuclear Information System (INIS)

Frisch, S.M.; Clark, E.J.; Werb, Z.

1987-01-01

Secreted proteinases are required for tumor metastasis, angiogenesis, and tissue remodeling during wound healing and embryonic growth. Thus, the regulation of the genes of secreted proteinases may serve as an interesting model for growth-controlled genes in general. The authors studied the genes of the secreted proteinases stromelysin and collagenase by using molecularly cloned cDNAs from each proteinase. Stromelysin cDNA was cloned by differential screening of a total cDNA library from rabbit synovial cells treated with phorbol 12-myristate 13-acetate, which yielded a clone of 1.2 kilobase pairs; collagenase cDNA was obtained by cloning reverse transcripts of anti-collagenase-immunoadsorbed polysomal mRNA, which yielded a clone of 0.8 kilobase pairs. Stromelysin and collagenase mRNA species of 2.2 and 2.4 kilobases, respectively, were detected on hybridization blots of RNA from phorbol 12-myristate 13-acetate-treated but not untreated rabbit synovial cells. Expression of stromelysin mRNA was also induced in rabbit alveolar macrophages and rabbit brain capillary endothelial cells treated with phorbol 12-myristate 13-acetate. Stromelysin and collagenase mRNA were both induced by phorbol 12-myristate 13-acetate and cytochalasin B at a constant ratio of the two gene products; this suggest coordinate regulation. The fact that induction was blocked after inhibition of protein synthesis by cycloheximide implicates an indirect signal transduction pathway that requires new protein synthesis

Two-dimensional gel proteome reference map of human small intestine

Directory of Open Access Journals (Sweden)

Canzonieri Vincenzo

2009-03-01

Full Text Available Abstract Background The small intestine is an important human organ that plays a central role in many physiological functions including digestion, absorption, secretion and defense. Duodenal pathologies include, for instance, the ulcer associated to Helicobacter Pylori infection, adenoma and, in genetically predisposed individuals, celiac disease. Alterations in the bowel reduce its capability to absorb nutrients, minerals and fat-soluble vitamins. Anemia and osteopenia or osteoporosis may develop as a consequence of vitamins malabsorption. Adenoma is a benign tumor that has the potential to become cancerous. Adult celiac disease patients present an overall risk of cancer that is almost twice than that found in the general population. These disease processes are not completely known. To date, a two dimensional (2D reference map of proteins expressed in human duodenal tissue is not yet available: the aim of our study was to characterize the 2D protein map, and to identify proteins of duodenal mucosa of adult individuals without duodenal illness, to create a protein database. This approach, may be useful for comparing similar protein samples in different laboratories and for the molecular characterization of intestinal pathologies without recurring to the use of surgical material. Results The enrolled population comprised five selected samples (3 males and 2 females, aged 19 to 42, taken from 20 adult subjects, on their first visit at the gastroenterology unit for a suspected celiac disease, who did not turn to be affected by any duodenal pathology after gastrointestinal and histological evaluations. Proteins extracted from the five duodenal mucosal specimens were singly separated by 2D gel electrophoresis. After image analysis of each 2D gel, 179 protein spots, representing 145 unique proteins, from 218 spots tested, were successfully identified by MALDI-TOF ms analysis. Normalized volumes, for each protein, have been reported for every gel

Growth hormone and prolactin in Andrias davidianus: cDNA cloning, tissue distribution and phylogenetic analysis.

Science.gov (United States)

Yang, Liping; Meng, Zining; Liu, Yun; Zhang, Yong; Liu, Xiaochun; Lu, Danqi; Huang, Junhai; Lin, Haoran

2010-01-15

The Chinese giant salamander (Andrias davidianus) is one of the largest and 'living fossil' species of amphibian. To obtain genetic information for this species, the cDNAs encoding growth hormone (adGH) and prolactin (adPRL) were cloned from a pituitary cDNA library. The isolated adGH cDNA consisted of 864 bp and encoded a propeptide of 215 amino acids, while the cDNA of adPRL was 1106 bp in length and encoded a putative peptide of 229 amino acids. Expression of the GH and PRL mRNA was only detected in the pituitary. Phylogenetic analyses were performed based on the isolated pituitary hormone sequences using maximum parsimony and neighbor-joining algorithms. The clustering results are similar to that based on the morphological characteristics or the rRNA genes, which indicate that the two orders (Anura and Caudata) of amphibian were monophyletic, and that A. davidianus was diverged early in the Caudate clade. These results indicated that both the GH and PRL sequence might be useful to study the phylogenies of relatively moderate evolved groups.

Agar/gelatin bilayer gel matrix fabricated by simple thermo-responsive sol-gel transition method.

Science.gov (United States)

Wang, Yifeng; Dong, Meng; Guo, Mengmeng; Wang, Xia; Zhou, Jing; Lei, Jian; Guo, Chuanhang; Qin, Chaoran

2017-08-01

We present a simple and environmentally-friendly method to generate an agar/gelatin bilayer gel matrix for further biomedical applications. In this method, the thermally responsive sol-gel transitions of agar and gelatin combined with the different transition temperatures are exquisitely employed to fabricate the agar/gelatin bilayer gel matrix and achieve separate loading for various materials (e.g., drugs, fluorescent materials, and nanoparticles). Importantly, the resulting bilayer gel matrix provides two different biopolymer environments (a polysaccharide environment vs a protein environment) with a well-defined border, which allows the loaded materials in different layers to retain their original properties (e.g., magnetism and fluorescence) and reduce mutual interference. In addition, the loaded materials in the bilayer gel matrix exhibit an interesting release behavior under the control of thermal stimuli. Consequently, the resulting agar/gelatin bilayer gel matrix is a promising candidate for biomedical applications in drug delivery, controlled release, fluorescence labeling, and bio-imaging. Copyright © 2017 Elsevier B.V. All rights reserved.

CDNA cloning, characterization and expression of an endosperm-specific barley peroxidase

DEFF Research Database (Denmark)

Rasmussen, Søren Kjærsgård; Welinder, K.G.; Hejgaard, J.

1991-01-01

A barley peroxidase (BP 1) of pI ca. 8.5 and M(r) 37000 has been purified from mature barley grains. Using antibodies towards peroxidase BP 1, a cDNA clone (pcR7) was isolated from cDNA expression library. The nucleotide sequence of pcR7 gave a derived amino acid sequence identical to the 158 C...

Heterogeneity of rat tropoelastin mRNA revealed by cDNA cloning

International Nuclear Information System (INIS)

Pierce, R.A.; Deak, S.B.; Stolle, C.A.; Boyd, C.D.

1990-01-01

A λgt11 library constructed from poly(A+) RNA isolated from aortic tissue of neonatal rats was screened for rat tropoelastin cDNAs. The first, screen, utilizing a human tropoelastin cDNA clone, provided rat tropoelastin cDNAs spanning 2.3 kb of carboxy-terminal coding sequence and extended into the 3'-untranslated region. A subsequent screen using a 5' rat tropoelastin cDNA clone yielded clones extending into the amino-terminal signal sequence coding region. Sequence analysis of these clones has provided the complete derived amino acid sequence of rat tropoelastin and allowed alignment and comparison with published bovine cDNA sequence. While the overall structure of rat tropoelastin is similar to bovine sequence, numerous substitutions, deletions, and insertions demonstrated considerable heterogeneity between species. In particular, the pentapeptide repeat VPGVG, characteristic of all tropoelastins analyzed to date, is replaced in rat tropoelastin by a repeating pentapeptide, IPGVG. The hexapeptide repeat VGVAPG, the bovine elastin receptor binding peptide, is not encoded by rat tropoelastin cDNAs. Variations in coding sequence between rat tropoelastin CDNA clones were also found which may represent mRNA heterogeneity produced by alternative splicing of the rat tropoelastin pre-mRNA

Polymer gel dosimeter with AQUAJOINT® as hydrogel matrix

Science.gov (United States)

Maeyama, Takuya; Ishida, Yasuhiro; Kudo, Yoshihiro; Fukasaku, Kazuaki; Ishikawa, Kenichi L.; Fukunishi, Nobuhisa

2018-05-01

We report a polymer gel dosimeter based on a new gel matrix (AQUAJOINT®) that is a thermo-irreversible hydrogel formed by mixing two types of water-based liquids at room temperature. Normoxic N-vinylpyrrolidone-based polymer gels were prepared with AQUAJOINT® instead of gelatin. This AQUAJOINT®-based gel dosimeter exhibits a 2.5-fold increase in sensitivity over a gelatin-based gel dosimeter and a linear dose-response in the dose range of 0-8 Gy. This gel has heat resistance in a jar and controlled gel properties such as viscoelastic and mechanical characters, which may be useful for deformable polymer gel dosimetry.

Gel polymer electrolytes for batteries

Science.gov (United States)

Balsara, Nitash Pervez; Eitouni, Hany Basam; Gur, Ilan; Singh, Mohit; Hudson, William

2014-11-18

Nanostructured gel polymer electrolytes that have both high ionic conductivity and high mechanical strength are disclosed. The electrolytes have at least two domains--one domain contains an ionically-conductive gel polymer and the other domain contains a rigid polymer that provides structure for the electrolyte. The domains are formed by block copolymers. The first block provides a polymer matrix that may or may not be conductive on by itself, but that can soak up a liquid electrolyte, thereby making a gel. An exemplary nanostructured gel polymer electrolyte has an ionic conductivity of at least 1.times.10.sup.-4 S cm.sup.-1 at 25.degree. C.

Development of a porcine skeletal muscle cDNA microarray: analysis of differential transcript expression in phenotypically distinct muscles

Directory of Open Access Journals (Sweden)

Stear Michael

2003-03-01

Full Text Available Abstract Background Microarray profiling has the potential to illuminate the molecular processes that govern the phenotypic characteristics of porcine skeletal muscles, such as hypertrophy or atrophy, and the expression of specific fibre types. This information is not only important for understanding basic muscle biology but also provides underpinning knowledge for enhancing the efficiency of livestock production. Results We report on the de novo development of a composite skeletal muscle cDNA microarray, comprising 5500 clones from two developmentally distinct cDNA libraries (longissimus dorsi of a 50-day porcine foetus and the gastrocnemius of a 3-day-old pig. Clones selected for the microarray assembly were of low to moderate abundance, as indicated by colony hybridisation. We profiled the differential expression of genes between the psoas (red muscle and the longissimus dorsi (white muscle, by co-hybridisation of Cy3 and Cy5 labelled cDNA derived from these two muscles. Results from seven microarray slides (replicates correctly identified genes that were expected to be differentially expressed, as well as a number of novel candidate regulatory genes. Quantitative real-time RT-PCR on selected genes was used to confirm the results from the microarray. Conclusion We have developed a porcine skeletal muscle cDNA microarray and have identified a number of candidate genes that could be involved in muscle phenotype determination, including several members of the casein kinase 2 signalling pathway.

Peptidomics combined with cDNA library unravel the diversity of centipede venom

DEFF Research Database (Denmark)

Rong, Mingqiang; Yang, Shilong; Wen, Bo

2015-01-01

of centipede venom. In the present study, we use peptidomics combined with cDNA library to uncover the diversity of centipede Scolopendra subspinipes mutilans L. Koch. 192 peptides were identified by LC-MS/MS and 79 precursors were deduced by cDNA library. Surprisingly, the signal peptides of centipede toxins...

Quantum circuit implementation of cyclic mutually unbiased bases

Energy Technology Data Exchange (ETDEWEB)

Seyfarth, Ulrich; Dittmann, Niklas; Alber, Gernot [Institut fuer Angewandte Physik, Technische Universitaet Darmstadt, 64289 Darmstadt (Germany)

2013-07-01

Complete sets of mutually unbiased bases (MUBs) play an important role in the areas of quantum state tomography and quantum cryptography. Sets which can be generated cyclically may eliminate certain side-channel attacks. To profit from the advantages of these MUBs we propose a method for deriving a quantum circuit that implements the generator of a set into an experimental setup. For some dimensions this circuit is minimal. The presented method is in principle applicable for a larger set of operations and generalizes recently published results.

Characteristic properties of Fibonacci-based mutually unbiased bases

Energy Technology Data Exchange (ETDEWEB)

Seyfarth, Ulrich; Alber, Gernot [Institut fuer Angewandte Physik, Technische Universitaet Darmstadt, 64289 Darmstadt (Germany); Ranade, Kedar [Institut fuer Quantenphysik, Universitaet Ulm, Albert-Einstein-Allee 11, 89069 Ulm (Germany)

2012-07-01

Complete sets of mutually unbiased bases (MUBs) offer interesting applications in quantum information processing ranging from quantum cryptography to quantum state tomography. Different construction schemes provide different perspectives on these bases which are typically also deeply connected to various mathematical research areas. In this talk we discuss characteristic properties resulting from a recently established connection between construction methods for cyclic MUBs and Fibonacci polynomials. As a remarkable fact this connection leads to construction methods which do not involve any relations to mathematical properties of finite fields.

Actuator device utilizing a conductive polymer gel

Science.gov (United States)

Chinn, Douglas A.; Irvin, David J.

2004-02-03

A valve actuator based on a conductive polymer gel is disclosed. A nonconductive housing is provided having two separate chambers separated by a porous frit. The conductive polymer is held in one chamber and an electrolyte solution, used as a source of charged ions, is held in the second chamber. The ends of the housing a sealed with a flexible elastomer. The polymer gel is further provide with electrodes with which to apply an electrical potential across the gel in order to initiate an oxidation reaction which in turn drives anions across the porous frit and into the polymer gel, swelling the volume of the gel and simultaneously contracting the volume of the electrolyte solution. Because the two end chambers are sealed the flexible elastomer expands or contracts with the chamber volume change. By manipulating the potential across the gel the motion of the elastomer can be controlled to act as a "gate" to open or close a fluid channel and thereby control flow through that channel.

Consistent errors in first strand cDNA due to random hexamer mispriming.

Directory of Open Access Journals (Sweden)

Thomas P van Gurp

Full Text Available Priming of random hexamers in cDNA synthesis is known to show sequence bias, but in addition it has been suggested recently that mismatches in random hexamer priming could be a cause of mismatches between the original RNA fragment and observed sequence reads. To explore random hexamer mispriming as a potential source of these errors, we analyzed two independently generated RNA-seq datasets of synthetic ERCC spikes for which the reference is known. First strand cDNA synthesized by random hexamer priming on RNA showed consistent position and nucleotide-specific mismatch errors in the first seven nucleotides. The mismatch errors found in both datasets are consistent in distribution and thermodynamically stable mismatches are more common. This strongly indicates that RNA-DNA mispriming of specific random hexamers causes these errors. Due to their consistency and specificity, mispriming errors can have profound implications for downstream applications if not dealt with properly.

A metrological study of autoradiographs from two-dimensional gel electrophoresis.

Science.gov (United States)

Valiron, O; Lefkovits, I; Garderet, P; Steinberg, C

1984-12-01

Samples prepared from a single batch of labeled cells were subjected to two-dimensional gel electrophoresis and autoradiography. Three factors were varied: total quantity of protein, quantity of labeled protein, and exposure time. The mean background absorbance of the film remained identical (about 0.5 A) for all the treated series, whatever the exposure time and whether or not there were unlabeled proteins in the sample. Hence any spot with a peak A of the same order of magnitude can be seen. The standard deviation was about 0.05 A. Thus, the measurement precision is 2.5% of full scale for digitalization over 0 to 2 A. We derived experimental calibration curves, which are neither linear nor logarithmic because of the film response and which can be used on randomly chosen spots.

Unbiased classification of spatial strategies in the Barnes maze.

Science.gov (United States)

Illouz, Tomer; Madar, Ravit; Clague, Charlotte; Griffioen, Kathleen J; Louzoun, Yoram; Okun, Eitan

2016-11-01

Spatial learning is one of the most widely studied cognitive domains in neuroscience. The Morris water maze and the Barnes maze are the most commonly used techniques to assess spatial learning and memory in rodents. Despite the fact that these tasks are well-validated paradigms for testing spatial learning abilities, manual categorization of performance into behavioral strategies is subject to individual interpretation, and thus to bias. We have previously described an unbiased machine-learning algorithm to classify spatial strategies in the Morris water maze. Here, we offer a support vector machine-based, automated, Barnes-maze unbiased strategy (BUNS) classification algorithm, as well as a cognitive score scale that can be used for memory acquisition, reversal training and probe trials. The BUNS algorithm can greatly benefit Barnes maze users as it provides a standardized method of strategy classification and cognitive scoring scale, which cannot be derived from typical Barnes maze data analysis. Freely available on the web at http://okunlab.wix.com/okunlab as a MATLAB application. eitan.okun@biu.ac.ilSupplementary information: Supplementary data are available at Bioinformatics online. © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Protein composition of wheat gluten polymer fractions determined by quantitative two-dimensional gel electrophoresis and tandem mass spectrometry

Science.gov (United States)

Flour proteins from the US bread wheat Butte 86 were extracted in 0.5% SDS using a two-step procedure with and without sonication and further separated by size exclusion chromatography into monomeric and polymeric fractions. Proteins in each fraction were analyzed by quantitative two-dimensional gel...

SU2 nonstandard bases: the case of mutually unbiased bases

International Nuclear Information System (INIS)

Olivier, Albouy; Kibler, Maurice R.

2007-02-01

This paper deals with bases in a finite-dimensional Hilbert space. Such a space can be realized as a subspace of the representation space of SU 2 corresponding to an irreducible representation of SU 2 . The representation theory of SU 2 is reconsidered via the use of two truncated deformed oscillators. This leads to replace the familiar scheme [j 2 , j z ] by a scheme [j 2 , v ra ], where the two-parameter operator v ra is defined in the universal enveloping algebra of the Lie algebra su 2 . The eigenvectors of the commuting set of operators [j 2 , v ra ] are adapted to a tower of chains SO 3 includes C 2j+1 (2j belongs to N * ), where C 2j+1 is the cyclic group of order 2j + 1. In the case where 2j + 1 is prime, the corresponding eigenvectors generate a complete set of mutually unbiased bases. Some useful relations on generalized quadratic Gauss sums are exposed in three appendices. (authors)

Reverse transcription using random pentadecamer primers increases yield and quality of resulting cDNA

DEFF Research Database (Denmark)

Stangegaard, Michael; Dufva, I.H.; Dufva, Hans Martin

2006-01-01

oligonucleotides (pentadecamers) consistently, yielded at least 2 fold as much cDNA as did random hexamers using either-poly(A) RNA or an amplified version of messenger RNA (aRNA) as a template. The cDNA generated using pentadecamers did not differ in size distribution or the amount of incorporated label compared...... with cDNA generated with random hexamers. The increased efficiency of priming using random pentadecamers resulted in reverse transcription of > 80% of the template aRNA, while random hexamers induced reverse transcription of only 40% of the template aRNA. This suggests a better coverage...... that random pentadecamers can replace random hexamers in reverse transcription reactions on both poly(A) RNA and amplified RNA, resulting in higher cDNA yields and quality....

Sol-gel bonding of silicon wafers

International Nuclear Information System (INIS)

Barbe, C.J.; Cassidy, D.J.; Triani, G.; Latella, B.A.; Mitchell, D.R.G.; Finnie, K.S.; Short, K.; Bartlett, J.R.; Woolfrey, J.L.; Collins, G.A.

2005-01-01

Sol-gel bonds have been produced between smooth, clean silicon substrates by spin-coating solutions containing partially hydrolysed silicon alkoxides. The two coated substrates were assembled and the resulting sandwich fired at temperatures ranging from 60 to 600 deg. C. The sol-gel coatings were characterised using attenuated total reflectance Fourier transform infrared spectroscopy, ellipsometry, and atomic force microscopy, while the corresponding bonded specimens were investigated using scanning electron microscopy and cross-sectional transmission electron microscopy. Mechanical properties were characterised using both microindentation and tensile testing. Bonding of silicon wafers has been successfully achieved at temperatures as low as 60 deg. C. At 300 deg. C, the interfacial fracture energy was 1.55 J/m 2 . At 600 deg. C, sol-gel bonding provided superior interfacial fracture energy over classical hydrophilic bonding (3.4 J/m 2 vs. 1.5 J/m 2 ). The increase in the interfacial fracture energy is related to the increase in film density due to the sintering of the sol-gel interface with increasing temperature. The superior interfacial fracture energy obtained by sol-gel bonding at low temperature is due to the formation of an interfacial layer, which chemically bonds the two sol-gel coatings on each wafer. Application of a tensile stress on the resulting bond leads to fracture of the samples at the silicon/sol-gel interface

3D MR gel dosimetry with lung equivalent gel

International Nuclear Information System (INIS)

Scherer, J.; Solleder, M.; Schiessl, I.; Bogner, L.; Herbst, M.

1998-01-01

The MR gel dosimetry is used to verify complex 3D treatment plans. Till now this method served only for dose evaluation in homogeneous phantoms. On the way to build a heterogeneous anthropomorphic gel phantom, a lung equivalent gel with the density 0.4 g/cm 3 was developed. First experiments show a 1.55 times higher dose reponse in the low density gel (LD gel). The comparison of a dose distribution in a gel/LD gel/gel slab phantom with Monte Carlo calculations shows good agreement within 5%. More over the accuray of the measuring device magnetic resonance imager was studied in respect to the now exclusive digital image processing with the software MRD (MR dosimetry). Because of the dimensions of the Fricke gel phantom an artefact correction, based on the data from the unirradiated phantom proved to be essential. (orig.) [de

Two-dimensional gel electrophoresis data for proteomic profiling of Sporothrix yeast cells

Directory of Open Access Journals (Sweden)

Anderson Messias Rodrigues

2015-03-01

Full Text Available Sporotrichosis is a chronic infection of the skin and subcutaneous tissues of human and other mammals caused by a complex of cryptic dimorphic fungi in the plant-associated order Ophiostomatales. With major differences between routes of transmission, Sporothrix infections are emerging as new threat in tropical and subtropical areas, particularly in form of outbreaks. The mechanisms underlying the pathogenesis and invasion of Sporothrix spp. are still poorly understood and many virulence factors remain unidentified. In this scenario, a global analysis of proteins expressed by clinical Sporothrix species combined with the identification of seroreactive proteins is overdue. Optimization of sample preparation and electrophoresis conditions are key steps toward reproducibility of gel-based proteomics assays. We provide the data generated using an efficient protocol of protein extraction for rapid and large-scale proteome analysis using two-dimensional gel electrophoresis. The protocol was established and optimized for pathogenic and non-pathogenic Sporothrix spp. including Sporothrix brasiliensis (CBS 132990, Sporothrix schenckii sensu stricto (CBS 132974, Sporothrix globosa (CBS 132922, and Sporothrix mexicana (CBS 120341. The data, supplied in this article, are related to the research article entitled “Immunoproteomic analysis reveals a convergent humoral response signature in the Sporothrix schenckii complex” (Rodrigues et al., 2014 [1].

Gel properties and interactions of Mesona blumes polysaccharide-soy protein isolates mixed gel: The effect of salt addition.

Science.gov (United States)

Wang, Wenjie; Shen, Mingyue; Liu, Suchen; Jiang, Lian; Song, Qianqian; Xie, Jianhua

2018-07-15

Effect of different salt ions on the gel properties and microstructure of Mesona blumes polysaccharide (MBP)-soy protein isolates (SPI) mixed gels were investigated. Sodium and calcium ions were chosen to explore their effects on the rheological behavior and gel properties of MBP-SPI mixed gels were evaluated by using rheological, X-ray diffraction, protein solubility determination, and microstructure analysis. Results showed that the addition of salt ions change the crystalline state of gels system, the crystal of gel was enhanced at low ion concentrations (0.005-0.01 M). The two peaks of gel characteristic at 8.9° and 19.9° almost disappeared at high salt ions concentrations (0.015-0.02 M), and new crystallization peaks appeared at around 30° and 45°. The elasticity, viscosity, gel strength, water holding capacity, and thermal stability of gel were increased at low ion concentration. Results showed that the main interactions which promoted gel formation and maintain the three-dimensional structure of the gel were electrostatic interactions, hydrophobic interactions, and disulfide interactions. Copyright © 2018 Elsevier Ltd. All rights reserved.

Encoding mutually unbiased bases in orbital angular momentum for quantum key distribution

CSIR Research Space (South Africa)

Dudley, Angela L

2013-07-01

Full Text Available We encode mutually unbiased bases (MUBs) using the higher-dimensional orbital angular momentum (OAM) degree of freedom associated with optical fields. We illustrate how these states are encoded with the use of a spatial light modulator (SLM). We...

Experimental studies of unbiased gluon jets from $e^{+}e^{-}$ annihilations using the jet boost algorithm

CERN Document Server

Abbiendi, G.; Akesson, P.F.; Alexander, G.; Allison, John; Amaral, P.; Anagnostou, G.; Anderson, K.J.; Arcelli, S.; Asai, S.; Axen, D.; Azuelos, G.; Bailey, I.; Barberio, E.; Barlow, R.J.; Batley, R.J.; Bechtle, P.; Behnke, T.; Bell, Kenneth Watson; Bell, P.J.; Bella, G.; Bellerive, A.; Benelli, G.; Bethke, S.; Biebel, O.; Boeriu, O.; Bock, P.; Boutemeur, M.; Braibant, S.; Brigliadori, L.; Brown, Robert M.; Buesser, K.; Burckhart, H.J.; Campana, S.; Carnegie, R.K.; Caron, B.; Carter, A.A.; Carter, J.R.; Chang, C.Y.; Charlton, David G.; Csilling, A.; Cuffiani, M.; Dado, S.; De Roeck, A.; De Wolf, E.A.; Desch, K.; Dienes, B.; Donkers, M.; Dubbert, J.; Duchovni, E.; Duckeck, G.; Duerdoth, I.P.; Etzion, E.; Fabbri, F.; Feld, L.; Ferrari, P.; Fiedler, F.; Fleck, I.; Ford, M.; Frey, A.; Furtjes, A.; Gagnon, P.; Gary, John William; Gaycken, G.; Geich-Gimbel, C.; Giacomelli, G.; Giacomelli, P.; Giunta, Marina; Goldberg, J.; Gross, E.; Grunhaus, J.; Gruwe, M.; Gunther, P.O.; Gupta, A.; Hajdu, C.; Hamann, M.; Hanson, G.G.; Harder, K.; Harel, A.; Harin-Dirac, M.; Hauschild, M.; Hawkes, C.M.; Hawkings, R.; Hemingway, R.J.; Hensel, C.; Herten, G.; Heuer, R.D.; Hill, J.C.; Hoffman, Kara Dion; Horvath, D.; Igo-Kemenes, P.; Ishii, K.; Jeremie, H.; Jovanovic, P.; Junk, T.R.; Kanaya, N.; Kanzaki, J.; Karapetian, G.; Karlen, D.; Kawagoe, K.; Kawamoto, T.; Keeler, R.K.; Kellogg, R.G.; Kennedy, B.W.; Kim, D.H.; Klein, K.; Klier, A.; Kluth, S.; Kobayashi, T.; Kobel, M.; Komamiya, S.; Kormos, Laura L.; Kramer, T.; Krieger, P.; von Krogh, J.; Kruger, K.; Kuhl, T.; Kupper, M.; Lafferty, G.D.; Landsman, H.; Lanske, D.; Layter, J.G.; Leins, A.; Lellouch, D.; Letts, J.; Levinson, L.; Lillich, J.; Lloyd, S.L.; Loebinger, F.K.; Lu, J.; Ludwig, J.; Macpherson, A.; Mader, W.; Marcellini, S.; Martin, A.J.; Masetti, G.; Mashimo, T.; Mattig, Peter; McDonald, W.J.; McKenna, J.; McMahon, T.J.; McPherson, R.A.; Meijers, F.; Menges, W.; Merritt, F.S.; Mes, H.; Michelini, A.; Mihara, S.; Mikenberg, G.; Miller, D.J.; Moed, S.; Mohr, W.; Mori, T.; Mutter, A.; Nagai, K.; Nakamura, I.; Nanjo, H.; Neal, H.A.; Nisius, R.; O'Neale, S.W.; Oh, A.; Okpara, A.; Oreglia, M.J.; Orito, S.; Pahl, C.; Pasztor, G.; Pater, J.R.; Patrick, G.N.; Pilcher, J.E.; Pinfold, J.; Plane, David E.; Poli, B.; Polok, J.; Pooth, O.; Przybycien, M.; Quadt, A.; Rabbertz, K.; Rembser, C.; Renkel, P.; Rick, H.; Roney, J.M.; Rosati, S.; Rozen, Y.; Runge, K.; Sachs, K.; Saeki, T.; Sarkisyan, E.K.G.; Schaile, A.D.; Schaile, O.; Scharff-Hansen, P.; Schieck, J.; Schoerner-Sadenius, Thomas; Schroder, Matthias; Schumacher, M.; Schwick, C.; Scott, W.G.; Seuster, R.; Shears, T.G.; Shen, B.C.; Sherwood, P.; Siroli, G.; Skuja, A.; Smith, A.M.; Sobie, R.; Soldner-Rembold, S.; Spano, F.; Stahl, A.; Stephens, K.; Strom, David M.; Strohmer, R.; Tarem, S.; Tasevsky, M.; Taylor, R.J.; Teuscher, R.; Thomson, M.A.; Torrence, E.; Toya, D.; Tran, P.; Trigger, I.; Trocsanyi, Z.; Tsur, E.; Turner-Watson, M.F.; Ueda, I.; Ujvari, B.; Vollmer, C.F.; Vannerem, P.; Vertesi, R.; Verzocchi, M.; Voss, H.; Vossebeld, J.; Waller, D.; Ward, C.P.; Ward, D.R.; Warsinsky, M.; Watkins, P.M.; Watson, A.T.; Watson, N.K.; Wells, P.S.; Wengler, T.; Wermes, N.; Wetterling, D.; Wilson, G.W.; Wilson, J.A.; Wolf, G.; Wyatt, T.R.; Yamashita, S.; Zer-Zion, D.; Zivkovic, Lidija

2004-01-01

We present the first experimental results based on the jet boost algorithm, a technique to select unbiased samples of gluon jets in e+e- annihilations, i.e. gluon jets free of biases introduced by event selection or jet finding criteria. Our results are derived from hadronic Z0 decays observed with the OPAL detector at the LEP e+e- collider at CERN. First, we test the boost algorithm through studies with Herwig Monte Carlo events and find that it provides accurate measurements of the charged particle multiplicity distributions of unbiased gluon jets for jet energies larger than about 5 GeV, and of the jet particle energy spectra (fragmentation functions) for jet energies larger than about 14 GeV. Second, we apply the boost algorithm to our data to derive unbiased measurements of the gluon jet multiplicity distribution for energies between about 5 and 18 GeV, and of the gluon jet fragmentation function at 14 and 18 GeV. In conjunction with our earlier results at 40 GeV, we then test QCD calculations for the en...

Antibiotic-containing hyaluronic acid gel as an antibacterial carrier: Usefulness of sponge and film-formed HA gel in deep infection.

Science.gov (United States)

Matsuno, Hiroaki; Yudoh, Kazuo; Hashimoto, Masamichi; Himeda, Yasukazu; Miyoshi, Teruzo; Yoshida, Kaoru; Kano, Syogo

2006-03-01

We have developed a novel bioabsorbable antibacterial carrier using hyaluronic acid (HA) gel for prevention and treatment of orthopedic infections. In this study, we investigated the in vivo antibacterial effects of two forms of this new material, an HA gel sponge and an HA gel film. A titanium cylinder was inserted into the intramedullary cavity of each rabbit femur, along with an HA gel sponge or HA gel film containing antibiotics. The HA gel sponge contained gentamycin, vancomycin, tobramycin, or minomycin. The HA gel film contained gentamycin or vancomycin. After 0, 7, and 14 days, the rabbit bone marrow was collected, and the antibacterial activity of the HA gel was determined by agar diffusion test. As a control, we used Septocoll, a commercially available antibacterial carrier. Both the HA gel sponge and HA gel film exhibited antibacterial activity. The present results indicate that HA gel containing antibiotics is a clinically useful bioabsorbable antibacterial carrier. Copyright 2006 Orthopaedic Research Society.

Human cDNA clones for an α subunit of G/sub i/ signal-transduction protein

International Nuclear Information System (INIS)

Bray, P.; Carter, A.; Guo, V.; Puckett, C.; Kamholz, J.; Spiegel, A.; Nirenberg, M.

1987-01-01

Two cDNA clones were obtained from a λgt11 cDNA human brain library that correspond to α/sub i/ subunits of G signal-transduction proteins (where α/sub i/ subunits refer to the α subunits of G proteins that inhibit adenylate cyclase). The nucleotide sequence of human brain α/sub i/ is highly homologous to that of bovine brain α/sub i/ and the predicted amino acid sequences are identical. However, human and bovine brain α/sub i/ cDNAs differ significantly from α/sub i/ cDNAs from human monocytes, rat glioma, and mouse macrophages in amino acid (88% homology) and nucleotide (71-75% homology) sequences. In addition, the nucleotide sequences of the 3' untranslated regions of human and bovine brain α/sub i/ cDNAs differ markedly from the sequences of human monocyte, rat glioma, and mouse macrophage α/sub i/ cDNAs. These results suggest there are at least two classes of α/sub i/ mRNA

The nucleotide sequence of human transition protein 1 cDNA

Energy Technology Data Exchange (ETDEWEB)

Luerssen, H; Hoyer-Fender, S; Engel, W [Universitaet Goettingen (West Germany)

1988-08-11

The authors have screened a human testis cDNA library with an oligonucleotide of 81 mer prepared according to a part of the published nucleotide sequence of the rat transition protein TP 1. They have isolated a cDNA clone with the length of 441 bp containing the coding region of 162 bp for human transition protein 1. There is about 84% homology in the coding region of the sequence compared to rat. The human cDNA-clone encodes a polypeptide of 54 amino acids of which 7 are different to that of rat.

Unbiased estimators for spatial distribution functions of classical fluids

Science.gov (United States)

Adib, Artur B.; Jarzynski, Christopher

2005-01-01

We use a statistical-mechanical identity closely related to the familiar virial theorem, to derive unbiased estimators for spatial distribution functions of classical fluids. In particular, we obtain estimators for both the fluid density ρ(r) in the vicinity of a fixed solute and the pair correlation g(r) of a homogeneous classical fluid. We illustrate the utility of our estimators with numerical examples, which reveal advantages over traditional histogram-based methods of computing such distributions.

Unbiased stereological methods used for the quantitative evaluation of guided bone regeneration

DEFF Research Database (Denmark)

Aaboe, Else Merete; Pinholt, E M; Schou, S

1998-01-01

The present study describes the use of unbiased stereological methods for the quantitative evaluation of the amount of regenerated bone. Using the principle of guided bone regeneration the amount of regenerated bone after placement of degradable or non-degradable membranes covering defects...

High-throughput screening of suppression subtractive hybridization cDNA libraries using DNA microarray analysis

CSIR Research Space (South Africa)

Van den Berg, N

2004-11-01

Full Text Available Efficient construction of cDNA libraries enriched for differentially expressed transcripts is an important first step in many biological investigations. We present a quantitative procedure for screening cDNA libraries constructed by suppression...

Construction and identification of subtracted cDNA library in bone marrow cells of radon-exposed mice

International Nuclear Information System (INIS)

Li Jianxiang; Nie Jihua; Tong Jian; Fu Chunling; Zhou Jianwei

2008-01-01

Objective: To construct and identify subtracted cDNA library in bone marrow cells of mice exposed to radon inhalation. Methods: Adult male BALB/c mice, weighing 18-22 g, were placed in a multi- functional radon chamber. One group of mice was exposed to radon up to the accumulative dose of 105 work level month (WLM). The control group of mice was housed in a room with an accumulative dose of 1 WLM. To construct a subtracted cDNA library enriched with differentially expressed genes, the SMART technique and the suppression subtractive hybridization were performed. The obtained forward and reverse cDNA fragments were directly inserted into pMD18-T vector and transformed into E. coli JM109. The inserting cDNA fragments were screened by the blue-and-white blot screening and nested PCR of bacterium liquid. Results: The 244 of 285 white bacteria clones obtained randomly were positive clones contained 100-1100 bp inserted cDNA fragments. Conclusions: The forward and reverse subtracted cDNA library in bone marrow cells of mice exposed to radon inhalation is successfully constructed. (authors)

Effect of acidity of in-office bleaching gels on tooth sensitivity and whitening: a two-center double-blind randomized clinical trial.

Science.gov (United States)

Loguercio, A D; Servat, F; Stanislawczuk, R; Mena-Serrano, A; Rezende, M; Prieto, M V; Cereño, V; Rojas, M F; Ortega, K; Fernandez, E; Reis, A

2017-12-01

The study aimed to compare the tooth sensitivity (TS) and bleaching efficacy of two hydrogen peroxide gels with different pHs (acid pH [Pola Office, SDI] and the neutral pH [Pola Office+, SDI]) used for in-office bleaching. Fifty-four patients from Brazil and Chile, with right superior incisor darker than A2, were selected for this double-blind, split-mouth randomized trial. Teeth were bleached in two sessions, with 1-week interval. Each session had three applications of 8 min each, according to the manufacturer's instructions. The color changes were evaluated by subjective (Vita Classical and Vita Bleachedguide) and objective (Easy shade spectrophotometer) methods. Participants recorded TS with 0-10 visual analog scale. Color change in shade guide units (SGU) and ΔE was analyzed by Student's t test (α = 0.05). The absolute risk and intensity of TS were evaluated by McNemar's test and Wilcoxon-paired test, respectively (α = 0.05). All groups achieved the same level of whitening after 30 days of clinical evaluation. The use of a neutral in-office bleaching gel significantly decreases the absolute risk of TS (28%, 95% CI 18-41) and intensity of TS when compared to the acid bleaching gel (absolute risk of 50%, 95% CI 37-63). The use of a neutral in-office bleaching agent gel produced the same whitening degree than an acid bleaching gel but with reduced risk and intensity of tooth sensitivity. Clinicians should opt to use in-office bleaching with a neutral gel than an acid product because the former causes a significant lower risk and intensity of tooth sensitivity.

Comparison effect of azithromycin gel 2% with clindamycin gel 1% in patients with acne.

Science.gov (United States)

Mokhtari, Fatemeh; Faghihi, Gita; Basiri, Akram; Farhadi, Sadaf; Nilforoushzadeh, Mohammadali; Behfar, Shadi

2016-01-01

Acne vulgaris is the most common skin disease. Local and systemic antimicrobial drugs are used for its treatment. But increasing resistance of Propionibacterium acnes to antibiotics has been reported. In a double-blind clinical trial, 40 patients with mild to moderate acne vulgaris were recruited. one side of the face was treated with Clindamycin Gel 1% and the other side with Azithromycin Topical Gel 2% BID for 8 weeks and then they were assessed. Average age was 21. 8 ± 7 years. 82.5% of them were female. Average number of papules, pustules and comedones was similarly reduced in both groups and, no significant difference was observed between the two groups (P > 0.05, repeated measurs ANOVA). The mean indexes of ASI and TLC also significantly decreased during treatment in both groups, no significant difference was observed between the two groups. (P > 0.05, repeated measurs ANOVA). Also, impact of both drugs on papules and pustules was 2-3 times greater than the effect on comedones. Average satisfaction score was not significant between the two groups (P = 0.6, repeated measurs ANOVA). finally, frequency distribution of complications was not significant between the two groups (P > 0.05, Fisher Exact test). Azithromycin gel has medical impact at least similar to Clindamycin Gel in treatment of mild to moderate acne vulgaris, and it may be consider as suitable drug for resistant acne to conventional topical therapy.

The cDNA sequence of a neutral horseradish peroxidase.

Science.gov (United States)

Bartonek-Roxå, E; Eriksson, H; Mattiasson, B

1991-02-16

A cDNA clone encoding a horseradish (Armoracia rusticana) peroxidase has been isolated and characterized. The cDNA contains 1378 nucleotides excluding the poly(A) tail and the deduced protein contains 327 amino acids which includes a 28 amino acid leader sequence. The predicted amino acid sequence is nine amino acids shorter than the major isoenzyme belonging to the horseradish peroxidase C group (HRP-C) and the sequence shows 53.7% identity with this isoenzyme. The described clone encodes nine cysteines of which eight correspond well with the cysteines found in HRP-C. Five potential N-glycosylation sites with the general sequence Asn-X-Thr/Ser are present in the deduced sequence. Compared to the earlier described HRP-C this is three glycosylation sites less. The shorter sequence and fewer N-glycosylation sites give the native isoenzyme a molecular weight of several thousands less than the horseradish peroxidase C isoenzymes. Comparison with the net charge value of HRP-C indicates that the described cDNA clone encodes a peroxidase which has either the same or a slightly less basic pI value, depending on whether the encoded protein is N-terminally blocked or not. This excludes the possibility that HRP-n could belong to either the HRP-A, -D or -E groups. The low sequence identity (53.7%) with HRP-C indicates that the described clone does not belong to the HRP-C isoenzyme group and comparison of the total amino acid composition with the HRP-B group does not place the described clone within this isoenzyme group. Our conclusion is that the described cDNA clone encodes a neutral horseradish peroxidase which belongs to a new, not earlier described, horseradish peroxidase group.

cDNA cloning of rat and human medium chain acyl-CoA dehydrogenase (MCAD)

International Nuclear Information System (INIS)

Matsubara, Y.; Kraus, J.P.; Rosenberg, L.E.; Tanaka, K.

1986-01-01

MCAD is one of three mitochondrial flavoenzymes which catalyze the first step in the β-oxidation of straight chain fatty acids. It is a tetramer with a subunit Mr of 45 kDa. MCAD is synthesized in the cytosol as a 49 kDa precursor polypeptide (pMCAD), imported into mitochondria, and cleaved to the mature form. Genetic deficiency of MCAD causes recurrent episodes of hypoglycemic coma accompanied by medium chain dicarboxylic aciduria. Employing a novel approach, the authors now report isolation of partial rat and human cDNA clones encoding pMCAD. mRNA encoding pMCAD was purified to near homogeneity by polysome immunoadsorption using polyclonal monospecific antibody. Single-stranded [ 32 P]labeled cDNA probe was synthesized using the enriched mRNA as template, and was used to screen directly 16,000 colonies from a total rat liver cDNA library constructed in pBR322. One clone (600 bp) was detected by in situ hybridization. Hybrid-selected translation with this cDNA yielded a 49 kDa polypeptide indistinguishable in size from rat pMCAD and immunoprecipitable with anti-MCAD antibody. Using the rat cDNA as probe, 43,000 colonies from a human liver cDNA library were screened. Four identical positive clones (400 bp) were isolated and positively identified by hybrid-selected translation and immunoprecipitation. The sizes of rat and human mRNAs encoding pMCAD were 2.2 kb and 2.4 kb, respectively, as determined by Northern blotting

Unbiased Strain-Typing of Arbovirus Directly from Mosquitoes Using Nanopore Sequencing: A Field-forward Biosurveillance Protocol.

Science.gov (United States)

Russell, Joseph A; Campos, Brittany; Stone, Jennifer; Blosser, Erik M; Burkett-Cadena, Nathan; Jacobs, Jonathan L

2018-04-03

The future of infectious disease surveillance and outbreak response is trending towards smaller hand-held solutions for point-of-need pathogen detection. Here, samples of Culex cedecei mosquitoes collected in Southern Florida, USA were tested for Venezuelan Equine Encephalitis Virus (VEEV), a previously-weaponized arthropod-borne RNA-virus capable of causing acute and fatal encephalitis in animal and human hosts. A single 20-mosquito pool tested positive for VEEV by quantitative reverse transcription polymerase chain reaction (RT-qPCR) on the Biomeme two3. The virus-positive sample was subjected to unbiased metatranscriptome sequencing on the Oxford Nanopore MinION and shown to contain Everglades Virus (EVEV), an alphavirus in the VEEV serocomplex. Our results demonstrate, for the first time, the use of unbiased sequence-based detection and subtyping of a high-consequence biothreat pathogen directly from an environmental sample using field-forward protocols. The development and validation of methods designed for field-based diagnostic metagenomics and pathogen discovery, such as those suitable for use in mobile "pocket laboratories", will address a growing demand for public health teams to carry out their mission where it is most urgent: at the point-of-need.

Efficacy Testing of H56 cDNA Tattoo Immunization against Tuberculosis in a Mouse Model.

Science.gov (United States)

Platteel, Anouk C M; Nieuwenhuizen, Natalie E; Domaszewska, Teresa; Schürer, Stefanie; Zedler, Ulrike; Brinkmann, Volker; Sijts, Alice J A M; Kaufmann, Stefan H E

2017-01-01

Tuberculosis (TB), caused by Mycobacterium tuberculosis ( Mtb ), remains a global threat. The only approved vaccine against TB, Mycobacterium bovis bacillus Calmette-Guérin (BCG), provides insufficient protection and, being a live vaccine, can cause disseminated disease in immunocompromised individuals. Previously, we found that intradermal cDNA tattoo immunization with cDNA of tetanus toxoid fragment C domain 1 fused to cDNA of the fusion protein H56, comprising the Mtb antigens Ag85B, ESAT-6, and Rv2660c, induced antigen-specific CD8 + T cell responses in vivo . As cDNA tattoo immunization would be safer than a live vaccine in immunocompromised patients, we tested the protective efficacy of intradermal tattoo immunization against TB with H56 cDNA, as well as with H56_E, a construct optimized for epitope processing in a mouse model. As Mtb antigens can be used in combination with BCG to boost immune responses, we also tested the protective efficacy of heterologous prime-boost, using dermal tattoo immunization with H56_E cDNA to boost BCG immunization in mice. Dermal H56 and H56_E cDNA immunization induced H56-specific CD4 + and CD8 + T cell responses and Ag85B-specific IgG antibodies, but did not reduce bacterial loads, although immunization with H56_E ameliorated lung pathology. Both subcutaneous and intradermal immunization with BCG resulted in broad cellular immune responses, with increased frequencies of CD4 + T effector memory cells, T follicular helper cells, and germinal center B cells, and resulted in reduced bacterial loads and lung pathology. Heterologous vaccination with BCG/H56_E cDNA induced increased H56-specific CD4 + and CD8 + T cell cytokine responses compared to vaccination with BCG alone, and lung pathology was significantly decreased in BCG/H56_E cDNA immunized mice compared to unvaccinated controls. However, bacterial loads were not decreased after heterologous vaccination compared to BCG alone. CD4 + T cells responding to Ag85B- and ESAT-6

Transdermal delivery of paeonol using cubic gel and microemulsion gel

Science.gov (United States)

Luo, Maofu; Shen, Qi; Chen, Jinjin

2011-01-01

Background The aim of this study was to develop new systems for transdermal delivery of paeonol, in particular microemulsion gel and cubic gel formulations. Methods Various microemulsion vehicles were prepared using isopropyl myristate as an oil phase, polyoxyethylated castor oil (Cremophor® EL) as a surfactant, and polyethylene glycol 400 as a cosurfactant. In the optimum microemulsion gel formulation, carbomer 940 was selected as the gel matrix, and consisted of 1% paeonol, 4% isopropyl myristate, 28% Cremophor EL/polyethylene glycol 400 (1:1), and 67% water. The cubic gel was prepared containing 3% paeonol, 30% water, and 67% glyceryl monooleate. Results A skin permeability test using excised rat skins indicated that both the cubic gel and microemulsion gel formulations had higher permeability than did the paeonol solution. An in vivo pharmacokinetic study done in rats showed that the relative bioavailability of the cubic gel and microemulsion gel was enhanced by about 1.51-fold and 1.28-fold, respectively, compared with orally administered paeonol suspension. Conclusion Both the cubic gel and microemulsion gel formulations are promising delivery systems to enhance the skin permeability of paeonol, in particular the cubic gel. PMID:21904450

Antimicrobial efficacy of alcohol-based hand gels.

Science.gov (United States)

Guilhermetti, M; Marques Wiirzler, L A; Castanheira Facio, B; da Silva Furlan, M; Campo Meschial, W; Bronharo Tognim, M C; Botelho Garcia, L; Luiz Cardoso, C

2010-03-01

In recent years, several commercial alcohol-based hand gels have appeared on the market to improve the hand-cleansing compliance of healthcare workers. Although the antimicrobial efficacy of these products has been reported in different countries, few studies have investigated this subject in Brazil. In this study, we assessed the antimicrobial efficacy of 12 alcohol-based hand gels produced in Brazil, containing 70% w/w or v/v ethyl alcohol as the active ingredient, according to the European Standard EN 1500 (EN 1500). The following alcohol gels were tested: Hand Gel, Voga Gel, Solumax Solugel, Doctor Clean, Rio Gel, Clear Gel, Sevengel, Hand CHC, Gel Bac, WBL-50 Gel, Sanigel and Soft Care Gel. In addition, 70% w/w ethyl alcohol and three alcohol-based hand rubs (Sterillium, Sterillium Gel, and Spitaderm), commonly used in Europe and effective according to EN 1500, were also tested. All the products tested, except for two, were approved by the EN 1500 test protocol with a 60s application. The results confirmed the antimicrobial efficacy of the majority of the alcohol gels produced in Brazil for hand hygiene of healthcare workers. Copyright 2009 The Hospital Infection Society. Published by Elsevier Ltd. All rights reserved.

Personalized recommendation based on unbiased consistence

Science.gov (United States)

Zhu, Xuzhen; Tian, Hui; Zhang, Ping; Hu, Zheng; Zhou, Tao

2015-08-01

Recently, in physical dynamics, mass-diffusion-based recommendation algorithms on bipartite network provide an efficient solution by automatically pushing possible relevant items to users according to their past preferences. However, traditional mass-diffusion-based algorithms just focus on unidirectional mass diffusion from objects having been collected to those which should be recommended, resulting in a biased causal similarity estimation and not-so-good performance. In this letter, we argue that in many cases, a user's interests are stable, and thus bidirectional mass diffusion abilities, no matter originated from objects having been collected or from those which should be recommended, should be consistently powerful, showing unbiased consistence. We further propose a consistence-based mass diffusion algorithm via bidirectional diffusion against biased causality, outperforming the state-of-the-art recommendation algorithms in disparate real data sets, including Netflix, MovieLens, Amazon and Rate Your Music.

Radioactive cDNA microarray in neurospsychiatry

International Nuclear Information System (INIS)

Choe, Jae Gol; Shin, Kyung Ho; Lee, Min Soo; Kim, Meyoung Kon

2003-01-01

Microarray technology allows the simultaneous analysis of gene expression patterns of thousands of genes, in a systematic fashion, under a similar set of experimental conditions, thus making the data highly comparable. In some cases arrays are used simply as a primary screen leading to downstream molecular characterization of individual gene candidates. In other cases, the goal of expression profiling is to begin to identify complex regulatory networks underlying developmental processes and disease states. Microarrays were originally used with cell lines or other simple model systems. More recently, microarrays have been used in the analysis of more complex biological tissues including neural systems and the brain. The application of cDNA arrays in neuropsychiatry has lagged behind other fields for a number of reasons. These include a requirement for a large amount of input probe RNA in fluorescent-glass based array systems and the cellular complexity introduced by multicellular brain and neural tissues. An additional factor that impacts the general use of microarrays in neuropsychiatry is the lack of availability of sequenced clone sets from model systems. While human cDNA clones have been widely available, high quality rat, mouse, and drosophilae, among others are just becoming widely available. A final factor in the application of cDNA microarrays in neuropsychiatry is cost of commercial arrays. As academic microarray facilitates become more commonplace custom made arrays will become more widely available at a lower cost allowing more widespread applications. In summary, microarray technology is rapidly having an impact on many areas of biomedical research. Radioisotope-nylon based microarrays offer alternatives that may in some cases be more sensitive, flexible, inexpensive, and universal as compared to other array formats, such as fluorescent-glass arrays. In some situations of limited RNA or exotic species, radioactive membrane microarrays may be the most

Radioactive cDNA microarray in neurospsychiatry

Energy Technology Data Exchange (ETDEWEB)

Choe, Jae Gol; Shin, Kyung Ho; Lee, Min Soo; Kim, Meyoung Kon [Korea University Medical School, Seoul (Korea, Republic of)

2003-02-01

Microarray technology allows the simultaneous analysis of gene expression patterns of thousands of genes, in a systematic fashion, under a similar set of experimental conditions, thus making the data highly comparable. In some cases arrays are used simply as a primary screen leading to downstream molecular characterization of individual gene candidates. In other cases, the goal of expression profiling is to begin to identify complex regulatory networks underlying developmental processes and disease states. Microarrays were originally used with cell lines or other simple model systems. More recently, microarrays have been used in the analysis of more complex biological tissues including neural systems and the brain. The application of cDNA arrays in neuropsychiatry has lagged behind other fields for a number of reasons. These include a requirement for a large amount of input probe RNA in fluorescent-glass based array systems and the cellular complexity introduced by multicellular brain and neural tissues. An additional factor that impacts the general use of microarrays in neuropsychiatry is the lack of availability of sequenced clone sets from model systems. While human cDNA clones have been widely available, high quality rat, mouse, and drosophilae, among others are just becoming widely available. A final factor in the application of cDNA microarrays in neuropsychiatry is cost of commercial arrays. As academic microarray facilitates become more commonplace custom made arrays will become more widely available at a lower cost allowing more widespread applications. In summary, microarray technology is rapidly having an impact on many areas of biomedical research. Radioisotope-nylon based microarrays offer alternatives that may in some cases be more sensitive, flexible, inexpensive, and universal as compared to other array formats, such as fluorescent-glass arrays. In some situations of limited RNA or exotic species, radioactive membrane microarrays may be the most

A Potato cDNA Encoding a Homologue of Mammalian Multidrug Resistant P-Glycoprotein

Science.gov (United States)

Wang, W.; Takezawa, D.; Poovaiah, B. W.

1996-01-01

A homologue of the multidrug resistance (MDR) gene was obtained while screening a potato stolon tip cDNA expression library with S-15-labeled calmodulin. The mammalian MDR gene codes for a membrane-bound P-glycoprotein (170-180 kDa) which imparts multidrug resistance to cancerous cells. The potato cDNA (PMDR1) codes for a polypeptide of 1313 amino acid residues (ca. 144 kDa) and its structural features are very similar to the MDR P-glycoprotein. The N-terminal half of the PMDR1-encoded protein shares striking homology with its C-terminal half, and each half contains a conserved ATP-binding site and six putative transmembrane domains. Southern blot analysis indicated that potato has one or two MDR-like genes. PMDR1 mRNA is constitutively expressed in all organs studied with higher expression in the stem and stolon tip. The PMDR1 expression was highest during tuber initiation and decreased during tuber development.

Human tissue factor: cDNA sequence and chromosome localization of the gene

International Nuclear Information System (INIS)

Scarpati, E.M.; Wen, D.; Broze, G.J. Jr.; Miletich, J.P.; Flandermeyer, R.R.; Siegel, N.R.; Sadler, J.E.

1987-01-01

A human placenta cDNA library in λgt11 was screened for the expression of tissue factor antigens with rabbit polyclonal anti-human tissue factor immunoglobulin G. Among 4 million recombinant clones screened, one positive, λHTF8, expressed a protein that shared epitopes with authentic human brain tissue factor. The 1.1-kilobase cDNA insert of λHTF8 encoded a peptide that contained the amino-terminal protein sequence of human brain tissue factor. Northern blotting identified a major mRNA species of 2.2 kilobases and a minor species of ∼ 3.2 kilobases in poly(A) + RNA of placenta. Only 2.2-kilobase mRNA was detected in human brain and in the human monocytic U937 cell line. In U937 cells, the quantity of tissue factor mRNA was increased several fold by exposure of the cells to phorbol 12-myristate 13-acetate. Additional cDNA clones were selected by hybridization with the cDNA insert of λHTF8. These overlapping isolates span 2177 base pairs of the tissue factor cDNA sequence that includes a 5'-noncoding region of 75 base pairs, an open reading frame of 885 base pairs, a stop codon, a 3'-noncoding region of 1141 base pairs, and a poly(a) tail. The open reading frame encodes a 33-kilodalton protein of 295 amino acids. The predicted sequence includes a signal peptide of 32 or 34 amino acids, a probable extracellular factor VII binding domain of 217 or 219 amino acids, a transmembrane segment of 23 acids, and a cytoplasmic tail of 21 amino acids. There are three potential glycosylation sites with the sequence Asn-X-Thr/Ser. The 3'-noncoding region contains an inverted Alu family repetitive sequence. The tissue factor gene was localized to chromosome 1 by hybridization of the cDNA insert of λHTF8 to flow-sorted human chromosomes

Performance comparison of a silica gel-water and activated carbon-methanol two beds adsorption chillers

Directory of Open Access Journals (Sweden)

Szelągowski Adam

2017-01-01

Full Text Available The aim of the study is to compare the efficiency of adsorption refrigerating equipment working with different working pairs. Adsorption cooling devices can operate with a relatively low temperature of heat sources while consuming only a small amount of electricity for the operation of auxiliary equipment. Refrigerants used in adsorption devices are substances that do not have a negative impact on the environment. All that makes that adsorption refrigeration seems to be a good solution for utilizing renewable and waste heat sources for cold production. To carry out the experiment the adsorption cooling device has been developed and researched in Institute of Heat Engineering at Warsaw University of Technology. The test bench consisted of two cylindrical adsorbers, condenser, evaporator, oil heater and two oil coolers. In order to perform the correct action it has been developed and implemented special control algorithm device, allowed to keep the temperature in the evaporator at a preset level. The unit tested for two sorption pairs: activated carbon – methanol, and silica gel – water. For activated carbon - methanol working pair it was obtained energy efficiency rating (EER equals to 0.14 and specific cooling power (SPC of 16 W/kg. For silica gel - water EER of refrigeration unit was 0.25 and SPC was equal to 208 W/kg.

A BIOINFORMATIC STRATEGY TO RAPIDLY CHARACTERIZE CDNA LIBRARIES

Science.gov (United States)

A Bioinformatic Strategy to Rapidly Characterize cDNA LibrariesG. Charles Ostermeier1, David J. Dix2 and Stephen A. Krawetz1.1Departments of Obstetrics and Gynecology, Center for Molecular Medicine and Genetics, & Institute for Scientific Computing, Wayne State Univer...

Fundamentals of Polymer Gel Dosimeters

Science.gov (United States)

McAuley, Kim B.

2006-12-01

The recent literature on polymer gel dosimetry contains application papers and basic experimental studies involving polymethacrylic-acid-based and polyacrylamide-based gel dosimeters. The basic studies assess the relative merits of these two most commonly used dosimeters, and explore the effects of tetrakis hydroxymethyl phosphonium chloride (THPC) antioxidant on dosimeter performance. Polymer gel dosimeters that contain THPC or other oxygen scavengers are called normoxic dosimeters, because they can be prepared under normal atmospheric conditions, rather than in a glove box that excludes oxygen. In this review, an effort is made to explain some of the underlying chemical phenomena that affect dosimeter performance using THPC, and that lead to differences in behaviour between dosimeters made using the two types of monomer systems. Progress on the development of new more effective and less toxic dosimeters is also reported.

Mutually unbiased bases and trinary operator sets for N qutrits

International Nuclear Information System (INIS)

Lawrence, Jay

2004-01-01

A compete orthonormal basis of N-qutrit unitary operators drawn from the Pauli group consists of the identity and 9 N -1 traceless operators. The traceless ones partition into 3 N +1 maximally commuting subsets (MCS's) of 3 N -1 operators each, whose joint eigenbases are mutually unbiased. We prove that Pauli factor groups of order 3 N are isomorphic to all MCS's and show how this result applies in specific cases. For two qutrits, the 80 traceless operators partition into 10 MCS's. We prove that 4 of the corresponding basis sets must be separable, while 6 must be totally entangled (and Bell-like). For three qutrits, 728 operators partition into 28 MCS's with less rigid structure, allowing for the coexistence of separable, partially entangled, and totally entangled (GHZ-like) bases. However a minimum of 16 GHZ-like bases must occur. Every basis state is described by an N-digit trinary number consisting of the eigenvalues of N observables constructed from the corresponding MCS

Kinetics of swelling of polyelectrolyte gels: Fixed degree of ionization

Science.gov (United States)

Sen, Swati; Kundagrami, Arindam

2015-12-01

The swelling kinetics of uncharged and charged polymer (polyelectrolyte) gels in salt-free conditions is studied in one dimension by solving the constitutive equation of motion (Newton's law for the elementary gel volume) of the displacement variable by two theoretical methods: one in which the classical definition of stress is used with the bulk modulus taken as a parameter, and the other in which a phenomenological expression of the osmotic stress as a function of polymer density and degree of ionization is taken as an input to the dynamics. The time-evolution profiles for spatially varying polymer density and stress, along with the location of the gel-solvent interface, are obtained from the two methods. We show that both the polymer density (volume fraction) and stress inside the gel follow expected behaviours of being maximum for the uniformly shrunken gel, and relaxing slowly to the lowest values as the gel approaches equilibrium. We further show that, by comparing the temporal profiles of the gel-solvent interface and other variables between the two methods, one may attempt to assign an effective bulk modulus to the polyelectrolyte gel as a function of the degree of ionization and other parameters of the gel such as hydrophobicity, cross-link density, and the temperature. The major result we get is that the effective bulk modulus of a polyelectrolyte gel increases monotonically with its degree of ionization. In the process of identifying the parameters for a monotonic swelling, we calculated using a well-known expression of the free energy the equilibrium results of two-phase co-existence and the critical point of a polyelectrolyte gel with a fixed degree of ionization.

Characterization and identification of early proteins in Chlamydia trachomatis serovar L2 by two-dimensional gel electrophoresis

DEFF Research Database (Denmark)

Lundemose, AG; Birkelund, Svend; Larsen, PM

1990-01-01

The synthesis of early proteins from Chlamydia trachomatis serovar L2 was analyzed by two-dimensional gel electrophoresis. By pulse-label experiments, the synthesis of seven proteins was observed at 2 to 8 h postinfection before the major outer membrane protein was detected at 8 to 10 h after...

Outcome at two years after dextrose gel treatment for neonatal hypoglycemia; Follow up of a randomized trial

Science.gov (United States)

Harris, Deborah L; Alsweiler, Jane M; Ansell, Judith M; Gamble, Greg D; Thompson, Ben; Wouldes, Trecia A; Yu, Tzu-Ying; Harding, Jane E

2015-01-01

Objective To determine neurodevelopmental outcome at two years’ corrected age in children randomized to treatment with dextrose gel or placebo for hypoglycemia soon after birth (The Sugar Babies Study). Study design This was a follow-up study of 184 children who had been hypoglycemic (dextrose (90/118, 76%) or placebo gel (94/119, 79%). Assessments were performed at Kahikatea House, Hamilton, New Zealand, and included neurological function and general health (Pediatrician assessed), cognitive, language, behaviour and motor skills (Bayley-III), executive function (clinical assessment and BRIEF-P), and vision (clinical examination and global motion perception). Co-primary outcomes were neurosensory impairment (cognitive, language or motor score below −1 SD or cerebral palsy or blind or deaf) and processing difficulty (executive function or global motion perception worse than 1.5 SD from the mean). Statistical tests were two sided with 5% significance level. Results Mean (±SD) birth weight was 3093 ± 803 g and mean gestation was 37.7 ±1.6 weeks. Sixty-six children (36%) had neurosensory impairment (1 severe, 6 moderate, 59 mild) with similar rates in both groups (dextrose 38% vs. placebo 34%, RR 1.11, 95% CI 0.75–1.63). Processing difficulty was also similar between groups (dextrose 10% vs. placebo 18%, RR 0.52, 95% CI 0.23–1.15). Conclusions Dextrose gel is safe for treatment of neonatal hypoglycemia, but neurosensory impairment is common amongst these children. PMID:26613985

Proteomic study of muscle sarcoplasmic proteins using AUT-PAGE/SDS-PAGE as two-dimensional gel electrophoresis.

Science.gov (United States)

Picariello, Gianluca; De Martino, Alessandra; Mamone, Gianfranco; Ferranti, Pasquale; Addeo, Francesco; Faccia, Michele; Spagnamusso, Salvatore; Di Luccia, Aldo

2006-03-20

In the present study, an alternative procedure for two-dimensional (2D) electrophoretic analysis in proteomic investigation of the most represented basic muscle water-soluble proteins is suggested. Our method consists of Acetic acid-Urea-Triton polyacrylamide gel (AUT-PAGE) analysis in the first dimension and standard sodium dodecyl sulphate polyacrylamide gel (SDS-PAGE) in the second dimension. Although standard two-dimensional Immobilized pH Gradient-Sodium Dodecyl-Sulphate (2D IPG-SDS) gel electrophoresis has been successfully used to study these proteins, most of the water-soluble proteins are spread on the alkaline part of the 2D map and are poorly focused. Furthermore, the similarity in their molecular weights impairs resolution of the classical approach. The addition of Triton X-100, a non-ionic detergent, into the gel induces a differential electrophoretic mobility of proteins as a result of the formation of mixed micelles between the detergent and the hydrophobic moieties of polypeptides, separating basic proteins with a criterion similar to reversed phase chromatography based on their hydrophobicity. The acid pH induces positive net charges, increasing with the isoelectric point of proteins, thus allowing enhanced resolution in the separation. By using 2D AUT-PAGE/SDS electrophoresis approach to separate water-soluble proteins from fresh pork and from dry-cured products, we could spread proteins over a greater area, achieving a greater resolution than that obtained by IPG in the pH range 3-10 and 6-11. Sarcoplasmic proteins undergoing proteolysis during the ripening of products were identified by Matrix Assisted Laser Desorption/Ionization-Time of Flight (MALDI-ToF) mass spectrometry peptide mass fingerprinting in a easier and more effective way. Two-dimensional AUT-PAGE/SDS electrophoresis has allowed to simplify separation of sarcoplasmic protein mixtures making this technique suitable in the defining of quality of dry-cured pork products by immediate

SU{sub 2} nonstandard bases: the case of mutually unbiased bases

Energy Technology Data Exchange (ETDEWEB)

Olivier, Albouy; Kibler, Maurice R. [Universite de Lyon, Institut de Physique Nucleaire de Lyon, Universite Lyon, CNRS/IN2P3, 43 bd du 11 novembre 1918, F-69622 Villeurbanne Cedex (France)

2007-02-15

This paper deals with bases in a finite-dimensional Hilbert space. Such a space can be realized as a subspace of the representation space of SU{sub 2} corresponding to an irreducible representation of SU{sub 2}. The representation theory of SU{sub 2} is reconsidered via the use of two truncated deformed oscillators. This leads to replace the familiar scheme [j{sub 2}, j{sub z}] by a scheme [j{sup 2}, v{sub ra}], where the two-parameter operator v{sub ra} is defined in the universal enveloping algebra of the Lie algebra su{sub 2}. The eigenvectors of the commuting set of operators [j{sup 2}, v{sub ra}] are adapted to a tower of chains SO{sub 3} includes C{sub 2j+1} (2j belongs to N{sup *}), where C{sub 2j+1} is the cyclic group of order 2j + 1. In the case where 2j + 1 is prime, the corresponding eigenvectors generate a complete set of mutually unbiased bases. Some useful relations on generalized quadratic Gauss sums are exposed in three appendices. (authors)

An improved method for RNA isolation and cDNA library construction from immature seeds of Jatropha curcas L

Directory of Open Access Journals (Sweden)

Kaur Jatinder

2010-05-01

Full Text Available Abstract Background RNA quality and quantity is sometimes unsuitable for cDNA library construction, from plant seeds rich in oil, polysaccharides and other secondary metabolites. Seeds of jatropha (Jatropha curcas L. are rich in fatty acids/lipids, storage proteins, polysaccharides, and a number of other secondary metabolites that could either bind and/or co-precipitate with RNA, making it unsuitable for downstream applications. Existing RNA isolation methods and commercial kits often fail to deliver high-quality total RNA from immature jatropha seeds for poly(A+ RNA purification and cDNA synthesis. Findings A protocol has been developed for isolating good quality total RNA from immature jatropha seeds, whereby a combination of the CTAB based RNA extraction method and a silica column of a commercial plant RNA extraction kit is used. The extraction time was reduced from two days to about 3 hours and the RNA was suitable for poly(A+ RNA purification, cDNA synthesis, cDNA library construction, RT-PCR, and Northern hybridization. Based on sequence information from selected clones and amplified PCR product, the cDNA library seems to be a good source of full-length jatropha genes. The method was equally effective for isolating RNA from mustard and rice seeds. Conclusions This is a simple CTAB + silica column method to extract high quality RNA from oil rich immature jatropha seeds that is suitable for several downstream applications. This method takes less time for RNA extraction and is equally effective for other tissues where the quality and quantity of RNA is highly interfered by the presence of fatty acids, polysaccharides and polyphenols.

Human pro. cap alpha. 1(III) collagen: cDNA sequence for the 3' end

Energy Technology Data Exchange (ETDEWEB)

Mankoo, B S; Dalgleish, R

1988-03-25

The authors have previously isolated two overlapping cDNA clones, pIII-21 and pIII-33, which encode the C-terminal end of human type III procollagen. They now present the sequence of 2520 bases encoded in these cDNAs which overlaps other previously published sequences for the same gene. The sequence presented differs from previously published sequences at five positions.

Cloning and expression of a cDNA encoding human sterol carrier protein 2

International Nuclear Information System (INIS)

Yamamoto, Ritsu; Kallen, C.B.; Babalola, G.O.; Rennert, H.; Strauss, J.F. III; Billheimer, J.T.

1991-01-01

The authors report the cloning and expression of a cDNA encoding human sterol carrier protein 2 (SCP 2 ). The 1.3-kilobase (kb) cDNA contains an open reading frame which encompasses a 143-amino acid sequence which is 89% identical to the rat SCP 2 amino acid sequence. The deduced amino acid sequence of the polypeptide reveals a 20-residue amino-terminal leader sequence in front of the mature polypeptide, which contains a carboxyl-terminal tripeptide (Ala-Lys-Leu) related to the peroxisome targeting sequence. The expressed cDNA in COS-7 cells yields a 15.3-kDa polypeptide and increased amounts of a 13.2-kDa polypeptide, both reacting with a specific rabbit antiserum to rat liver SCP 2 . The cDNA insert hybridizes with 3.2- and 1.8-kb mRNA species in human liver poly(A) + RNA. In human fibroblasts and placenta the 1.8-kb mRNA was most abundant. Southern blot analysis suggests either that there are multiple copies of the SCP 2 gene in the human genome or that the SCP 2 gene is very large. Coexpression of the SCP 2 cDNA with expression vectors for cholesterol side-chain cleavage enzyme and adrenodoxin resulted in a 2.5-fold enhancement of progestin synthesis over that obtained with expression of the steroidogenic enzyme system alone. These findings are concordant with the notion that SCP 2 plays a role in regulating steroidogenesis, among other possible functions

Black-Box Search by Unbiased Variation

DEFF Research Database (Denmark)

Lehre, Per Kristian; Witt, Carsten

2012-01-01

The complexity theory for black-box algorithms, introduced by Droste, Jansen, and Wegener (Theory Comput. Syst. 39:525–544, 2006), describes common limits on the efficiency of a broad class of randomised search heuristics. There is an obvious trade-off between the generality of the black-box model...... and the strength of the bounds that can be proven in such a model. In particular, the original black-box model provides for well-known benchmark problems relatively small lower bounds, which seem unrealistic in certain cases and are typically not met by popular search heuristics.In this paper, we introduce a more...... restricted black-box model for optimisation of pseudo-Boolean functions which we claim captures the working principles of many randomised search heuristics including simulated annealing, evolutionary algorithms, randomised local search, and others. The key concept worked out is an unbiased variation operator...

Construction and application of a bovine immune-endocrine cDNA microarray.

Science.gov (United States)

Tao, Wenjing; Mallard, Bonnie; Karrow, Niel; Bridle, Byram

2004-09-01

A variety of commercial DNA arrays specific for humans and rodents are widely available; however, microarrays containing well-characterized genes to study pathway-specific gene expression are not as accessible for domestic animals, such as cattle, sheep and pigs. Therefore, a small-scale application-targeted bovine immune-endocrine cDNA array was developed to evaluate genetic pathways involved in the immune-endocrine axis of cattle during periods of altered homeostasis provoked by physiological or environmental stressors, such as infection, vaccination or disease. For this purpose, 167 cDNA sequences corresponding to immune, endocrine and inflammatory response genes were collected and categorized. Positive controls included 5 housekeeping genes (glyceraldehydes-3-phosphate dehydrogenase, hypoxanthine phosphoribosyltransferase, ribosomal protein L19, beta-actin, beta2-microglobulin) and bovine genomic DNA. Negative controls were a bacterial gene (Rhodococcus equi 17-kDa virulence-associated protein) and a partial sequence of the plasmid pACYC177. In addition, RNA extracted from un-stimulated, as well as superantigen (Staphylococcus aureus enterotoxin-A, S. aureus Cowan Pansorbin Cells) and mitogen-stimulated (LPS, ConA) bovine blood leukocytes was mixed, reverse transcribed and PCR amplified using gene-specific primers. The endocrine-associated genes were amplified from cDNA derived from un-stimulated bovine hypothalamus, pituitary, adrenal and thyroid gland tissues. The array was constructed in 4 repeating grids of 180 duplicated spots by coupling the PCR amplified 213-630 bp gene fragments onto poly-l-lysine coated glass slides. The bovine immune-endocrine arrays were standardized and preliminary gene expression profiles generated using Cy3 and Cy5 labelled cDNA from un-stimulated and ConA (5 microg/ml) stimulated PBMC of 4 healthy Holstein cows (2-4 replicate arrays/cow) in a time course study. Mononuclear cell-derived cytokine and chemokine (IL-2, IL-1alpha

Construction of a cDNA microarray derived from the ascidian Ciona intestinalis.

Science.gov (United States)

Azumi, Kaoru; Takahashi, Hiroki; Miki, Yasufumi; Fujie, Manabu; Usami, Takeshi; Ishikawa, Hisayoshi; Kitayama, Atsusi; Satou, Yutaka; Ueno, Naoto; Satoh, Nori

2003-10-01

A cDNA microarray was constructed from a basal chordate, the ascidian Ciona intestinalis. The draft genome of Ciona has been read and inferred to contain approximately 16,000 protein-coding genes, and cDNAs for transcripts of 13,464 genes have been characterized and compiled as the "Ciona intestinalis Gene Collection Release I". In the present study, we constructed a cDNA microarray of these 13,464 Ciona genes. A preliminary experiment with Cy3- and Cy5-labeled probes showed extensive differential gene expression between fertilized eggs and larvae. In addition, there was a good correlation between results obtained by the present microarray analysis and those from previous EST analyses. This first microarray of a large collection of Ciona intestinalis cDNA clones should facilitate the analysis of global gene expression and gene networks during the embryogenesis of basal chordates.

Thixotropic corrosive gels for nuclear decontamination

International Nuclear Information System (INIS)

Bargues, St.

1998-01-01

The aim of this thesis was the development of corrosive gels for metallic surface decontamination. These gels formulation, based on a powerful oxidant (the cerium IV), the nitric acid, a mineral charge (silica) and a non ionic surface-active, has been developed according to the specific constraints of the nuclear industry. The objective was to prepare thixotropic gels becoming liquid after shacking to allow an easy pulverization and coming again solid to permit a perfect adhesion on the metallic surface. This rheological study of the gels has been completed by an evaluation of their corrosive properties. The last part of the work presents an industrial utilization during two years. (A.L.B.)

Structure of gels layers with cells

Science.gov (United States)

Pokusaev, B. G.; Karlov, S. P.; Vyazmin, A. V.; Nekrasov, D. A.; Zakharov, N. S.; Khramtsov, D. P.; Skladnev, D. A.; Tyupa, D. V.

2017-11-01

The structure of two-layer agarose gels containing yeast cells is investigated experimentally by spectrometry, to shed a light on the theoretical foundations for the development of bioreactors by the method of 3D bioprinting. Due to division, cells overcome the layer of the dispersion phase separating successively applied layers of the agarose gel. However a gel layer of 100 μm thick with a high concentration of silver nanoparticles completely excludes the infiltration of yeast cells through it. A special sort of agarose is suggested where the concentration of silver nanoparticles formed by cells from salt of silver can serve as an indicator of the state of the yeast cells in the volume of the gel.

cDNA fingerprinting of osteoprogenitor cells to isolate differentiation stage-specific genes.

OpenAIRE

Candeliere, G A; Rao, Y; Floh, A; Sandler, S D; Aubin, J E

1999-01-01

A cDNA fingerprinting strategy was developed to identify genes based on their differential expression pattern during osteoblast development. Preliminary biological and molecular staging of cDNA pools prepared by global amplification PCR allowed discrim-inating choices to be made in selection of expressed sequence tags (ESTs) to be isolated. Sequencing of selected ESTs confirmed that both known and novel genes can be isolated from any developmental stage of interest, e.g. from primitive progen...

Contaminant containment using polymer gel barriers

NARCIS (Netherlands)

Darwish, M.I.M.; Rowe, R.K.; Maarel, van der J.R.C.; Pel, L.; Huinink, H.P.; Zitha, P.L.J.

2004-01-01

Polymer gels are well known in the oil industry, but their potential for use as barriers to contaminant transport has not previously received significant study. As a first step, this paper examines the potential for a polyelectrolyte gel to serve as a barrier to the migration of sodium chloride. Two

Experimental study on improved two-bed silica gel-water adsorption chiller

Energy Technology Data Exchange (ETDEWEB)

Xia Zaizhong [Institute of Refrigeration and Cryogenics, Shanghai Jiao Tong University, Shanghai 200030 (China)], E-mail: xzz@sjtu.edu.cn; Wang Dechang; Zhang Jincui [College of Electromechanical Engineering, Qingdao University, Qingdao 266071 (China)

2008-06-15

A novel silica gel-water adsorption chiller with two chambers has been built in Shanghai Jiao Tong University (SJTU). This chiller combines two single bed systems (basic system) without any vacuum valves. One adsorber, one condenser and one evaporator are housed in the same chamber to constitute one adsorption/desorption unit. In this work, the chiller is developed and improved. The improved chiller is composed of three vacuum chambers: two adsorption/desorption vacuum chambers (the same structure as the former chiller) and one heat pipe working vacuum chamber. The evaporators of these two adsorption/desorption units are combined by a heat pipe. So, no valves are installed in the chilled water sub system and one vacuum valve connects the two adsorption/desorption chambers together to improve its performance. The performance of the chiller is tested. As the results, the refrigerating capacity and the COP of the chiller are, respectively, 8.69 kW and 0.388 for the heat source temperature of 82.5 deg. C, the cooling water temperature of 30.4 deg. C and the chilled water outlet temperature of 11.9 deg. C. For a chilled water outlet temperature of 16.5 deg. C, the COP reaches 0.432, while the refrigerating capacity is near 11 kW. There is an improvement of at least 12% for the COP compared with the former chillers.

Experimental study on improved two-bed silica gel-water adsorption chiller

Energy Technology Data Exchange (ETDEWEB)

Xia, Zaizhong [Institute of Refrigeration and Cryogenics, Shanghai Jiao Tong University, Shanghai 200030 (China); Wang, Dechang; Zhang, Jincui [College of Electromechanical Engineering, Qingdao University, Qingdao 266071 (China)

2008-06-15

A novel silica gel-water adsorption chiller with two chambers has been built in Shanghai Jiao Tong University (SJTU). This chiller combines two single bed systems (basic system) without any vacuum valves. One adsorber, one condenser and one evaporator are housed in the same chamber to constitute one adsorption/desorption unit. In this work, the chiller is developed and improved. The improved chiller is composed of three vacuum chambers: two adsorption/desorption vacuum chambers (the same structure as the former chiller) and one heat pipe working vacuum chamber. The evaporators of these two adsorption/desorption units are combined by a heat pipe. So, no valves are installed in the chilled water sub system and one vacuum valve connects the two adsorption/desorption chambers together to improve its performance. The performance of the chiller is tested. As the results, the refrigerating capacity and the COP of the chiller are, respectively, 8.69 kW and 0.388 for the heat source temperature of 82.5 C, the cooling water temperature of 30.4 C and the chilled water outlet temperature of 11.9 C. For a chilled water outlet temperature of 16.5 C, the COP reaches 0.432, while the refrigerating capacity is near 11 kW. There is an improvement of at least 12% for the COP compared with the former chillers. (author)

Avoiding cross hybridization by choosing nonredundant targets on cDNA arrays

DEFF Research Database (Denmark)

Nielsen, Henrik Bjørn; Knudsen, Steen

2002-01-01

PROBEWIZ designs PCR primers for amplifying probes for cDNA arrays. The probes are designed to have minimal homology to other expressed sequences from a given organism. The primer selection is based on user-defined penalties for homology, primer quality, and proximity to the 3' end.......PROBEWIZ designs PCR primers for amplifying probes for cDNA arrays. The probes are designed to have minimal homology to other expressed sequences from a given organism. The primer selection is based on user-defined penalties for homology, primer quality, and proximity to the 3' end....

cDNA cloning and mRNA expression of heat shock protein 70 gene ...

African Journals Online (AJOL)

In this study, the full-length heat shock protein 70 of Tegillarca granosa was cloned from cDNA library by rapid amplification of cDNA end (RACE). The open reading frame (ORF) of heat shock protein 70 was 1968 bp, and it encoded a protein of 655 amino acids with a predicted molecular weight of 71.48 kDa and an ...

Pyroelectric photovoltaic spatial solitons in unbiased photorefractive crystals

International Nuclear Information System (INIS)

Jiang, Qichang; Su, Yanli; Ji, Xuanmang

2012-01-01

A new type of spatial solitons i.e. pyroelectric photovoltaic spatial solitons based on the combination of pyroelectric and photovoltaic effect is predicted theoretically. It shows that bright, dark and grey spatial solitons can exist in unbiased photovoltaic photorefractive crystals with appreciable pyroelectric effect. Especially, the bright soliton can form in self-defocusing photovoltaic crystals if it gives larger self-focusing pyroelectric effect. -- Highlights: ► A new type of spatial soliton i.e. pyroelectric photovoltaic spatial soliton is predicted. ► The bright, dark and grey pyroelectric photovoltaic spatial soliton can form. ► The bright soliton can also exist in self-defocusing photovoltaic crystals.

On the mathematical foundations of mutually unbiased bases

Science.gov (United States)

Thas, Koen

2018-02-01

In order to describe a setting to handle Zauner's conjecture on mutually unbiased bases (MUBs) (stating that in C^d, a set of MUBs of the theoretical maximal size d + 1 exists only if d is a prime power), we pose some fundamental questions which naturally arise. Some of these questions have important consequences for the construction theory of (new) sets of maximal MUBs. Partial answers will be provided in particular cases; more specifically, we will analyze MUBs with associated operator groups that have nilpotence class 2, and consider MUBs of height 1. We will also confirm Zauner's conjecture for MUBs with associated finite nilpotent operator groups.

Molecular characterization of two isoforms of ZFAND3 cDNA from the Japanese quail and the leopard gecko, and different expression patterns between testis and ovary.

Science.gov (United States)

Otake, Shigeo; Endo, Daisuke; Park, Min Kyun

2011-11-15

Zing finger AN1-type domain 3 (ZFAND3), also known as testis expressed sequence 27 (Tex27), is a gene found in the mouse testis, but its physiological function is unknown. We identified the full-length sequences of two isoforms (short and long) of ZFAND3 cDNA from Japanese quail and leopard gecko. This is the first cloning of avian and reptilian ZFAND3 cDNA. The two isoforms are generated by alternative polyadenylation in the 3'UTR and have the same ORF sequences encoding identical proteins. There were highly conserved regions in the 3'UTR of the long form near the polyadenylation sites from mammals to amphibians, suggesting that the features for determining the stability of mRNA or translation efficiency differ between isoforms. The deduced amino acid sequence of ZFAND3 has two putative zinc finger domains, an A20-like zinc finger domain at the N-terminal and an AN1-like zinc finger domain at the C-terminal. Sequence analysis revealed an additional exon in the genomic structures of the avian and reptilian ZFAND3 genes which is not present in mammals, amphibians, or fish, and this exon produces additional amino acid residues in the A20-like zinc finger domain. Expression analysis in Japanese quail revealed that the expression level of ZFAND3 mRNA was high in not only the testis but also the ovary, and ZFAND3 mRNA was expressed in both spermatides of the testis and oocytes of the ovary. While the short form mRNA was mainly expressed in the testis, the expression level of the long form mRNA was high in the ovary. These results suggest that ZFAND3 has physiological functions related to germ cell maturation and regulatory mechanisms that differ between the testis and ovary. Copyright © 2011 Elsevier B.V. All rights reserved.

An asymptotically unbiased minimum density power divergence estimator for the Pareto-tail index

DEFF Research Database (Denmark)

Dierckx, Goedele; Goegebeur, Yuri; Guillou, Armelle

2013-01-01

We introduce a robust and asymptotically unbiased estimator for the tail index of Pareto-type distributions. The estimator is obtained by fitting the extended Pareto distribution to the relative excesses over a high threshold with the minimum density power divergence criterion. Consistency...

Electrochemical behavior of ionically crosslinked polyampholytic gel electrolytes

International Nuclear Information System (INIS)

Chen Wanyu; Tang Haitao; Ou Ziwei; Wang Hong; Yang Yajiang

2007-01-01

An ionic complex of anionic and cationic monomers was obtained by protonation of (N,N-diethylamino)ethylmethacrylate (DEA) with acrylic acid (AAc). Free radical copolymerization of the ionic complex and acrylamide (AAm), yielded the ionically crosslinked polyampholytic gel electrolytes [poly(AAc-DEA-AAm), designated as PADA] using two types of organic solvents containing a lithium salt. The PADA gel electrolyte exhibited good thermal stability shown by the DSC thermogram. The impedance analysis at temperatures ranging from -30 to 75 deg. C indicated that the ionic conductivities of the PADA gel electrolytes were rather close to those of liquid electrolytes. The temperature dependence of the ionic conductivities was found to be in accord with the Arrhenius equation. Moreover, the ionic conductivities of PADA gel electrolytes increased with an increase of the molar ratios of cationic/anionic monomers. The ionic conductivities of PADA gels prepared in solvent mixtures of propylene carbonate, ethyl methyl ether and dioxolane (3:1:1, v/v) were higher than those of PADA gels prepared in propylene carbonate only. Significantly, the ionic conductivities of two kinds of PADA gel electrolytes were in the range of 10 -3 and 10 -4 S cm -1 even at -30 deg. C. The electrochemical windows of PADA gel electrolytes measured by cyclic voltammetry were in the range from -1 V to 4.5 V

Acoustic evaluation of polymer gel dosimeters

International Nuclear Information System (INIS)

Mather, M.L.; De Deene, Y.; Baldock, C.; Whittaker, A.K.

2002-01-01

Advances in radiotherapy treatment techniques such as intensity modulated radiotherapy are placing increasing demands on radiation dosimetry for verification of dose distributions in 3D. In response, polymer gel dosimeters that are capable of recording dose distributions in 3D are currently being developed. Recently, a new technique for evaluation of absorbed dose distributions in these dosimeters using ultrasound was introduced. The current work aims to demonstrate the potential of ultrasound as an evaluation technique for polymer gel dosimeters and to investigate the ultrasound properties of two different dosimeter formulations, PAG and MAGIC gels

Sol-Gel Glasses

Science.gov (United States)

Mukherjee, S. P.

1985-01-01

Multicomponent homogeneous, ultrapure noncrystalline gels/gel derived glasses are promising batch materials for the containerless glass melting experiments in microgravity. Hence, ultrapure, homogeneous gel precursors could be used to: (1) investigate the effect of the container induced nucleation on the glass forming ability of marginally glass forming compositions; and (2) investigate the influence of gravity on the phase separation and coarsening behavior of gel derived glasses in the liquid-liquid immiscibility zone of the nonsilicate systems having a high density phase. The structure and crystallization behavior of gels in the SiO2-GeO2 as a function of gel chemistry and thermal treatment were investigated. As are the chemical principles involved in the distribution of a second network former in silica gel matrix being investigated. The procedures for synthesizing noncrystalline gels/gel-monoliths in the SiO2-GeO2, GeO2-PbO systems were developed. Preliminary investigations on the levitation and thermal treatment of germania silicate gel-monoliths in the Pressure Facility Acoustic Levitator were done.

Circulating tumor cell detection: A direct comparison between negative and unbiased enrichment in lung cancer.

Science.gov (United States)

Xu, Yan; Liu, Biao; Ding, Fengan; Zhou, Xiaodie; Tu, Pin; Yu, Bo; He, Yan; Huang, Peilin

2017-06-01

Circulating tumor cells (CTCs), isolated as a 'liquid biopsy', may provide important diagnostic and prognostic information. Therefore, rapid, reliable and unbiased detection of CTCs are required for routine clinical analyses. It was demonstrated that negative enrichment, an epithelial marker-independent technique for isolating CTCs, exhibits a better efficiency in the detection of CTCs compared with positive enrichment techniques that only use specific anti-epithelial cell adhesion molecules. However, negative enrichment techniques incur significant cell loss during the isolation procedure, and as it is a method that uses only one type of antibody, it is inherently biased. The detection procedure and identification of cell types also relies on skilled and experienced technicians. In the present study, the detection sensitivity of using negative enrichment and a previously described unbiased detection method was compared. The results revealed that unbiased detection methods may efficiently detect >90% of cancer cells in blood samples containing CTCs. By contrast, only 40-60% of CTCs were detected by negative enrichment. Additionally, CTCs were identified in >65% of patients with stage I/II lung cancer. This simple yet efficient approach may achieve a high level of sensitivity. It demonstrates a potential for the large-scale clinical implementation of CTC-based diagnostic and prognostic strategies.

Mass spectrometry-based cDNA profiling as a potential tool for human body fluid identification.

Science.gov (United States)

Donfack, Joseph; Wiley, Anissa

2015-05-01

Several mRNA markers have been exhaustively evaluated for the identification of human venous blood, saliva, and semen in forensic genetics. As new candidate human body fluid specific markers are discovered, evaluated, and reported in the scientific literature, there is an increasing trend toward determining the ideal markers for cDNA profiling of body fluids of forensic interest. However, it has not been determined which molecular genetics-based technique(s) should be utilized to assess the performance of these markers. In recent years, only a few confirmatory, mRNA/cDNA-based methods have been evaluated for applications in body fluid identification. The most frequently described methods tested to date include quantitative polymerase chain reaction (qPCR) and capillary electrophoresis (CE). However these methods, in particular qPCR, often favor narrow multiplex PCR due to the availability of a limited number of fluorescent dyes/tags. In an attempt to address this technological constraint, this study explored matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) for human body fluid identification via cDNA profiling of venous blood, saliva, and semen. Using cDNA samples at 20pg input phosphoglycerate kinase 1 (PGK1) amounts, body fluid specific markers for the candidate genes were amplified in their corresponding body fluid (i.e., venous blood, saliva, or semen) and absent in the remaining two (100% specificity). The results of this study provide an initial indication that MALDI-TOF MS is a potential fluorescent dye-free alternative method for body fluid identification in forensic casework. However, the inherent issues of low amounts of mRNA, and the damage caused to mRNA by environmental exposures, extraction processes, and storage conditions are important factors that significantly hinder the implementation of cDNA profiling into forensic casework. Published by Elsevier Ireland Ltd.

Cloning, sequencing and expression of cDNA encoding growth ...

Indian Academy of Sciences (India)

Unknown

of medicine, animal husbandry, fish farming and animal ..... northern pike (Esox lucius) growth hormone; Mol. Mar. Biol. ... prolactin 1-luciferase fusion gene in African catfish and ... 1988 Cloning and sequencing of cDNA that encodes goat.

Human uroporphyrinogen III synthase: Molecular cloning, nucleotide sequence, and expression of a full-length cDNA

International Nuclear Information System (INIS)

Tsai, Shihfeng; Bishop, D.F.; Desnick, R.J.

1988-01-01

Uroporphyrinogen III synthase, the fourth enzyme in the heme biosynthetic pathway, is responsible for conversion of the linear tetrapyrrole, hydroxymethylbilane, to the cyclic tetrapyrrole, uroporphyrinogen III. The deficient activity of URO-synthase is the enzymatic defect in the autosomal recessive disorder congenital erythropoietic porphyria. To facilitate the isolation of a full-length cDNA for human URO-synthase, the human erythrocyte enzyme was purified to homogeneity and 81 nonoverlapping amino acids were determined by microsequencing the N terminus and four tryptic peptides. Two synthetic oligonucleotide mixtures were used to screen 1.2 x 10 6 recombinants from a human adult liver cDNA library. Eight clones were positive with both oligonucleotide mixtures. Of these, dideoxy sequencing of the 1.3 kilobase insert from clone pUROS-2 revealed 5' and 3' untranslated sequences of 196 and 284 base pairs, respectively, and an open reading frame of 798 base pairs encoding a protein of 265 amino acids with a predicted molecular mass of 28,607 Da. The isolation and expression of this full-length cDNA for human URO-synthase should facilitate studies of the structure, organization, and chromosomal localization of this heme biosynthetic gene as well as the characterization of the molecular lesions causing congenital erythropoietic porphyria

Isolation and characterization of cDNA clones for carrot extensin and a proline-rich 33-kDa protein

International Nuclear Information System (INIS)

Chen, J.; Varner, J.E.

1985-01-01

Extensins are hydroxyproline-rich glycoproteins associated with most dicotyledonous plant cell walls. To isolate cDNA clones encoding extensin, the authors started by isolating poly(A) + RNA from carrot root tissue, and then translating the RNA in vitro, in the presence of tritiated leucine or proline. A 33-kDa peptide was identified in the translation products as a putative extensin precursor. From a cDNA library constructed with poly(A) + RNA from wounded carrots, one cDNA clone (pDC5) was identified that specifically hybridized to poly(A) + RNA encoding this 33-kDa peptide. They isolated three cDNA clones (pDC11, pDC12, and pDC16) from another cDNA library using pCD5 as a probe. DNA sequence data, RNA hybridization analysis, and hybrid released in vitro translation indicate that the cDNA clones pDC11 encodes extensin and that cDNA clones pDC12 and pDC16 encode the 33-kDa peptide, which as yet has an unknown identity and function. The assumption that the 33-kDa peptide was an extensin precursor was invalid. RNA hybridization analysis showed that RNA encoded by both clone types is accumulated upon wounding

Identification of cysteine proteases and screening of cysteine protease inhibitors in biological samples by a two-dimensional gel system of zymography and reverse zymography.

Science.gov (United States)

Saitoh, Eiichi; Yamamoto, Shinya; Okamoto, Eishiro; Hayakawa, Yoshimi; Hoshino, Takashi; Sato, Ritsuko; Isemura, Satoko; Ohtsubo, Sadami; Taniguchi, Masayuki

2007-11-18

We have developed a two-dimensional (2D-) gel system of zymography and reverse zymography for the detection and characterization of proteases and protease inhibitors. Isoelectric focusing (IEF) agarose gels with pH gradients were employed for separation in the first-dimension and sodium dodecyl sulfate (SDS)-polyacrylamide gel copolymerized with gelatin used for the second dimension. Proteases and protease inhibitors separated by IEF gel were applied on the second gel without trichloroacetic acid (TCA) fixation. Protease activity in the 2D-gel was visualized as transparent spots where gelatin substrate was digested after commassie brilliant blue (CBB) staining. Some of the transparent spots from the skin mucus extract of rainbow trout were determined to be a cysteine protease through use of E-64 or CA-074. In the reverse zymography technique, the gel was incubated with papain solution at 37 degrees C for 18 h. Cysteine protease inhibitors from broad bean seeds were detected as clear blue spots after CBB staining. The amino (N-) terminal sequences of four papain inhibitor spots thus detected were demonstrated to be identical to that of favin beta chain, a broad bean lectin. Taken together, our system can be considered to be an efficient technique for discovering and characterizing new proteases and protease inhibitors in biological samples. This is the first report describing a 2D-gel system of zymography and reverse zymography.

On shear rheology of gel propellants

Energy Technology Data Exchange (ETDEWEB)

Rahimi, Shai; Peretz, Arie [RAFAEL, MANOR Propulsion and Explosive Systems Division, Haifa (Israel); Natan, Benveniste [Faculty of Aerospace Engineering, Technion - Israel Institute of Technology, Haifa (Israel)

2007-04-15

Selected fuel, oxidizer and simulant gels were prepared and rheologically characterized using a rotational rheometer. For fuel gelation both organic and inorganic gellants were utilized, whereas oxidizers and simulants were gelled with addition of silica and polysaccharides, respectively. The generalized Herschel-Bulkley constitutive model was found to most adequately represent the gels studied. Hydrazine-based fuels, gelled with polysaccharides, were characterized as shear-thinning pseudoplastic fluids with low shear yield stress, whereas inhibited red-fuming nitric acid (IRFNA) and hydrogen peroxide oxidizers, gelled with silica, were characterized as yield thixotropic fluids with significant shear yield stress. Creep tests were conducted on two rheological types of gels with different gellant content and the results were fitted by Burgers-Kelvin viscoelastic constitutive model. The effect of temperature on the rheological properties of gel propellant simulants was also investigated. A general rheological classification of gel propellants and simulants is proposed. (Abstract Copyright [2007], Wiley Periodicals, Inc.)

Isolation and sequence of cDNA encoding a cytochrome P-450 from an insecticide-resistant strain of the house fly, Musca domestica.

OpenAIRE

Feyereisen, R; Koener, J F; Farnsworth, D E; Nebert, D W

1989-01-01

A cDNA expression library from phenobarbital-treated house fly (Musca domestica) was screened with rabbit antisera directed against partially purified house fly cytochrome P-450. Two overlapping clones with insert lengths of 1.3 and 1.5 kilobases were isolated. The sequence of a 1629-base-pair (bp) cDNA was obtained, with an open reading frame (nucleotides 81-1610) encoding a P-450 protein of 509 residues (Mr = 58,738). The insect P-450 protein contains a hydrophobic NH2 terminus and a 22-res...

Cloning of the cDNA for a human homologue of the Drosophila white gene and mapping to chromosome 21q22.3.

OpenAIRE

Chen, H.; Rossier, C.; Lalioti, M. D.; Lynn, A.; Chakravarti, A.; Perrin, G.; Antonarakis, S. E.

1996-01-01

In an effort to contribute to the transcript map of human chromosome 21 and the understanding of the pathophysiology of trisomy 21, we have used exon trapping to identify fragments of chromosome 21 genes. Two trapped exons, from pools of chromosome 21-specific cosmids, showed homology to the Drosophila white (w) gene. We subsequently cloned the corresponding cDNA for a human homologue of the Drosophila w gene (hW) from human retina and fetal brain cDNA libraries. The gene belongs to the ATP-b...

Detection of Aspartic Proteinase Activities Using Gel Zymography.

Science.gov (United States)

Perera, Handunge Kumudu Irani

2017-01-01

Gel zymography is a two-stage process where the proteins from the test sample are first separated by electrophoresis followed by the detection of the activity of hydrolytic enzymes. Many zymography procedures use sodium dodecyl sulfate (SDS) polyacrylamide gels copolymerized with an appropriate substrate. The procedure described here uses native polyacrylamide gel electrophoresis (PAGE) in the absence of both SDS and substrate. In order to visualize aspartic proteinase activity, the gel is impregnated in bovine hemoglobin at pH 3.0 for 15 min after the electrophoresis procedure. Subsequently, the gel is incubated in a humid container in the absence of hemoglobin for 1 h at 37 °C. At the end, the gel is stained with amido black and destained. Clear areas against a dark background corresponding to aspartic proteinase activities can be detected.

PCR amplification on microarrays of gel immobilized oligonucleotides

Science.gov (United States)

Strizhkov, Boris; Tillib, Sergei; Mikhailovich, Vladimir; Mirzabekov, Andrei

2003-11-04

The invention relates two general methods for performing PCR amplification, combined with the detection and analysis of the PCR products on a microchip. In the first method, the amplification occurs both outside and within a plurality of gel pads on a microchip, with at least one oligonucleotide primer immobilized in a gel pad. In the second method, PCR amplification also takes place within gel pads on a microchip, but the pads are surrounded by a hydrophobic liquid such as that which separates the individual gel pads into environments which resemble micro-miniaturized test tubes.

Cloning and expression of a cDNA coding for a human monocyte-derived plasminogen activator inhibitor.

OpenAIRE

Antalis, T M; Clark, M A; Barnes, T; Lehrbach, P R; Devine, P L; Schevzov, G; Goss, N H; Stephens, R W; Tolstoshev, P

1988-01-01

Human monocyte-derived plasminogen activator inhibitor (mPAI-2) was purified to homogeneity from the U937 cell line and partially sequenced. Oligonucleotide probes derived from this sequence were used to screen a cDNA library prepared from U937 cells. One positive clone was sequenced and contained most of the coding sequence as well as a long incomplete 3' untranslated region (1112 base pairs). This cDNA sequence was shown to encode mPAI-2 by hybrid-select translation. A cDNA clone encoding t...

GelTouch

DEFF Research Database (Denmark)

Miruchna, Viktor; Walter, Robert; Lindlbauer, David

2015-01-01

We present GelTouch, a gel-based layer that can selectively transition between soft and stiff to provide tactile multi-touch feedback. It is flexible, transparent when not activated, and contains no mechanical, electromagnetic, or hydraulic components, resulting in a compact form factor (a 2mm thin...... touchscreen layer for our prototype). The activated areas can be morphed freely and continuously, without being limited to fixed, predefined shapes. GelTouch consists of a poly(N-isopropylacrylamide) gel layer which alters its viscoelasticity when activated by applying heat (>32 C). We present three different...

Collision energy alteration during mass spectrometric acquisition is essential to ensure unbiased metabolomic analysis

CSIR Research Space (South Africa)

Madala, NE

2012-08-01

Full Text Available Metabolomics entails identification and quantification of all metabolites within a biological system with a given physiological status; as such, it should be unbiased. A variety of techniques are used to measure the metabolite content of living...

Differential representation of sunflower ESTs in enriched organ-specific cDNA libraries in a small scale sequencing project

Directory of Open Access Journals (Sweden)

Heinz Ruth A

2003-09-01

Full Text Available Abstract Background Subtractive hybridization methods are valuable tools for identifying differentially regulated genes in a given tissue avoiding redundant sequencing of clones representing the same expressed genes, maximizing detection of low abundant transcripts and thus, affecting the efficiency and cost effectiveness of small scale cDNA sequencing projects aimed to the specific identification of useful genes for breeding purposes. The objective of this work is to evaluate alternative strategies to high-throughput sequencing projects for the identification of novel genes differentially expressed in sunflower as a source of organ-specific genetic markers that can be functionally associated to important traits. Results Differential organ-specific ESTs were generated from leaf, stem, root and flower bud at two developmental stages (R1 and R4. The use of different sources of RNA as tester and driver cDNA for the construction of differential libraries was evaluated as a tool for detection of rare or low abundant transcripts. Organ-specificity ranged from 75 to 100% of non-redundant sequences in the different cDNA libraries. Sequence redundancy varied according to the target and driver cDNA used in each case. The R4 flower cDNA library was the less redundant library with 62% of unique sequences. Out of a total of 919 sequences that were edited and annotated, 318 were non-redundant sequences. Comparison against sequences in public databases showed that 60% of non-redundant sequences showed significant similarity to known sequences. The number of predicted novel genes varied among the different cDNA libraries, ranging from 56% in the R4 flower to 16 % in the R1 flower bud library. Comparison with sunflower ESTs on public databases showed that 197 of non-redundant sequences (60% did not exhibit significant similarity to previously reported sunflower ESTs. This approach helped to successfully isolate a significant number of new reported sequences

NORMAL NASAL GENE EXPRESSION LEVELS USING CDNA ARRAY TECHNOLOGY

Science.gov (United States)

Normal Nasal Gene Expression Levels Using cDNA Array Technology. The nasal epithelium is a target site for chemically-induced toxicity and carcinogenicity. To detect and analyze genetic events which contribute to nasal tumor development, we first defined the gene expressi...

The gel electrophoresis markup language (GelML) from the Proteomics Standards Initiative.

Science.gov (United States)

Gibson, Frank; Hoogland, Christine; Martinez-Bartolomé, Salvador; Medina-Aunon, J Alberto; Albar, Juan Pablo; Babnigg, Gyorgy; Wipat, Anil; Hermjakob, Henning; Almeida, Jonas S; Stanislaus, Romesh; Paton, Norman W; Jones, Andrew R

2010-09-01

The Human Proteome Organisation's Proteomics Standards Initiative has developed the GelML (gel electrophoresis markup language) data exchange format for representing gel electrophoresis experiments performed in proteomics investigations. The format closely follows the reporting guidelines for gel electrophoresis, which are part of the Minimum Information About a Proteomics Experiment (MIAPE) set of modules. GelML supports the capture of metadata (such as experimental protocols) and data (such as gel images) resulting from gel electrophoresis so that laboratories can be compliant with the MIAPE Gel Electrophoresis guidelines, while allowing such data sets to be exchanged or downloaded from public repositories. The format is sufficiently flexible to capture data from a broad range of experimental processes, and complements other PSI formats for MS data and the results of protein and peptide identifications to capture entire gel-based proteome workflows. GelML has resulted from the open standardisation process of PSI consisting of both public consultation and anonymous review of the specifications.

Stacking gels: A method for maximising output for pulsed-field gel electrophoresis

Directory of Open Access Journals (Sweden)

Heng See

2009-01-01

Full Text Available Pulsed field gel electrophoresis (PFGE, the gold standard of molecular typing methods, has a major disadvantage of an unusually long electrophoretic time. From the original protocol of 6 days, it was modified to 3 days and subsequently to a single day. We describe the procedure of stacking five to six gels one on top of another in order to increase and maximize the output in a shorter time without compromising the resolution and reproducibility. All the variables that affect pulsed field gels during electrophoresis were taken into consideration. We firstly optimized the parameters to be used and secondly determined whether stacking of five to six gels had any effect on the molecular separation during electrophoresis in comparison with a single gel run. DNA preparation, restriction, electrophoresis, staining and gel documentation was carried out based on previously published methods. Gels were analysed using BioNumerics and dice coefficient and unweighted pair group methods were used to generate dendrograms based on 1.5% tolerance values. Identical band profiles and band resolution-separation were seen in the PFGE patterns with single gel and multiple stacking gels. Cluster analysis further strengthened the fact that results from stacking gels were reproducible and comparable with a single gel run. This method of stacking gels saves time and maximizes the output at the same time. The run time for a single gel was about 28 hours, but with six stacked gels the run time was 54 hours compared with 28 x 6 = 168 hours if they were run separately as single gels thus saving time of 67.86%. Beside the big factor of saving time, stacking gels save resources (electricity, reagents, water, chemicals and working time by increasing the sample throughput in a shorter time without compromising on quality of data. But optimization of working parameters is vital depending on the PFGE system used.

Reconstruction of high-dimensional states entangled in orbital angular momentum using mutually unbiased measurements

CSIR Research Space (South Africa)

Giovannini, D

2013-06-01

Full Text Available : QELS_Fundamental Science, San Jose, California United States, 9-14 June 2013 Reconstruction of High-Dimensional States Entangled in Orbital Angular Momentum Using Mutually Unbiased Measurements D. Giovannini1, ⇤, J. Romero1, 2, J. Leach3, A...

ESTs, cDNA microarrays, and gene expression profiling: tools for dissecting plant physiology and development.

Science.gov (United States)

Alba, Rob; Fei, Zhangjun; Payton, Paxton; Liu, Yang; Moore, Shanna L; Debbie, Paul; Cohn, Jonathan; D'Ascenzo, Mark; Gordon, Jeffrey S; Rose, Jocelyn K C; Martin, Gregory; Tanksley, Steven D; Bouzayen, Mondher; Jahn, Molly M; Giovannoni, Jim

2004-09-01

Gene expression profiling holds tremendous promise for dissecting the regulatory mechanisms and transcriptional networks that underlie biological processes. Here we provide details of approaches used by others and ourselves for gene expression profiling in plants with emphasis on cDNA microarrays and discussion of both experimental design and downstream analysis. We focus on methods and techniques emphasizing fabrication of cDNA microarrays, fluorescent labeling, cDNA hybridization, experimental design, and data processing. We include specific examples that demonstrate how this technology can be used to further our understanding of plant physiology and development (specifically fruit development and ripening) and for comparative genomics by comparing transcriptome activity in tomato and pepper fruit.

Pattern analysis approach reveals restriction enzyme cutting abnormalities and other cDNA library construction artifacts using raw EST data

Directory of Open Access Journals (Sweden)

Zhou Sun

2012-05-01

Full Text Available Abstract Background Expressed Sequence Tag (EST sequences are widely used in applications such as genome annotation, gene discovery and gene expression studies. However, some of GenBank dbEST sequences have proven to be “unclean”. Identification of cDNA termini/ends and their structures in raw ESTs not only facilitates data quality control and accurate delineation of transcription ends, but also furthers our understanding of the potential sources of data abnormalities/errors present in the wet-lab procedures for cDNA library construction. Results After analyzing a total of 309,976 raw Pinus taeda ESTs, we uncovered many distinct variations of cDNA termini, some of which prove to be good indicators of wet-lab artifacts, and characterized each raw EST by its cDNA terminus structure patterns. In contrast to the expected patterns, many ESTs displayed complex and/or abnormal patterns that represent potential wet-lab errors such as: a failure of one or both of the restriction enzymes to cut the plasmid vector; a failure of the restriction enzymes to cut the vector at the correct positions; the insertion of two cDNA inserts into a single vector; the insertion of multiple and/or concatenated adapters/linkers; the presence of 3′-end terminal structures in designated 5′-end sequences or vice versa; and so on. With a close examination of these artifacts, many problematic ESTs that have been deposited into public databases by conventional bioinformatics pipelines or tools could be cleaned or filtered by our methodology. We developed a software tool for Abnormality Filtering and Sequence Trimming for ESTs (AFST, http://code.google.com/p/afst/ using a pattern analysis approach. To compare AFST with other pipelines that submitted ESTs into dbEST, we reprocessed 230,783 Pinus taeda and 38,709 Arachis hypogaea GenBank ESTs. We found 7.4% of Pinus taeda and 29.2% of Arachis hypogaea GenBank ESTs are “unclean” or abnormal, all of which could be cleaned

Application of Singh et al., unbiased estimator in a dual to ratio-cum ...

African Journals Online (AJOL)

This paper applied an unbiased estimator in a dual to ratio–cum-product estimator in sample surveys to double sampling design. Its efficiency over the conventional biased double sampling design estimator was determined based on the conditions attached to its supremacy. Three different data sets were used to testify to ...

Light scattering from a binary-liquid entanglement gel

Science.gov (United States)

Xia, K.-Q.; Maher, J. V.

1987-09-01

Light-scattering experiments have been carried out on an entanglement gel with a binary-liquid mixture as solvent. The onset temperature for critical opalescence has a composition dependence which is similar to the coexistence curve of the free-liquid mixture. This system resembles previously reported work on the cross-linked gel polyacrylamide in two ways: (1) As temperature is lowered toward the critical temperature of the free-liquid mixture, the binary-fluid gel exhibits a strong and increasing light scattering over a broad temperature region of several kelvins, and (2) no appreciable temporal fluctuations are observed throughout this temperature region. Two added features are observed in the present, entanglement-gel measurements: (a) Gel samples with solvent composition both near and off the critical composition of the free-liquid mixture exhibit similar light-scattering behavior, and (b) a Lorentzian-squared fit to the light-scattering angular distributions yields a characteristic wave number which does not change with temperature and an amplitude which shows a very strong dependence on the temperature.

cDNA cloning and sequencing of human fibrillarin, a conserved nucleolar protein recognized by autoimmune antisera

International Nuclear Information System (INIS)

Aris, J.P.; Blobel, G.

1991-01-01

The authors have isolated a 1.1-kilobase cDNA clone that encodes human fibrillarin by screening a hepatoma library in parallel with DNA probes derived from the fibrillarin genes of Saccharomyces cerevisiae (NOP1) and Xenopus laevis. RNA blot analysis indicates that the corresponding mRNA is ∼1,300 nucleotides in length. Human fibrillarin expressed in vitro migrates on SDS gels as a 36-kDa protein that is specifically immunoprecipitated by antisera from humans with scleroderma autoimmune disease. Human fibrillarin contains an amino-terminal repetitive domain ∼75-80 amino acids in length that is rich in glycine and arginine residues and is similar to amino-terminal domains in the yeast and Xenopus fibrillarins. The occurrence of a putative RNA-binding domain and an RNP consensus sequence within the protein is consistent with the association of fibrillarin with small nucleolar RNAs. Protein sequence alignments show that 67% of amino acids from human fibrillarin are identical to those in yeast fibrillarin and that 81% are identical to those in Xenopus fibrillarin. This identity suggests the evolutionary conservation of an important function early in the pathway for ribosome biosynthesis

Modelling the behaviour of the push-pull gel dosimeter

International Nuclear Information System (INIS)

Bosi, S.G.; Davies, J.B.; Gorjiara, T.; Baldock, C.

2010-01-01

Full text: Recent development of a gel dosimeter based on the radiobleaching pigment, genipin, allows development of a new 3D optically scanned gel dosimeter-the p ush-pull g el. This gel would contain two spectrally complementary pigments, one which darkens with dose and another (e.g. genipin) which bleaches. The two pigments deal separately with the high and low dose ends of the dosimeter's dynamic range. The bleaching pigment would be optimised for high sensitivity and the darkening pigment for low. Employing dual pigments, optimised independently, relaxes the need for compromise between sensitivity at low dose and accuracy at high dose. Such a gel, after exposure, would be read using two successive optical CT scans, at two different wavelengths. The reduction in sensitivity of the darkening pigment (allowed by the use of push-pull) would reduce the occurrence of regions of high optical attenuation which can generate optical CT artefacts. Simulated optical CT reconstructions of the optical density map (Fig. La) scanned at the darkening pigment wavelength of a hypothetical push-pull gel, confirms the reduction in susceptibility to artefacts. Fig. I b shows a profile through the map with no stray light added. The centre of the profile in Fig. I d shows a cupping artefact produced by 10 ppm of stray light. The similarity of Fig. Ic and b show that a 30% sensitivity reduction allowed by push-pull, renders the artefact negligible. This paper presents the results of' these simulations of a push-pull gel scanned using optical CT and also some results of experiments with genipin gel. (author)

Generation of a reliable full-length cDNA of infectiousTembusu virus using a PCR-based protocol.

Science.gov (United States)

Liang, Te; Liu, Xiaoxiao; Cui, Shulin; Qu, Shenghua; Wang, Dan; Liu, Ning; Wang, Fumin; Ning, Kang; Zhang, Bing; Zhang, Dabing

2016-02-02

Full-length cDNA of Tembusu virus (TMUV) cloned in a plasmid has been found instable in bacterial hosts. Using a PCR-based protocol, we generated a stable full-length cDNA of TMUV. Different cDNA fragments of TMUV were amplified by reverse transcription (RT)-PCR, and cloned into plasmids. Fragmented cDNAs were amplified and assembled by fusion PCR to produce a full-length cDNA using the recombinant plasmids as templates. Subsequently, a full-length RNA was transcribed from the full-length cDNA in vitro and transfected into BHK-21 cells; infectious viral particles were rescued successfully. Following several passages in BKH-21 cells, the rescued virus was compared with the parental virus by genetic marker checks, growth curve determinations and animal experiments. These assays clearly demonstrated the genetic and biological stabilities of the rescued virus. The present work will be useful for future investigations on the molecular mechanisms involved in replication and pathogenesis of TMUV. Copyright © 2015 Elsevier B.V. All rights reserved.

Sol-gel growth of vanadium dioxide

International Nuclear Information System (INIS)

Speck, K.R.

1990-01-01

This thesis examines the chemical reactivity of vanadium (IV) tetrakis(t-butoxide) as a precursor for the sol-gel synthesis of vanadium dioxide. Hydrolysis and condensation of the alkoxide was studied by FTIR spectroscopy. Chemical modification of the vanadium tetraalkoxide by alcohol interchange was studied using 51 V NMR and FTIR. Vanadium dioxide thin films and powders were made from vanadium tetrakis(t-butoxide) by standard sol-gel techniques. Post-deposition heating under nitrogen was necessary to transform amorphous gels into vanadium dioxide. Crystallization of films and powders was studied by FTIR, DSC, TGA, and XRD. Gel-derived vanadium dioxide films undergo a reversible semiconductor-to-metal phase transition near 68C, exhibiting characteristic resistive and spectral changes. The electrical resistance decreased by two to three orders of magnitude and the infrared transmission sharply dropped as the material was cycled through this thermally induced phase transition. The sol-gel method was also used to make doped vanadium dioxide films. Films were doped with tungsten and molybdenum ions to effectively lower the temperature at which the transition occurs

Subtractive cloning of cDNA from Aspergillus oryzae differentially regulated between solid-state culture and liquid (submerged) culture.

Science.gov (United States)

Akao, Takeshi; Gomi, Katsuya; Goto, Kuniyasu; Okazaki, Naoto; Akita, Osamu

2002-07-01

In solid-state cultures (SC), Aspergillus oryzae shows characteristics such as high-level production and secretion of enzymes and hyphal differentiation with asexual development which are absent in liquid (submerged) culture (LC). It was predicted that many of the genes involved in the characteristics of A. oryzae in SC are differentially expressed between SC and LC. We generated two subtracted cDNA libraries with bi-directional cDNA subtractive hybridizations to isolate and identify such genes. Among them, we identified genes upregulated in or specific to SC, such as the AOS ( A. oryzae SC-specific gene) series, and those downregulated or not expressed in SC, such as the AOL ( A. oryzae LC-specific) series. Sequencing analyses revealed that the AOS series and the AOL series contain genes encoding extra- and intracellular enzymes and transport proteins. However, half were functionally unclassified by nucleotide sequences. Also, by expression profile, the AOS series comprised two groups. These gene products' molecular functions and physiological roles in SC await further investigation.

An unbiased method to build benchmarking sets for ligand-based virtual screening and its application to GPCRs.

Science.gov (United States)

Xia, Jie; Jin, Hongwei; Liu, Zhenming; Zhang, Liangren; Wang, Xiang Simon

2014-05-27

Benchmarking data sets have become common in recent years for the purpose of virtual screening, though the main focus had been placed on the structure-based virtual screening (SBVS) approaches. Due to the lack of crystal structures, there is great need for unbiased benchmarking sets to evaluate various ligand-based virtual screening (LBVS) methods for important drug targets such as G protein-coupled receptors (GPCRs). To date these ready-to-apply data sets for LBVS are fairly limited, and the direct usage of benchmarking sets designed for SBVS could bring the biases to the evaluation of LBVS. Herein, we propose an unbiased method to build benchmarking sets for LBVS and validate it on a multitude of GPCRs targets. To be more specific, our methods can (1) ensure chemical diversity of ligands, (2) maintain the physicochemical similarity between ligands and decoys, (3) make the decoys dissimilar in chemical topology to all ligands to avoid false negatives, and (4) maximize spatial random distribution of ligands and decoys. We evaluated the quality of our Unbiased Ligand Set (ULS) and Unbiased Decoy Set (UDS) using three common LBVS approaches, with Leave-One-Out (LOO) Cross-Validation (CV) and a metric of average AUC of the ROC curves. Our method has greatly reduced the "artificial enrichment" and "analogue bias" of a published GPCRs benchmarking set, i.e., GPCR Ligand Library (GLL)/GPCR Decoy Database (GDD). In addition, we addressed an important issue about the ratio of decoys per ligand and found that for a range of 30 to 100 it does not affect the quality of the benchmarking set, so we kept the original ratio of 39 from the GLL/GDD.

Implementation of MRI gel dosimetry in radiation therapy

International Nuclear Information System (INIS)

Baeck, S.Aa.J.

1998-12-01

Gel dosimetry was used together with magnetic resonance imaging (MRI) to measure three-dimensional absorbed dose distributions in radiation therapy. Two different dosimeters were studied: ferrous- and monomer gel, based on the principles of radiation-induced oxidation and polymerisation, respectively. Single clinical electron and photon beams were evaluated and gel dose distributions were mainly within 2% of conventional detector results. The ferrous-gel was also used for clinical proton beams. A decrease in signal per absorbed dose was found close to the end of the range of the protons (15-20%). This effect was explained as a linear energy transfer dependence, further supported with Monte Carlo simulations. A method for analysing and comparing data from treatment planning system (TPS) and gel measurements was developed. The method enables a new pixel by pixel evaluation, isodose comparison and dose volume histogram verification. Two standard clinical radiation therapy procedures were examined using the developed TPS verification method. The treatment regimes included several beams of different radiation qualities. The TPS calculated data were in very good agreement with the dose distribution measured by the ferrous-gel. However, in a beam abutment region, larger dose difference was found. Beam adjustment errors and a minor TPS underestimation of the lateral scatter contribution outside the primary electron beam may explain the discrepancy. The overall uncertainty in the ferrous-gel dose determination was considerably reduced using an optimised MRI acquisition protocol and a new MRI scanner. The relative dose uncertainty was found to be better than 3.3% for all dose levels (95% confidence level). Using the method developed for comparing measured gel data with calculated treatment plans, the gel dosimetry method was proven to be a useful tool for radiation treatment planning verification

Implementation of MRI gel dosimetry in radiation therapy

Energy Technology Data Exchange (ETDEWEB)

Baeck, S.Aa.J

1998-12-01

Gel dosimetry was used together with magnetic resonance imaging (MRI) to measure three-dimensional absorbed dose distributions in radiation therapy. Two different dosimeters were studied: ferrous- and monomer gel, based on the principles of radiation-induced oxidation and polymerisation, respectively. Single clinical electron and photon beams were evaluated and gel dose distributions were mainly within 2% of conventional detector results. The ferrous-gel was also used for clinical proton beams. A decrease in signal per absorbed dose was found close to the end of the range of the protons (15-20%). This effect was explained as a linear energy transfer dependence, further supported with Monte Carlo simulations. A method for analysing and comparing data from treatment planning system (TPS) and gel measurements was developed. The method enables a new pixel by pixel evaluation, isodose comparison and dose volume histogram verification. Two standard clinical radiation therapy procedures were examined using the developed TPS verification method. The treatment regimes included several beams of different radiation qualities. The TPS calculated data were in very good agreement with the dose distribution measured by the ferrous-gel. However, in a beam abutment region, larger dose difference was found. Beam adjustment errors and a minor TPS underestimation of the lateral scatter contribution outside the primary electron beam may explain the discrepancy. The overall uncertainty in the ferrous-gel dose determination was considerably reduced using an optimised MRI acquisition protocol and a new MRI scanner. The relative dose uncertainty was found to be better than 3.3% for all dose levels (95% confidence level). Using the method developed for comparing measured gel data with calculated treatment plans, the gel dosimetry method was proven to be a useful tool for radiation treatment planning verification 103 refs, 20 figs, 6 tabs

Isolation and characterization of cDNA clones for human erythrocyte β-spectrin

International Nuclear Information System (INIS)

Prchal, J.T.; Morley, B.J.; Yoon, S.H.; Coetzer, T.L.; Palek, J.; Conboy, J.G.; Kan, Y.W.

1987-01-01

Spectrin is an important structural component of the membrane skeleton that underlies and supports the erythrocyte plasma membrane. It is composed of nonidentical α (M/sub r/ 240,000) and β (M/sub r/ 220,000) subunits, each of which contains multiple homologous 106-amino acid segments. The authors report here the isolation and characterization of a human erythroid-specific β-spectrin cDNA clone that encodes parts of the β-9 through β-12 repeat segments. This cDNA was used as a hybridization probe to assign the β-spectrin gene to human chromosome 14 and to begin molecular analysis of the gene and its mRNA transcripts. RNA transfer blot analysis showed that the reticulocyte β-spectrin mRNA is 7.8 kilobases in length. Southern blot analysis of genomic DNA revealed the presence of restriction fragment length polymorphisms (RFLPs) within the β-spectrin gene locus. The isolation of human spectrin cDNA probes and the identification of closely linked RFLPs will facilitate analysis of mutant spectrin genes causing congenital hemolytic anemias associated with quantitative and qualitative spectrin abnormalities

Expression of wild-type and mutant medium-chain acyl-CoA dehydrogenase (MCAD) cDNA in eucaryotic cells

DEFF Research Database (Denmark)

Jensen, T G; Andresen, B S; Bross, P

1992-01-01

An effective EBV-based expression system for eucaryotic cells has been developed and used for the study of the mitochondrial enzyme medium-chain acyl-CoA dehydrogenase (MCAD). 1325 bp of PCR-generated MCAD cDNA, containing the entire coding region, was placed between the SV40 early promoter...... and polyadenylation signals in the EBV-based vector. Both wild-type MCAD cDNA and cDNA containing the prevalent disease-causing mutation A to G at position 985 of the MCAD cDNA were tested. In transfected COS-7 cells, the steady state amount of mutant MCAD protein was consistently lower than the amount of wild......-type human enzyme. The enzyme activity in extracts from cells harbouring the wild-type MCAD cDNA was dramatically higher than in the controls (harbouring the vector without the MCAD gene) while only a slightly higher activity was measured with the mutant MCAD. The mutant MCAD present behaves like wild...

Encapsulation of biological species in sol-gel matrices

International Nuclear Information System (INIS)

Finnie, K.S.; Bartlett, J.R.; Woolfrey, J.L.

2000-01-01

Two examples are given of the gelation of silica sols containing bio catalysts, resulting in their encapsulation in porous matrices. Urease was encapsulated in gels made from a mixture of TMOS and alkyltrimethoxysilane. Enzyme activities, monitored by measuring the rate of production of ammoniacal nitrogen as urea was decomposed, ranged up to 60% of that of the unencapsulated species. Anaerobic sulphate-reducing bacteria were encapsulated in a gel produced from colloidal silica, thus avoiding contact with alcohol. The detection of H 2 S produced in the doped gel indicated that the bacteria were able to continue normal metabolic function within the gel matrix. A gel initially doped with ∼ 5 x 10 5 cells cm -3 , exhibited an optimum sulphate reduction rate of 11 ug h -1 cm -3 ; this reduction rate was quickly re-established after storage of the gel for 14 weeks. Copyright (2000) The Australian Ceramic Society

An Unbiased Distance-based Outlier Detection Approach for High-dimensional Data

DEFF Research Database (Denmark)

Nguyen, Hoang Vu; Gopalkrishnan, Vivekanand; Assent, Ira

2011-01-01

than a global property. Different from existing approaches, it is not grid-based and dimensionality unbiased. Thus, its performance is impervious to grid resolution as well as the curse of dimensionality. In addition, our approach ranks the outliers, allowing users to select the number of desired...... outliers, thus mitigating the issue of high false alarm rate. Extensive empirical studies on real datasets show that our approach efficiently and effectively detects outliers, even in high-dimensional spaces....

Monofunctional stealth nanoparticle for unbiased single molecule tracking inside living cells.

Science.gov (United States)

Lisse, Domenik; Richter, Christian P; Drees, Christoph; Birkholz, Oliver; You, Changjiang; Rampazzo, Enrico; Piehler, Jacob

2014-01-01

On the basis of a protein cage scaffold, we have systematically explored intracellular application of nanoparticles for single molecule studies and discovered that recognition by the autophagy machinery plays a key role for rapid metabolism in the cytosol. Intracellular stealth nanoparticles were achieved by heavy surface PEGylation. By combination with a generic approach for nanoparticle monofunctionalization, efficient labeling of intracellular proteins with high fidelity was accomplished, allowing unbiased long-term tracking of proteins in the outer mitochondrial membrane.

Enhanced detergent extraction for analysis of membrane proteomes by two-dimensional gel electrophoresis

Directory of Open Access Journals (Sweden)

Hsu Kimberly K

2005-06-01

Full Text Available Abstract Background The analysis of hydrophobic membrane proteins by two-dimensional gel electrophoresis has long been hampered by the concept of inherent difficulty due to solubility issues. We have optimized extraction protocols by varying the detergent composition of the solubilization buffer with a variety of commercially available non-ionic and zwitterionic detergents and detergent-like phospholipids. Results After initial analyses by one-dimensional SDS-PAGE, quantitative two-dimensional analyses of human erythrocyte membranes, mouse liver membranes, and mouse brain membranes, extracted with buffers that included the zwitterionic detergent MEGA 10 (decanoyl-N-methylglucamide and the zwitterionic lipid LPC (1-lauroyl lysophosphatidylcholine, showed selective improvement over extraction with the common 2-DE detergent CHAPS (3 [(3-cholamidopropyldimethylammonio]-1-propanesulfonate. Mixtures of the three detergents showed additive improvements in spot number, density, and resolution. Substantial improvements in the analysis of a brain membrane proteome were observed. Conclusion This study demonstrates that an optimized detergent mix, coupled with rigorous sample handling and electrophoretic protocols, enables simple and effective analysis of membrane proteomes using two-dimensional electrophoresis.

Cloning a cDNA for the lysosomal alpha-glucosidase

NARCIS (Netherlands)

KONINGS, A.; HUPKES, P.; Versteeg, R.; Grosveld, G.; Reuser, A.; Galjaard, H.

1984-01-01

Messenger RNA was isolated from monkey testes and size-fractionated on sucrose gradients. In vitro translation of these mRNA fractions resulted in nascent, labeled alpha-glucosidase that could be precipitated with anti human alpha-glucosidase antiserum. A cDNA library was constructed from the most

Importance of the efficiency of double-stranded DNA formation in cDNA synthesis for the imprecision of microarray expression analysis.

Science.gov (United States)

Thormar, Hans G; Gudmundsson, Bjarki; Eiriksdottir, Freyja; Kil, Siyoen; Gunnarsson, Gudmundur H; Magnusson, Magnus Karl; Hsu, Jason C; Jonsson, Jon J

2013-04-01

The causes of imprecision in microarray expression analysis are poorly understood, limiting the use of this technology in molecular diagnostics. Two-dimensional strandness-dependent electrophoresis (2D-SDE) separates nucleic acid molecules on the basis of length and strandness, i.e., double-stranded DNA (dsDNA), single-stranded DNA (ssDNA), and RNA·DNA hybrids. We used 2D-SDE to measure the efficiency of cDNA synthesis and its importance for the imprecision of an in vitro transcription-based microarray expression analysis. The relative amount of double-stranded cDNA formed in replicate experiments that used the same RNA sample template was highly variable, ranging between 0% and 72% of the total DNA. Microarray experiments showed an inverse relationship between the difference between sample pairs in probe variance and the relative amount of dsDNA. Approximately 15% of probes showed between-sample variation (P cDNA synthesized can be an important component of the imprecision in T7 RNA polymerase-based microarray expression analysis. © 2013 American Association for Clinical Chemistry

Identification and Molecular Characterization of the cDNA Encoding Cucumis melo Allergen, Cuc m 3, a Plant Pathogenesis-Related Protein

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Mojtaba Sankian

2014-05-01

Full Text Available Background: Melon (Cucumis melo allergy is one of the most common food allergies, characterized by oral allergy syndrome. To date, two allergen molecules, Cuc m 1 and Cuc m 2, have been fully characterized in melon pulp, but there are few reports about the molecular characteristics of Cuc m 3. Methods:The Cuc m 3 cDNA has been characterized by rapid amplification of cDNA ends (RACE, which revealed a 456 base-pair (bp fragment encoding a 151-amino acid polypeptide with a predicted molecular mass of 16.97 kDa, and identified 79 and 178 bp untranslated sequences at the 5′ and 3´ ends, respectively. Results: In silico analysis showed strong similarities between Cuc m 3 and other plant pathogen-related protein 1s from cucumber, grape, bell pepper, and tomato. Conclusion: Here we report the identification and characterization of the Cuc m 3 cDNA, which will be utilized for further analyses of structural and allergenic features of this allergen

Multiplex cDNA quantification method that facilitates the standardization of gene expression data

Science.gov (United States)

Gotoh, Osamu; Murakami, Yasufumi; Suyama, Akira

2011-01-01

Microarray-based gene expression measurement is one of the major methods for transcriptome analysis. However, current microarray data are substantially affected by microarray platforms and RNA references because of the microarray method can provide merely the relative amounts of gene expression levels. Therefore, valid comparisons of the microarray data require standardized platforms, internal and/or external controls and complicated normalizations. These requirements impose limitations on the extensive comparison of gene expression data. Here, we report an effective approach to removing the unfavorable limitations by measuring the absolute amounts of gene expression levels on common DNA microarrays. We have developed a multiplex cDNA quantification method called GEP-DEAN (Gene expression profiling by DCN-encoding-based analysis). The method was validated by using chemically synthesized DNA strands of known quantities and cDNA samples prepared from mouse liver, demonstrating that the absolute amounts of cDNA strands were successfully measured with a sensitivity of 18 zmol in a highly multiplexed manner in 7 h. PMID:21415008

cDNA cloning, sequence analysis, and chromosomal localization of the gene for human carnitine palmitoyltransferase

International Nuclear Information System (INIS)

Finocchiaro, G.; Taroni, F.; Martin, A.L.; Colombo, I.; Tarelli, G.T.; DiDonato, S.; Rocchi, M.

1991-01-01

The authors have cloned and sequenced a cDNA encoding human liver carnitine palmitoyltransferase an inner mitochondrial membrane enzyme that plays a major role in the fatty acid oxidation pathway. Mixed oligonucleotide primers whose sequences were deduced from one tryptic peptide obtained from purified CPTase were used in a polymerase chain reaction, allowing the amplification of a 0.12-kilobase fragment of human genomic DNA encoding such a peptide. A 60-base-pair (bp) oligonucleotide synthesized on the basis of the sequence from this fragment was used for the screening of a cDNA library from human liver and hybridized to a cDNA insert of 2255 bp. This cDNA contains an open reading frame of 1974 bp that encodes a protein of 658 amino acid residues including 25 residues of an NH 2 -terminal leader peptide. The assignment of this open reading frame to human liver CPTase is confirmed by matches to seven different amino acid sequences of tryptic peptides derived from pure human CPTase and by the 82.2% homology with the amino acid sequence of rat CPTase. The NH 2 -terminal region of CPTase contains a leucine-proline motif that is shared by carnitine acetyl- and octanoyltransferases and by choline acetyltransferase. The gene encoding CPTase was assigned to human chromosome 1, region 1q12-1pter, by hybridization of CPTase cDNA with a DNA panel of 19 human-hanster somatic cell hybrids

Improving calibration accuracy in gel dosimetry

International Nuclear Information System (INIS)

Oldham, M.; McJury, M.; Webb, S.; Baustert, I.B.; Leach, M.O.

1998-01-01

A new method of calibrating gel dosimeters (applicable to both Fricke and polyacrylamide gels) is presented which has intrinsically higher accuracy than current methods, and requires less gel. Two test-tubes of gel (inner diameter 2.5 cm, length 20 cm) are irradiated separately with a 10x10cm 2 field end-on in a water bath, such that the characteristic depth-dose curve is recorded in the gel. The calibration is then determined by fitting the depth-dose measured in water, against the measured change in relaxivity with depth in the gel. Increased accuracy is achieved in this simple depth-dose geometry by averaging the relaxivity at each depth. A large number of calibration data points, each with relatively high accuracy, are obtained. Calibration data over the full range of dose (1.6-10 Gy) is obtained by irradiating one test-tube to 10 Gy at dose maximum (D max ), and the other to 4.5 Gy at D max . The new calibration method is compared with a 'standard method' where five identical test-tubes of gel were irradiated to different known doses between 2 and 10 Gy. The percentage uncertainties in the slope and intercept of the calibration fit are found to be lower with the new method by a factor of about 4 and 10 respectively, when compared with the standard method and with published values. The gel was found to respond linearly within the error bars up to doses of 7 Gy, with a slope of 0.233±0.001 s -1 Gy -1 and an intercept of 1.106±0.005 Gy. For higher doses, nonlinear behaviour was observed. (author)

Dissection of the inflammatory bowel disease transcriptome using genome-wide cDNA microarrays.

Directory of Open Access Journals (Sweden)

Christine M Costello

2005-08-01

Full Text Available BACKGROUND: The differential pathophysiologic mechanisms that trigger and maintain the two forms of inflammatory bowel disease (IBD, Crohn disease (CD, and ulcerative colitis (UC are only partially understood. cDNA microarrays can be used to decipher gene regulation events at a genome-wide level and to identify novel unknown genes that might be involved in perpetuating inflammatory disease progression. METHODS AND FINDINGS: High-density cDNA microarrays representing 33,792 UniGene clusters were prepared. Biopsies were taken from the sigmoid colon of normal controls (n = 11, CD patients (n = 10 and UC patients (n = 10. 33P-radiolabeled cDNA from purified poly(A+ RNA extracted from biopsies (unpooled was hybridized to the arrays. We identified 500 and 272 transcripts differentially regulated in CD and UC, respectively. Interesting hits were independently verified by real-time PCR in a second sample of 100 individuals, and immunohistochemistry was used for exemplary localization. The main findings point to novel molecules important in abnormal immune regulation and the highly disturbed cell biology of colonic epithelial cells in IBD pathogenesis, e.g., CYLD (cylindromatosis, turban tumor syndrome and CDH11 (cadherin 11, type 2. By the nature of the array setup, many of the genes identified were to our knowledge previously uncharacterized, and prediction of the putative function of a subsection of these genes indicate that some could be involved in early events in disease pathophysiology. CONCLUSION: A comprehensive set of candidate genes not previously associated with IBD was revealed, which underlines the polygenic and complex nature of the disease. It points out substantial differences in pathophysiology between CD and UC. The multiple unknown genes identified may stimulate new research in the fields of barrier mechanisms and cell signalling in the context of IBD, and ultimately new therapeutic approaches.

Critical point relascope sampling for unbiased volume estimation of downed coarse woody debris

Science.gov (United States)

Jeffrey H. Gove; Michael S. Williams; Mark J. Ducey; Mark J. Ducey

2005-01-01

Critical point relascope sampling is developed and shown to be design-unbiased for the estimation of log volume when used with point relascope sampling for downed coarse woody debris. The method is closely related to critical height sampling for standing trees when trees are first sampled with a wedge prism. Three alternative protocols for determining the critical...

Behavior of water gel permissibles in coal mines

Energy Technology Data Exchange (ETDEWEB)

Ribovich, J.; Watson, R.W.

1979-01-01

This paper presents an overview of the successful use of water gel permissibles, since their introduction in 1970, with an emphasis on some problems of behavior in their field application. One is the metal clip or ties used to seal the water gel package. Miners are occasionally hit by the 2.4 mm metal wire expelled from boreholes in normal shooting operations. Manufacturers have consequently reduced the mass of the metal ties by 25-50%, providing a partial solution. Another problem concerned two reported instances of misfired water gel burning after coal shooting operations. Extensive tests have offered no satisfactory solution as yet, except to point up a principal shortcoming of water gels: that of pressure desensitization. Two other factors affecting sensitivity, and possibly resulting in misfires, are low temperature and aging. There is a decline in air-gap sensitivity with prolonged storage but all approved water gels seem capable of meeting requirements for long periods under normal storage conditions. Low temperature tests indicate the majority of water gels become insensitive to a No. 6 EBC at temperatures easily attained in cold weather storage. An attempt was made to determine the minimum temperature at which a nitroglycerin-sensitized permisslbe could be initiated with a No. 8 cap; a particularly granular one was initiated as low as minus 196 C. (5 refs.) (In English)

Analysis of glycoprotein-derived oligosaccharides in glycoproteins detected on two-dimensional gel by capillary electrophoresis using on-line concentration method.

Science.gov (United States)

Kamoda, Satoru; Nakanishi, Yasuharu; Kinoshita, Mitsuhiro; Ishikawa, Rika; Kakehi, Kazuaki

2006-02-17

Capillary electrophoresis (CE) is an effective tool to analyze carbohydrate mixture derived from glycoproteins with high resolution. However, CE has a disadvantage that a few nanoliters of a sample solution are injected to a narrow capillary. Therefore, we have to prepare a sample solution of high concentration for CE analysis. In the present study, we applied head column field-amplified sample stacking method to the analysis of N-linked oligosaccharides derived from glycoprotein separated by two-dimensional gel electrophoresis. Model studies demonstrated that we achieved 60-360 times concentration effect on the analysis of carbohydrate chains labeled with 3-aminobenzoic acid (3-AA). The method was applied to the analysis of N-linked oligosaccharides from glycoproteins separated and detected on PAGE gel. Heterogeneity of alpha1-acid glycoprotein (AGP), i.e. glycoforms, was examined by 2D-PAGE and N-linked oligosaccharides were released by in-gel digestion with PNGase F. The released oligosaccharides were derivatized with 3-AA and analyzed by CE. The results showed that glycoforms having lower pI values contained a larger amount of tetra- and tri-antennary oligosaccharides. In contrast, glycoforms having higher pI values contained bi-antennary oligosaccharides abundantly. The result clearly indicated that the spot of a glycoprotein glycoform detected by Coomassie brilliant blue staining on 2D-PAGE gel is sufficient for quantitative profiling of oligosaccharides.

Identification of immune protective genes of Eimeria maxima through cDNA expression library screening.

Science.gov (United States)

Yang, XinChao; Li, MengHui; Liu, JianHua; Ji, YiHong; Li, XiangRui; Xu, LiXin; Yan, RuoFeng; Song, XiaoKai

2017-02-16

Eimeria maxima is one of the most prevalent Eimeria species causing avian coccidiosis, and results in huge economic loss to the global poultry industry. Current control strategies, such as anti-coccidial medication and live vaccines have been limited because of their drawbacks. The third generation anticoccidial vaccines including the recombinant vaccines as well as DNA vaccines have been suggested as a promising alternative strategy. To date, only a few protective antigens of E. maxima have been reported. Hence, there is an urgent need to identify novel protective antigens of E. maxima for the development of neotype anticoccidial vaccines. With the aim of identifying novel protective genes of E. maxima, a cDNA expression library of E. maxima sporozoites was constructed using Gateway technology. Subsequently, the cDNA expression library was divided into 15 sub-libraries for cDNA expression library immunization (cDELI) using parasite challenged model in chickens. Protective sub-libraries were selected for the next round of screening until individual protective clones were obtained, which were further sequenced and analyzed. Adopting the Gateway technology, a high-quality entry library was constructed, containing 9.2 × 10 6 clones with an average inserted fragments length of 1.63 kb. The expression library capacity was 2.32 × 10 7 colony-forming units (cfu) with an average inserted fragments length of 1.64 Kb. The expression library was screened using parasite challenged model in chickens. The screening yielded 6 immune protective genes including four novel protective genes of EmJS-1, EmRP, EmHP-1 and EmHP-2, and two known protective genes of EmSAG and EmCKRS. EmJS-1 is the selR domain-containing protein of E. maxima whose function is unknown. EmHP-1 and EmHP-2 are the hypothetical proteins of E. maxima. EmRP and EmSAG are rhomboid-like protein and surface antigen glycoproteins of E. maxima respectively, and involved in invasion of the parasite. Our

Rethinking economy-wide rebound measures: An unbiased proposal

International Nuclear Information System (INIS)

Guerra, Ana-Isabel; Sancho, Ferran

2010-01-01

In spite of having been first introduced in the last half of the ninetieth century, the debate about the possible rebound effects from energy efficiency improvements is still an open question in the economic literature. This paper contributes to the existing research on this issue proposing an unbiased measure for economy-wide rebound effects. The novelty of this economy-wide rebound measure stems from the fact that not only actual energy savings but also potential energy savings are quantified under general equilibrium conditions. Our findings indicate that the use of engineering savings instead of general equilibrium potential savings downward biases economy-wide rebound effects and upward-biases backfire effects. The discrepancies between the traditional indicator and our proposed measure are analysed in the context of the Spanish economy.

Modelling of the inhomogeneous interior of polymer gels

International Nuclear Information System (INIS)

Shew, C-Y; Iwaki, Takafumi

2006-01-01

A simple model has been investigated to elucidate the mean squared displacement (MSD) of probe molecules in cross-linked polymer gels. In the model, we assume that numerous cavities distribute in the inhomogeneous interior of a gel, and probe molecules are confined within these cavities. The individual probe molecules trapped in a gel are treated as Brownian particles confined to a spherical harmonic potential. The harmonic potential is chosen to model the effective potential experienced by the probe particle in the cavity of a gel. Each field strength is corresponding to the characteristic of one type of effective cavity. Since the statistical distribution of different effective cavity sizes is unknown, several distribution functions are examined. Meanwhile, the calculated averaged MSDs are compared to the experimental data by Nisato et al (2000 Phys. Rev. E 61 2879). We find that the theoretical results of the MSD are sensitive to the shape of the distribution function. For low cross-linked gels, the best fit is obtained when the interior cavities of a gel follow a bimodal distribution. Such a result may be attributed to the presence of at least two distinct classes of cavity in gels. For high cross-linked gels, the cavities in the gel can be depicted by a single-modal uniform distribution function, suggesting that the range of cavity sizes becomes smaller. These results manifest the voids inside a gel, and the shape of distribution functions may provide the insight into the inhomogeneous interior of a gel

Applications and Properties of Ionic Liquid- Based Gels and Soft Solid Composites

Science.gov (United States)

Voss, Bret Alan McGinness

2011-12-01

Solid-liquid composites (gels) have a combination of properties that afford new material applications in which high solute diffusion is desirable. These composites have a soft-solid mechanical integrity and will not flow under gravity, but entrain a liquid matrix (i.e. 60-98 mass %) which allows for high diffusion and high reactivity. Room temperature ionic liquid (RTILs) are molten organic salts with a melting point below room temperature and negligible vapor pressure. If the RTILs are used as the liquid component of a gel, then the gel matrix will not evaporate (unlike other organic solvents) and may be used for long term applications. This thesis research applies RTIL gels for two new applications; carbon dioxide/nitrogen separation and chemical warfare agent (CWA) barrier and decontamination. Separating CO2 from the flue gas of coal and gas fired power-plants is an increasingly economically and environmentally important gas separation. In this first study, RTIL gels are cast in a supported membrane and gas permeability and ideal selectivity are measured. The RTIL matrix has an inherent affinity for CO2 and provides a high diffusion, hence high permeability (i.e. 500-700 barrer). The solidifying component is a low molecular-weight organic gelator (LMOG) which through physical bonding interactions (i.e. hydrogen bonding, pi-pi stacking and van der Walls forces) forms an entangled network which provides mechanical stability (i.e. increase trans-membrane pressure required to expel selective material from the support). In these studies two LMOGs and five RTILs are used to make supported gel membranes and determine gas permeability and temperature dependent trends. The second application for RTIL gels is a decontaminating barrier for CWAs and toxic industrial compounds (TICs). In these studies a layer of RTIL gel is applied on top of a substrate contaminated with a CWA simulant (i.e. chloroethylethylsulfide, CEES). The gel performs well as a barrier, preventing CEES

Cloning and sequencing of cDNA encoding human DNA topoisomerase II and localization of the gene to chromosome region 17q21-22

International Nuclear Information System (INIS)

Tsai-Pflugfelder, M.; Liu, L.F.; Liu, A.A.; Tewey, K.M.; Whang-Peng, J.; Knutsen, T.; Huebner, K.; Croce, C.M.; Wang, J.C.

1988-01-01

Two overlapping cDNA clones encoding human DNA topoisomerase II were identified by two independent methods. In one, a human cDNA library in phage λ was screened by hybridization with a mixed oligonucleotide probe encoding a stretch of seven amino acids found in yeast and Drosophila DNA topoisomerase II; in the other, a different human cDNA library in a λgt11 expression vector was screened for the expression of antigenic determinants that are recognized by rabbit antibodies specific to human DNA topoisomerase II. The entire coding sequences of the human DNA topoisomerase II gene were determined from these and several additional clones, identified through the use of the cloned human TOP2 gene sequences as probes. Hybridization between the cloned sequences and mRNA and genomic DNA indicates that the human enzyme is encoded by a single-copy gene. The location of the gene was mapped to chromosome 17q21-22 by in situ hybridization of a cloned fragment to metaphase chromosomes and by hybridization analysis with a panel of mouse-human hybrid cell lines, each retaining a subset of human chromosomes

Investigation of two different anoxia models by 2-dimensional gel electrophoresis

DEFF Research Database (Denmark)

Wulff, Tune; Jessen, Flemming; Hoffmann, Else Kay

2006-01-01

anoxia obtained by NaN3 is a widely used model for simulating anoxia (Ossum et al., 2004). The effects of anoxia were studied by protein expression analysis using 2-dimensional gel electrophoresis followed by MS/MS. In this way we were able to separate more than 1500 protein spots with an apparent range...

Monoterpene engineering in a woody plant Eucalyptus camaldulensis using a limonene synthase cDNA.

Science.gov (United States)

Ohara, Kazuaki; Matsunaga, Etsuko; Nanto, Kazuya; Yamamoto, Kyoko; Sasaki, Kanako; Ebinuma, Hiroyasu; Yazaki, Kazufumi

2010-01-01

Metabolic engineering aimed at monoterpene production has become an intensive research topic in recent years, although most studies have been limited to herbal plants including model plants such as Arabidopsis. The genus Eucalyptus includes commercially important woody plants in terms of essential oil production and the pulp industry. This study attempted to modify the production of monoterpenes, which are major components of Eucalyptus essential oil, by introducing two expression constructs containing Perilla frutescens limonene synthase (PFLS) cDNA, whose gene products were designed to be localized in either the plastid or cytosol, into Eucalyptus camaldulensis. The expression of the plastid-type and cytosol-type PFLS cDNA in transgenic E. camaldulensis was confirmed by real-time polymerase chain reaction (PCR). Gas chromatography with a flame ionization detector analyses of leaf extracts revealed that the plastidic and cytosolic expression of PFLS yielded 2.6- and 4.5-times more limonene than that accumulated in wild-type E. camaldulensis, respectively, while the ectopic expression of PFLS had only a small effect on the emission of limonene from the leaves of E. camaldulensis. Surprisingly, the high level of PFLS in Eucalyptus was accompanied by a synergistic increase in the production of 1,8-cineole and alpha-pinene, two major components of Eucalyptus monoterpenes. This genetic engineering of monoterpenes demonstrated a new potential for molecular breeding in woody plants.

Characterization of the cDNA encoding human nucleophosmin and studies of its role in normal and abnormal growth

International Nuclear Information System (INIS)

Chan, Waiyee; Liu, Qingrong; Borjigin, J.; Busch, H.; Rennert, O.M.; Tease, L.A.; Chan, Puikwong

1989-01-01

A cDNA encoding human nucleophosmin (protein B23) was obtained by screening a human placental cDNA library in δgtll first with monoclonal antibody to rat nucleophosmin and then with confirmed partial cDNA of human nucleophosmin as probes. The cDNA had 1,311 bp with a coding sequence encoding a protein of 294 amino acids. The identity of the cDNA was confirmed by the presence of encoded amino acid sequences identical with those determined by sequencing pure rat nucleophosmin (a total of 138 amino acids). The most striking feature of the sequence is an acidic cluster located in the middle of the molecule. The cluster consists of 26 Asp/Glu and 1 Phe and Ala. Comparison of human nucleophosmin and Xenopus nucleolar protein NO38 shows 64.3% sequence identity. The N-terminal 130 amino acids of human nucleophosmin also bear 50% identity with that of Xenopus nucleoplasmin. Northern blot analysis of rat liver total RNA with a partial nucleophosmin cDNA as probe demonstrated a homogeneous mRNA band of about 1.6 kb. Similar observations were made in hypertrophic rat liver and Novikoff hepatoma. When the protein levels were compared with Western blot immunoassays, Navikoff hepatoma showed 20 times more nucleophosmin, while only about 5 times more nucleophosmin was observed in hypertrophic rat liver than in unstimulated normal liver

Sequence Dependent Electrophoretic Separations of DNA in Pluronic F127 Gels

Science.gov (United States)

You, Seungyong; van Winkle, David H.

2010-03-01

Two-dimensional (2-D) electrophoresis has successfully been used to visualize the separation of DNA fragments of the same length. We electrophorese a double-stranded DNA ladder in an Agarose gel for the first dimension and in gels of Pluronic F127 for the second dimension at room temperature. The 1000 bp band that travels together as a single band in an Agarose gel is split into two bands in Pluronic gels. The slower band follows the exponential decay trend that the other ladder constituents do. After sequencing the DNA fragments, the faster band has an apparently random sequence, while the slower band and the others have two A-tracts in each 250 bp segment. The A-tracts consist of a series of at least five adenine bases pairing with thymine bases. This result leads to the conclusion that the migration of the DNA molecules bent with A-tracts is more retarded in Pluronic gels than the wild-type of DNA molecules.

The New Peabody Picture Vocabulary Test-III: An Illusion of Unbiased Assessment?

Science.gov (United States)

Stockman, Ida J

2000-10-01

This article examines whether changes in the ethnic minority composition of the standardization sample for the latest edition of the Peabody Picture Vocabulary Test (PPVT-III, Dunn & Dunn, 1997) can be used as the sole explanation for children's better test scores when compared to an earlier edition, the Peabody Picture Vocabulary Test-Revised (PPVT-R, Dunn & Dunn, 1981). Results from a comparative analysis of these two test editions suggest that other factors may explain improved performances. Among these factors are the number of words and age levels sampled, the types of words and pictures used, and characteristics of the standardization sample other than its ethnic minority composition. This analysis also raises questions regarding the usefulness of converting scores from one edition to the other and the type of criteria that could be used to evaluate whether the PPVT-III is an unbiased test of vocabulary for children from diverse cultural and linguistic backgrounds.

cDNA, genomic cloning and sequence analysis of ribosomal protein ...

African Journals Online (AJOL)

enoh

2012-03-13

Mar 13, 2012 ... cDNA and the genomic sequence of RPS4X were cloned successfully from ... S4 genes plays a role in Turner syndrome; however, this ..... Project of Educational Committee of Sichuan Province ... Molecular biology of the cell.

Thorium inorganic gels

International Nuclear Information System (INIS)

Genet, M.; Brandel, V.

1988-01-01

The optimum pH and concentration values of thorium salts and oxoacids or oxoacid salts which lead to transparent and stable inorganic gels have been determined. The isotherm drying process of the gel at 50 0 C leads successively to a partly dehydrated gel, then, to the formation of an unusual liquid phase and, finally to a dry amorphous solid phase which is still transparent. This kind of transparent inorganic gels and amorphous phase can be used as matrices for spectroscopic studies [fr

MPO cDNA clone identifies an RFLP with PstI

Energy Technology Data Exchange (ETDEWEB)

Miki, T; Weil, S C; Rosner, G L; Reid, M S; Kidd, K K

1988-02-25

A myeloperoxidase (MPO) cDNA clone (pHMP7: 270 base pair insert in the vector pGEM-1reverse arrow was isolated from a library created from human promyelocytic (HL-60) cell mRNA. PstI (CTGCA/G) (New England Biolabs) identifies a simple two-allele polymorphism with bands at either 2.2 kb (Al) or 2.0 kb (A2). There are three constant bands at 2.8 kb, 0.95 kb and 0.6 kb. Preliminary family data show evidence of linkage to several markers in proximal 17q, with MPO closest to the Growth Hormone cluster at 17q22-q24. Autosomal condominant segregation was observed in four large reference pedigrees with several informative matings.

The micropolyurethane foam-coated Diagon/Gel4Two implant in aesthetic and reconstructive breast surgery – 3-year results of an ongoing study

Directory of Open Access Journals (Sweden)

Brunnert, Klaus E.

2015-12-01

Full Text Available Background: Breast implants are worldwide in use since 1962. Initially there were some problems with capsular contracture and the palpability of the rim of the implant. In 1968 this led to the introduction of the micropolyurethane foam-coating and then in 1970 to the first micropolyurethane foam-coated implant by F.A. Ashley. As a result of additional technical refinements in manufacturing this new implant design significantly reduced complications i.e. capsular contracture and implant rotation. Methods: This study reports a single surgeon’s experience with aesthetic and reconstructive breast surgery, in primary and secondary cases with the sole use of micropolyurethane foam-coated Diagon/gel4Two implants, partly in combination with the additional use of synthetic meshes, acellular dermal matrices and lipofilling. The trial is a prospective, single center cohort study designed to demonstrate the safety and effectiveness of the new implant design in primary and secondary aesthetic and reconstructive breast surgery. The reported data provide an interim report of the implantations performed from November 2010 to December 2013.Results: 90 patients were admitted to the study with 152 implants. The majority of the implants (n=95, 62.5% were used in reoperative cases for either oncological (n=52, 34.2% or aesthetic reasons (n=43, 28.3%. The median age of the study cohort was 45 years; the median body mass index was 21; the median observation time is 41 months. There was a very low complication rate, both short term within 6 weeks after the implantation of the silicone gel implant and in the follow up in November 2015. There were no serious complications needing explantation, no capsular fibrosis or implant rotation or rupture so far. There were only 4 minor complications (1.97%. There was 1 local recurrence 4 years after skin and nipple sparing mastectomy.Conclusion: The micropolyurethane foam-coated Diagon/gel4Two implant is a very reliable silicone

An Unbiased Survey of 500 Nearby Stars for Debris Disks: A JCMT Legacy Program

NARCIS (Netherlands)

Matthews, B.C.; Greaves, J.S.; Holland, W.S.; Wyatt, M.C.; Barlow, M.J.; Bastien, P.; Beichman, C.A.; Biggs, A.; Butner, H.M.; Dent, W.R.F.; Francesco, J. Di; Dominik, C.; Fissel, L.; Friberg, P.; Gibb, A.G.; Halpern, M.; Ivison, R.J.; Jayawardhana, R.; Jenness, T.; Johnstone, D.; Kavelaars, J.J.; Marshall, J.L.; Phillips, N.; Schieven, G.; Snellen, I.A.G.; Walker, H.J.; Ward-Thompson, D.; Weferling, B.; White, G.J.; Yates, J.; Zhu, M.; Craigon, A.

2007-01-01

We present the scientific motivation and observing plan for an upcoming detection survey for debris disks using the James Clerk Maxwell Telescope. The SCUBA-2 Unbiased Nearby Stars (SUNS) survey will observe 500 nearby main-sequence and subgiant stars (100 of each of the A, F, G, K, and M spectral

Ordering phenomena in ABA triblock copolymer gels

DEFF Research Database (Denmark)

Reynders, K.; Mischenko, N.; Kleppinger, R.

1997-01-01

Temperature and concentration dependencies of the degree of order in ABA triblock copolymer gels are discussed. Two factors can influence the ordering phenomena: the conformation of the midblocks (links of the network) and the polydispersity of the endblock domains (nodes of the network). The lat......Temperature and concentration dependencies of the degree of order in ABA triblock copolymer gels are discussed. Two factors can influence the ordering phenomena: the conformation of the midblocks (links of the network) and the polydispersity of the endblock domains (nodes of the network...

Molecular characterization of MHC-DRB cDNA in water buffalo (Bubalus bubalis

Directory of Open Access Journals (Sweden)

Soumen Naskar

2012-01-01

Full Text Available In the present study, water buffalo MHC (Bubu-DRB cDNA was cloned and characterized. The 1022 base long-amplified cDNA product encompassed a single open reading frame of 801 bases that coded for 266 amino acids. The Bubu-DRB sequence showed maximum homology with the BoLA-DRB3*0101 allele of cattle. A total of seven amino acid residues were found to be unique for the Bubu-DRB sequence. The majority of amino acid substitutions was observed in the β1 domain. Residues associated with important functions were mostly conserved. Water buffalo DRB was phylogenetically closer to goat DRB*A.

Integration of gel-based and gel-free proteomic data for functional analysis of proteins through Soybean Proteome Database

KAUST Repository

Komatsu, Setsuko

2017-05-10

The Soybean Proteome Database (SPD) stores data on soybean proteins obtained with gel-based and gel-free proteomic techniques. The database was constructed to provide information on proteins for functional analyses. The majority of the data is focused on soybean (Glycine max ‘Enrei’). The growth and yield of soybean are strongly affected by environmental stresses such as flooding. The database was originally constructed using data on soybean proteins separated by two-dimensional polyacrylamide gel electrophoresis, which is a gel-based proteomic technique. Since 2015, the database has been expanded to incorporate data obtained by label-free mass spectrometry-based quantitative proteomics, which is a gel-free proteomic technique. Here, the portions of the database consisting of gel-free proteomic data are described. The gel-free proteomic database contains 39,212 proteins identified in 63 sample sets, such as temporal and organ-specific samples of soybean plants grown under flooding stress or non-stressed conditions. In addition, data on organellar proteins identified in mitochondria, nuclei, and endoplasmic reticulum are stored. Furthermore, the database integrates multiple omics data such as genomics, transcriptomics, metabolomics, and proteomics. The SPD database is accessible at http://proteome.dc.affrc.go.jp/Soybean/. Biological significanceThe Soybean Proteome Database stores data obtained from both gel-based and gel-free proteomic techniques. The gel-free proteomic database comprises 39,212 proteins identified in 63 sample sets, such as different organs of soybean plants grown under flooding stress or non-stressed conditions in a time-dependent manner. In addition, organellar proteins identified in mitochondria, nuclei, and endoplasmic reticulum are stored in the gel-free proteomics database. A total of 44,704 proteins, including 5490 proteins identified using a gel-based proteomic technique, are stored in the SPD. It accounts for approximately 80% of all

Integration of gel-based and gel-free proteomic data for functional analysis of proteins through Soybean Proteome Database.

Science.gov (United States)

Komatsu, Setsuko; Wang, Xin; Yin, Xiaojian; Nanjo, Yohei; Ohyanagi, Hajime; Sakata, Katsumi

2017-06-23

The Soybean Proteome Database (SPD) stores data on soybean proteins obtained with gel-based and gel-free proteomic techniques. The database was constructed to provide information on proteins for functional analyses. The majority of the data is focused on soybean (Glycine max 'Enrei'). The growth and yield of soybean are strongly affected by environmental stresses such as flooding. The database was originally constructed using data on soybean proteins separated by two-dimensional polyacrylamide gel electrophoresis, which is a gel-based proteomic technique. Since 2015, the database has been expanded to incorporate data obtained by label-free mass spectrometry-based quantitative proteomics, which is a gel-free proteomic technique. Here, the portions of the database consisting of gel-free proteomic data are described. The gel-free proteomic database contains 39,212 proteins identified in 63 sample sets, such as temporal and organ-specific samples of soybean plants grown under flooding stress or non-stressed conditions. In addition, data on organellar proteins identified in mitochondria, nuclei, and endoplasmic reticulum are stored. Furthermore, the database integrates multiple omics data such as genomics, transcriptomics, metabolomics, and proteomics. The SPD database is accessible at http://proteome.dc.affrc.go.jp/Soybean/. The Soybean Proteome Database stores data obtained from both gel-based and gel-free proteomic techniques. The gel-free proteomic database comprises 39,212 proteins identified in 63 sample sets, such as different organs of soybean plants grown under flooding stress or non-stressed conditions in a time-dependent manner. In addition, organellar proteins identified in mitochondria, nuclei, and endoplasmic reticulum are stored in the gel-free proteomics database. A total of 44,704 proteins, including 5490 proteins identified using a gel-based proteomic technique, are stored in the SPD. It accounts for approximately 80% of all predicted proteins from

Integration of gel-based and gel-free proteomic data for functional analysis of proteins through Soybean Proteome Database

KAUST Repository

Komatsu, Setsuko; Wang, Xin; Yin, Xiaojian; Nanjo, Yohei; Ohyanagi, Hajime; Sakata, Katsumi

2017-01-01

The Soybean Proteome Database (SPD) stores data on soybean proteins obtained with gel-based and gel-free proteomic techniques. The database was constructed to provide information on proteins for functional analyses. The majority of the data is focused on soybean (Glycine max ‘Enrei’). The growth and yield of soybean are strongly affected by environmental stresses such as flooding. The database was originally constructed using data on soybean proteins separated by two-dimensional polyacrylamide gel electrophoresis, which is a gel-based proteomic technique. Since 2015, the database has been expanded to incorporate data obtained by label-free mass spectrometry-based quantitative proteomics, which is a gel-free proteomic technique. Here, the portions of the database consisting of gel-free proteomic data are described. The gel-free proteomic database contains 39,212 proteins identified in 63 sample sets, such as temporal and organ-specific samples of soybean plants grown under flooding stress or non-stressed conditions. In addition, data on organellar proteins identified in mitochondria, nuclei, and endoplasmic reticulum are stored. Furthermore, the database integrates multiple omics data such as genomics, transcriptomics, metabolomics, and proteomics. The SPD database is accessible at http://proteome.dc.affrc.go.jp/Soybean/. Biological significanceThe Soybean Proteome Database stores data obtained from both gel-based and gel-free proteomic techniques. The gel-free proteomic database comprises 39,212 proteins identified in 63 sample sets, such as different organs of soybean plants grown under flooding stress or non-stressed conditions in a time-dependent manner. In addition, organellar proteins identified in mitochondria, nuclei, and endoplasmic reticulum are stored in the gel-free proteomics database. A total of 44,704 proteins, including 5490 proteins identified using a gel-based proteomic technique, are stored in the SPD. It accounts for approximately 80% of all

Automated and unbiased image analyses as tools in phenotypic classification of small-spored Alternaria species

DEFF Research Database (Denmark)

Andersen, Birgitte; Hansen, Michael Edberg; Smedsgaard, Jørn

2005-01-01

often has been broadly applied to various morphologically and chemically distinct groups of isolates from different hosts. The purpose of this study was to develop and evaluate automated and unbiased image analysis systems that will analyze different phenotypic characters and facilitate testing...

Unbiased multi-fidelity estimate of failure probability of a free plane jet

Science.gov (United States)

Marques, Alexandre; Kramer, Boris; Willcox, Karen; Peherstorfer, Benjamin

2017-11-01

Estimating failure probability related to fluid flows is a challenge because it requires a large number of evaluations of expensive models. We address this challenge by leveraging multiple low fidelity models of the flow dynamics to create an optimal unbiased estimator. In particular, we investigate the effects of uncertain inlet conditions in the width of a free plane jet. We classify a condition as failure when the corresponding jet width is below a small threshold, such that failure is a rare event (failure probability is smaller than 0.001). We estimate failure probability by combining the frameworks of multi-fidelity importance sampling and optimal fusion of estimators. Multi-fidelity importance sampling uses a low fidelity model to explore the parameter space and create a biasing distribution. An unbiased estimate is then computed with a relatively small number of evaluations of the high fidelity model. In the presence of multiple low fidelity models, this framework offers multiple competing estimators. Optimal fusion combines all competing estimators into a single estimator with minimal variance. We show that this combined framework can significantly reduce the cost of estimating failure probabilities, and thus can have a large impact in fluid flow applications. This work was funded by DARPA.

Cost-effective sequencing of full-length cDNA clones powered by a de novo-reference hybrid assembly.

Science.gov (United States)

Kuroshu, Reginaldo M; Watanabe, Junichi; Sugano, Sumio; Morishita, Shinichi; Suzuki, Yutaka; Kasahara, Masahiro

2010-05-07

Sequencing full-length cDNA clones is important to determine gene structures including alternative splice forms, and provides valuable resources for experimental analyses to reveal the biological functions of coded proteins. However, previous approaches for sequencing cDNA clones were expensive or time-consuming, and therefore, a fast and efficient sequencing approach was demanded. We developed a program, MuSICA 2, that assembles millions of short (36-nucleotide) reads collected from a single flow cell lane of Illumina Genome Analyzer to shotgun-sequence approximately 800 human full-length cDNA clones. MuSICA 2 performs a hybrid assembly in which an external de novo assembler is run first and the result is then improved by reference alignment of shotgun reads. We compared the MuSICA 2 assembly with 200 pooled full-length cDNA clones finished independently by the conventional primer-walking using Sanger sequencers. The exon-intron structure of the coding sequence was correct for more than 95% of the clones with coding sequence annotation when we excluded cDNA clones insufficiently represented in the shotgun library due to PCR failure (42 out of 200 clones excluded), and the nucleotide-level accuracy of coding sequences of those correct clones was over 99.99%. We also applied MuSICA 2 to full-length cDNA clones from Toxoplasma gondii, to confirm that its ability was competent even for non-human species. The entire sequencing and shotgun assembly takes less than 1 week and the consumables cost only approximately US$3 per clone, demonstrating a significant advantage over previous approaches.

UNBIASED INCLINATION DISTRIBUTIONS FOR OBJECTS IN THE KUIPER BELT

International Nuclear Information System (INIS)

Gulbis, A. A. S.; Elliot, J. L.; Adams, E. R.; Benecchi, S. D.; Buie, M. W.; Trilling, D. E.; Wasserman, L. H.

2010-01-01

Using data from the Deep Ecliptic Survey (DES), we investigate the inclination distributions of objects in the Kuiper Belt. We present a derivation for observational bias removal and use this procedure to generate unbiased inclination distributions for Kuiper Belt objects (KBOs) of different DES dynamical classes, with respect to the Kuiper Belt plane. Consistent with previous results, we find that the inclination distribution for all DES KBOs is well fit by the sum of two Gaussians, or a Gaussian plus a generalized Lorentzian, multiplied by sin i. Approximately 80% of KBOs are in the high-inclination grouping. We find that Classical object inclinations are well fit by sin i multiplied by the sum of two Gaussians, with roughly even distribution between Gaussians of widths 2.0 +0.6 -0.5 0 and 8.1 +2.6 -2.1 0 . Objects in different resonances exhibit different inclination distributions. The inclinations of Scattered objects are best matched by sin i multiplied by a single Gaussian that is centered at 19.1 +3.9 -3.6 0 with a width of 6.9 +4.1 -2.7 0 . Centaur inclinations peak just below 20 0 , with one exceptionally high-inclination object near 80 0 . The currently observed inclination distribution of the Centaurs is not dissimilar to that of the Scattered Extended KBOs and Jupiter-family comets, but is significantly different from the Classical and Resonant KBOs. While the sample sizes of some dynamical classes are still small, these results should begin to serve as a critical diagnostic for models of solar system evolution.

Construction and biological activities of the first infectious cDNA clones of the genus Foveavirus

Energy Technology Data Exchange (ETDEWEB)

Meng, Baozhong, E-mail: bmeng@uoguelph.ca [Department of Molecular and Cellular Biology, University of Guelph, 50 Stone Road, Guelph, Ontario, Canada N1G2W1 (Canada); Venkataraman, Srividhya; Li, Caihong; Wang, Weizhou [Department of Molecular and Cellular Biology, University of Guelph, 50 Stone Road, Guelph, Ontario, Canada N1G2W1 (Canada); Dayan-Glick, Cathy; Mawassi, Munir [The Plant Pathology Department-The Virology Unit, Plant Protection Institute, Agricultural Research Organization, The Volcani Center, Bet-Dagan 50250 (Israel)

2013-01-20

Grapevine rupestris stem pitting-associated virus (GRSPaV, genus Foveavirus, family Betaflexiviridae) is one of the most prevalent viruses in grapevines and is associated with three distinct diseases: rupestris stem pitting, vein necrosis and Syrah decline. Little is known about the biology and pathological properties of GRSPaV. In this work, we engineered a full-length infectious cDNA clone for GRSPaV and a GFP-tagged variant, both under the transcriptional control of Cauliflower mosaic virus 35 S promoter. We demonstrated that these cDNA clones were infectious in grapevines and Nicotiana benthamiana through fluorescence microscopy, RT-PCR, Western blotting and immuno electron microscopy. Interestingly, GRSPaV does not cause systemic infection in four of the most commonly used herbaceous plants, even in the presence of the movement proteins of two other viruses which are known to complement numerous movement-defective viruses. These infectious clones are the first of members of Foveavirus which would allow further investigations into mechanisms governing different aspects of replication for GRSPaV and perhaps related viruses.

Two-dimensional gel electrophoresis pattern (pH 6-11) and identification of water-soluble barley seed and malt proteins by mass spectrometry

DEFF Research Database (Denmark)

Bak-Jensen, K.S.; Laugesen, S.; Roepstorff, P.

2004-01-01

A protocol was established for two-dimensional gel electrophoresis (2-DE) of barley seed and malt proteins in the pH range of 6-11. Proteins extracted from flour in a low-salt buffer were focused after cup-loading onto IPG strips. Successful separation in the second dimension was achieved using...... gradient gels in a horizontal SDS-PAGE system. Silver staining of gels visualized around 380 (seed) and 500 (malt) spots. Thirty-seven different proteins from seeds were identified in 60 spots, among these 46 were visualized also in the malt 2-D pattern. Proteins were identified by peptide mass...... in defence against pathogens (21 spots), 4 in storage, folding, and synthesis of proteins, and in nitrogen metabolism (5 spots), 6 in carbohydrate metabolism (11 spots), and 4 in stress and detoxification (9 spots). Six proteins (7 spots) were not grouped in these categories, and 3 were not ascribed...

Efficient method of protein extraction from Theobroma cacao L. roots for two-dimensional gel electrophoresis and mass spectrometry analyses.

Science.gov (United States)

Bertolde, F Z; Almeida, A-A F; Silva, F A C; Oliveira, T M; Pirovani, C P

2014-07-04

Theobroma cacao is a woody and recalcitrant plant with a very high level of interfering compounds. Standard protocols for protein extraction were proposed for various types of samples, but the presence of interfering compounds in many samples prevented the isolation of proteins suitable for two-dimensional gel electrophoresis (2-DE). An efficient method to extract root proteins for 2-DE was established to overcome these problems. The main features of this protocol are: i) precipitation with trichloroacetic acid/acetone overnight to prepare the acetone dry powder (ADP), ii) several additional steps of sonication in the ADP preparation and extractions with dense sodium dodecyl sulfate and phenol, and iii) adding two stages of phenol extractions. Proteins were extracted from roots using this new protocol (Method B) and a protocol described in the literature for T. cacao leaves and meristems (Method A). Using these methods, we obtained a protein yield of about 0.7 and 2.5 mg per 1.0 g lyophilized root, and a total of 60 and 400 spots could be separated, respectively. Through Method B, it was possible to isolate high-quality protein and a high yield of roots from T. cacao for high-quality 2-DE gels. To demonstrate the quality of the extracted proteins from roots of T. cacao using Method B, several protein spots were cut from the 2-DE gels, analyzed by tandem mass spectrometry, and identified. Method B was further tested on Citrus roots, with a protein yield of about 2.7 mg per 1.0 g lyophilized root and 800 detected spots.

A study of correlations between the release of drugs from petrolatum-based gels containing nonionic surfactants and some physical and physico-chemical characteristics of the gel systems.

Science.gov (United States)

Colo, G D; Nannipieri, E; Serafini, M F; Vitale, D

1986-06-01

Synopsis The in vitro release of benzocaine and 2-ethyIhexyl p-di-methylaminobenzoate (EH-PABA) from petrolatum-based gels either containing two nonionic surfactants, or not, was compared with some physical and/or physico-chemical characteristics of the drugs, the gels and the drug-gel systems. The surfactants had no effect on the release of EH-PABA, the less polar drug, whereas they decreased the release of benzocaine. Moreover, the release data show a complex dependence of diffusive properties of ben-zocaine on drug and surfactant concentration. Benzocaine appears to form mixed micelles with each of the two surfactants and/or undergoes self-aggregation phenomena within surfactant micelles. The results indicate that drug diffusion is influenced by gel porosity, drug molecular size and polarity and molecular interactions. Etude des corrélations entre la disponibilité des medicaments dans les gels a base de vaseline contenant des surfactifs non ioniques et quelques propriétés physiques et physicochimiques des gels.

MOLECULAR CHARACTERIZATION OF ENDOCRINE DISRUPTION IN FISH USING CDNA ARRAYS.

Science.gov (United States)

We are developing cDNA macroarrays to measure the induction of gene expression in sheepshead minnows and largemouth bass exposed to anthropogenic chemicals that can mimic the action of endogenous hormones. For sheepshead minnows exposed in aqua, we observed similar genetic profil...

Transcription analysis of apple fruit development using cDNA microarrays

NARCIS (Netherlands)

Soglio, V.; Costa, F.; Molthoff, J.W.; Weemen-Hendriks, M.; Schouten, H.J.; Gianfranceschi, L.

2009-01-01

The knowledge of the molecular mechanisms underlying fruit quality traits is fundamental to devise efficient marker-assisted selection strategies and to improve apple breeding. In this study, cDNA microarray technology was used to identify genes whose expression changes during fruit development and

Isolation and expression of a pea vicilin cDNA in the yeast Saccharomyces cerevisiae.

OpenAIRE

Watson, M D; Lambert, N; Delauney, A; Yarwood, J N; Croy, R R; Gatehouse, J A; Wright, D J; Boulter, D

1988-01-01

A cDNA clone containing the complete coding sequence for vicilin from pea (Pisum sativum L.) was isolated. It specifies a 50,000-Mr protein that in pea is neither post-translationally processed nor glycosylated. The cDNA clone was expressed in yeast from a 2 micron plasmid by using the yeast phosphoglycerate kinase promoter and initiator codon. The resultant fusion protein, which contains the first 16 amino acid residues of phosphoglycerate kinase in addition to the vicilin sequence, was puri...

Detection of hepatitis C virus RNA using reverse transcription PCR

International Nuclear Information System (INIS)

Yap, S.F.

1998-01-01

Detection of the viral genome (HCV RNA) is by a combination of cDNA synthesis and PCR followed by gel analysis and/or hybridization assay. In principle, cDNA is synthesized using the viral RNA as template and the enzyme, reverse transcriptase. The cDNA is then amplified by PCR and the product detected. Agarose gel electrophoresis provides a rapid and simple detection method; however, it is non-quantitative. The assay protocol described in this paper is adapted from that published by Chan et al. Comments on various aspects of the assay are based on experience with the method in our laboratory

Colloid molecular weight estimation by gel chromatography/acrylamide gel electrophoresis

International Nuclear Information System (INIS)

Liberatore, F.A.; Dearborn, C.; Nigam, S.; Poon, C.; Camin, L.; Liteplo, M.

1984-01-01

Size or molecular weight (MW) estimation of radiolabeled collides in aqueous solutions has long been a problem. The authors have prepared several minimicroaggregated albumin colloids (mμAA) by heat denaturation of stannous-containing HSA solutions at pH 7.0, 7.5, and 8.5). The resulting colloids were labeled with Tc-99m and compared with Au-198 colloid and Tc-99m-antimony sulfide colloid (Tc-99m-Sb/sub 2/S3) by gel chromatography and gel electrophoresis. Tc-99mm-mμAA aggregated at pH 7.0 and the Au-198 colloid appeared in the external void volume of a BioRad A5.0 agarose column indicating an apparent MW of > 5 x 10/sup 6/ daltons. The pH7.5 Tc-99m-mμAA, migrated within the filtration range of the column as did a small fraction of Tc-99m-Sb/sub 2/S/sub 3/, suggesting that the MW is between 6 x 10/sup 4/ - 5 x 10/sup 6/ daltons. The Tc-99m-mμAA, aggregated at pH 8.5, had an apparent MW on gel filtration similar to that of untreated albumin, MW 6.6 x 10-/sup 4/ daltons. The mobilities of the colloids, on acrylamide disc gel electrophoresis, were consistent with the results on gel chromatography. The largest colloids, Au-198 colloid and pH 7.0 Tc-99m-mμAA, barely entered the separating gel; intermediate sized colloids, a small fraction of Tc-99m-Sb/sub 2/S/sub 3/ and pH 7.5 Tc-99m-mμAA migrated farther into the separating gel; while pH 8.5 Tc-99m-mμAA had mobility approaching that of untreated albumin. Lymphoscintigraphy studies using these colloids in animals showed the predicted, particle size-related differences in migration and clearance. The authors conclude that gel chromatography and gel electrophoresis are useful methods for estimating the apparent size of the colloidal particles

Oral dextrose gel for the treatment of hypoglycaemia in newborn infants.

Science.gov (United States)

Weston, Philip J; Harris, Deborah L; Battin, Malcolm; Brown, Julie; Hegarty, Joanne E; Harding, Jane E

2016-05-04

Neonatal hypoglycaemia, a common condition, can be associated with brain injury. It is frequently managed by providing infants with an alternative source of glucose, given enterally with formula or intravenously with dextrose solution. This often requires that mother and baby are cared for in separate environments and may inhibit breast feeding. Dextrose gel is simple and inexpensive and can be administered directly to the buccal mucosa for rapid correction of hypoglycaemia, in association with continued breast feeding and maternal care. To assess the effectiveness of dextrose gel in correcting hypoglycaemia and in reducing long-term neurodevelopmental impairment. We searched MEDLINE, EMBASE, the Cochrane Central Register of Controlled Trials (CENTRAL), the Cumulative Index to Nursing and Allied Health Literature (CINAHL) and Web of Science from inception of the database to February 2016. We also searched international clinical trials networks and handsearched proceedings of specific scientific meetings. Randomised and quasi-randomised studies comparing dextrose gel versus placebo, no treatment or other therapies for treatment of neonatal hypoglycaemia. Two review authors independently assessed trial quality and extracted data and did not assess publications for which they themselves were study authors. We included two trials involving 312 infants. No data were available for correction of hypoglycaemia for each hypoglycaemic event. We found no evidence of a difference between dextrose gel and placebo gel for major neurosensory disability at two-year follow-up (risk ratio (RR) 6.27, 95% confidence interval (CI) 0.77 to 51.03; one trial, n = 184; quality of evidence very low). Dextrose gel compared with placebo gel or no gel did not alter the need for intravenous treatment for hypoglycaemia (typical RR 0.78, 95% CI 0.46 to 1.32; two trials, 312 infants; quality of evidence very low). Infants treated with dextrose gel were less likely to be separated from their

Sillica Gel-Amine from Geothermal Sludge

Science.gov (United States)

Muljani, S.; Pujiastuti, C.; Wicaksono, P.; Lutfianingrum, R.

2018-01-01

Silica Gel-Amine (SGA) has been made from geothermal sludge by grafting amine method. Sodium silicate solution is prepared by extracted geothermal sludge powder using sodium hidroxide solution then acidification in the range of pH 5 - 9 by using tartaric acid 1N. The grafting process uses 1 ml of ammonia solution and 10 ml of toluene at a rate of 0.1 ml min-1 accompanied by a reflux process. The amine grafting is done in two methods. The first method is grafting amine in silicate solution and the second method is grafting amine in washed gel. Product SGA was confirmed by FTIR, TGA-DTG and BET characterization. The results show that the pH affects the amount of amine that is grafted onto silica gel. Differences in grafting method affect the size of the pore and surface area. SGA product prepared by grafting washed gel at pH 8 have pore diameter of 12.06 nm, surface area of 173.44 m2g-1, and mass of decomposed amine compound 0.4 mg. In the presence of amine groups on the silica gel surface, these adsorbents may be able to selectively adsorb CO2 gas from natural gas.

Characterization of cDNA for PMT: a Partial Nicotine Biosynthesis-Related Gene Isolated from Indonesian Local Tobacco (Nicotiana tabacum cv. Sindoro1

Directory of Open Access Journals (Sweden)

SESANTI BASUKI

2013-12-01

Full Text Available Nicotine is the major alkaloid compound in cultivated tobacco (Nicotiana tabacum that could potentially be converted into carcinogenic compound (nor-nicotine. The PMT gene encoding putrescine N-methyltransferase (PMT is one of the two key genes that play a prominent role in nicotine biosynthesis. The aimed of this study was to isolate and characterize the cDNA sequence originated from Indonesian local tobacco cv. Sindoro1 (Ntpmt_Sindoro1. The results showed that the Ntpmt_Sindoro1 was 1124 bp in length. This cDNA fragment encodes for 374 amino acid residues. The predicted polypeptide from the cDNA is a hidrophilic protein, and has a predicted molecular weight of 40.95 kDa. The predicted amino acids sequence also showed high similarity to the PMT gene product Nicotiana sp. available in the GenBank data base. The amino acid sequences also exert conserved residues specifically exhibited only by PMT gene originated from N. tabacum. Clustering analysis revealed that Ntpmt_Sindoro1 belongs to the same clade as the PMT3 gene, a member of the N. tabacum PMT gene family. The Ntpmt_Sindoro1 cDNA sequence covering exon1-exon8 of the PMT gene fragment has been registered in the GenBank data base, under the accession number JX978277.

FIELD VALIDATION OF A SHEEPSHEAD MINNOW ESTROGEN-RESPONSIVE CDNA MACROARRAY

Science.gov (United States)

Hemmer, Michael J., Iris Knoebl, Becky L. Hemmer, Patrick Larkin, Peggy S. Harris and Nancy D. Denslow. In press. Field Validation of a Sheepshead Minnow Estrogen-Responsive cDNA Macroarray (Abstract). To be presented at the SETAC Fourth World Congress, 14-18 November 2004, Portl...

Correlating gene expression to physiological parameters and environmental conditons during cold acclimation of Pinus sylvestris, identification of molecular markers using cDNA microarrays

NARCIS (Netherlands)

Joosen, R.V.L.; Lammers, M.; Balk, P.A.; Bronnum, P.; Konings, M.C.J.M.; Perks, M.; Stattin, E.; Wordragen, van M.F.; Geest, van der A.H.M.

2006-01-01

Scots pine (Pinus sylvestris L.) seedlings were grown under different conditions (three field locations, two seasons and two climate room regimes), and then analyzed for freezing tolerance of shoots and roots and for transcript abundance in apical buds based on a cDNA microarray containing about

Aldehyde-Selective Wacker-Type Oxidation of Unbiased Alkenes Enabled by a Nitrite Co-Catalyst

KAUST Repository

Wickens, Zachary K.; Morandi, Bill; Grubbs, Robert H.

2013-01-01

Breaking the rules: Reversal of the high Markovnikov selectivity of Wacker-type oxidations was accomplished using a nitrite co-catalyst. Unbiased aliphatic alkenes can be oxidized with high yield and aldehyde selectivity, and several functional groups are tolerated. 18O-labeling experiments indicate that the aldehydic O atom is derived from the nitrite salt.

Aldehyde-Selective Wacker-Type Oxidation of Unbiased Alkenes Enabled by a Nitrite Co-Catalyst

KAUST Repository

Wickens, Zachary K.

2013-09-13

Breaking the rules: Reversal of the high Markovnikov selectivity of Wacker-type oxidations was accomplished using a nitrite co-catalyst. Unbiased aliphatic alkenes can be oxidized with high yield and aldehyde selectivity, and several functional groups are tolerated. 18O-labeling experiments indicate that the aldehydic O atom is derived from the nitrite salt.

Colloidal probe dynamics in gelatin solution during the sol-gel transition.

Science.gov (United States)

Hong, Wei; Xu, Guozhi; Ou, Xiaogang; Sun, Weixiang; Wang, Tao; Tong, Zhen

2018-05-16

The dynamics of the colloidal probes in a gelatin solution during the time-dependent sol-gel transition was investigated by multi-particle tracking. The relationship between the relaxation of the medium at the critical gel point and the mean square displacement of the probes was elucidated. Based on this understanding, the critical gel point of gelatin and the corresponding critical exponent n were unambiguously determined by the loss angle criterion and the time-cure superposition. The shift factors of the latter are further used to estimate the time/length-scale evolution of the gelatin during the sol-gel transition. The growth of the medium length scale crossed with the two measuring length scales successively at the pre-gel regime. Coinciding with the length-scale crossovers, the probability density function (PDF) of the probe displacements displayed two transient peaks of non-Gaussianity. In the post-gel regime, the third peak of Gaussianity suggested inhomogeneity in the gel network. The non-Gaussianity results from the bifurcation of diffusivity. The present work showed that the non-Gaussian dynamics of the probes are not the direct equivalence of that of the medium, but an effect of length-scale coupling.

Use of Non-Normalized, Non-Amplified cDNA for 454-Based RNA Sequencing of Fleshy Melon Fruit

Directory of Open Access Journals (Sweden)

Vitaly Portnoy

2011-03-01

Full Text Available The melon ( L. fruit is an important crop and model system for the genomic study of both fleshy fruit development and the Cucurbitaceae family. To obtain an accurate representation of the melon fruit transcriptome based on expressed sequence tag (EST abundance in 454-pyrosequencing data, we prepared double-stranded complementary DNA (cDNA of melon without the usual amplification and normalization steps. A purification step was also included to eliminate small fragments. Complementary DNAs were obtained from 14 individual fruit libraries derived from two genotypes, separated into flesh and peel tissues, and sampled throughout fruit development. Pyrosequencing was performed using Genome Sequencer FLX (GS FLX technology, resulting in 1,215,359 reads, with mean length of >200 nucleotides. The global digital expression data was validated by comparative reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR of 40 selected genes and expression patterns were similar for the two methods. The results indicate that high-quality, nonbiased cDNA for next-generation sequencing can be prepared from mature, fleshy fruit, which are notorious for difficulties in ribonucleic acid (RNA preparation.

cDNA, genomic sequence cloning and overexpression of ribosomal ...

African Journals Online (AJOL)

RPS16 of eukaryote is a component of the 40S small ribosomal subunit encoded by RPS16 gene and is also a homolog of prokaryotic RPS9. The cDNA and genomic sequence of RPS16 was cloned successfully for the first time from the Giant Panda (Ailuropoda melanoleuca) using reverse transcription-polymerase chain ...

Complete cDNA sequence coding for human docking protein

Energy Technology Data Exchange (ETDEWEB)

Hortsch, M; Labeit, S; Meyer, D I

1988-01-11

Docking protein (DP, or SRP receptor) is a rough endoplasmic reticulum (ER)-associated protein essential for the targeting and translocation of nascent polypeptides across this membrane. It specifically interacts with a cytoplasmic ribonucleoprotein complex, the signal recognition particle (SRP). The nucleotide sequence of cDNA encoding the entire human DP and its deduced amino acid sequence are given.

An unbiased stereological method for efficiently quantifying the innervation of the heart and other organs based on total length estimations

DEFF Research Database (Denmark)

Mühlfeld, Christian; Papadakis, Tamara; Krasteva, Gabriela

2010-01-01

Quantitative information about the innervation is essential to analyze the structure-function relationships of organs. So far, there has been no unbiased stereological tool for this purpose. This study presents a new unbiased and efficient method to quantify the total length of axons in a given...... reference volume, illustrated on the left ventricle of the mouse heart. The method is based on the following steps: 1) estimation of the reference volume; 2) randomization of location and orientation using appropriate sampling techniques; 3) counting of nerve fiber profiles hit by a defined test area within...

Fiscal 1998 achievement report. Industrial technology research and development project. (Strategic human cDNA genome application technology development); 1998 nendo senryakuteki hito cDNA genome oyo gijutsu kaihatsu seika hokokusho

Energy Technology Data Exchange (ETDEWEB)

NONE

2000-03-01

A human genome related project named above was started, and studies were conducted for base sequence determination and function analysis for approximately 10,000 kinds of full-length or long-chain human cDNA clones owned by research organizations in this country. The Institute of Medical Science of University of Tokyo and Helix Research Institute dealt with a full-length human cDNA library constructed by oligo-capping, and determined the base sequences of all specimens in the library. The Kazusa DNA Research Institute determined partial sequences for long-chain clones which are not shorter than 4-5kbp, and determined entire sequences for some bases. The obtained base sequence data were subjected to homology analysis, the base sequences were converted into amino acid sequences, and functions of proteins were predicted. In the analysis of gene functions, ATAC-PCR (adaptor tagged competitive-polymerase chain reaction) was applied to the clones covered by this project, and a database was prepared by use of the results of analyses of frequency-related information. For the preparation of a comprehensive gene expression profile, technologies for cDNA microarray construction were established. (NEDO)

Comparative gel-based phosphoproteomics in response to signaling molecules

KAUST Repository

Marondedze, Claudius; Lilley, Kathryn S.; Thomas, Ludivine

2013-01-01

The gel-based proteomics approach is a valuable technique for studying the characteristics of proteins. This technique has diverse applications ranging from analysis of a single protein to the study of the total cellular proteins. Further, protein quality and to some extent distribution can be first assessed by means of one-dimensional gel electrophoresis and then more informatively, for comparative analysis, using the two-dimensional gel electrophoresis technique. Here, we describe how to take advantage of the availability of fluorescent dyes to stain for a selective class of proteins on the same gel for the detection of both phospho- and total proteomes. This enables the co-detection of phosphoproteins as well as total proteins from the same gel and is accomplished by utilizing two different fluorescent stains, the ProQ-Diamond, which stains only phosphorylated proteins, and Sypro Ruby, which stains the entire subset of proteins. This workflow can be applied to gain insights into the regulatory mechanisms induced by signaling molecules such as cyclic nucleotides through the quantification and subsequent identification of responsive phospho- and total proteins. © Springer Science+Business Media New York 2013.

Comparative gel-based phosphoproteomics in response to signaling molecules

KAUST Repository

Marondedze, Claudius

2013-09-03

The gel-based proteomics approach is a valuable technique for studying the characteristics of proteins. This technique has diverse applications ranging from analysis of a single protein to the study of the total cellular proteins. Further, protein quality and to some extent distribution can be first assessed by means of one-dimensional gel electrophoresis and then more informatively, for comparative analysis, using the two-dimensional gel electrophoresis technique. Here, we describe how to take advantage of the availability of fluorescent dyes to stain for a selective class of proteins on the same gel for the detection of both phospho- and total proteomes. This enables the co-detection of phosphoproteins as well as total proteins from the same gel and is accomplished by utilizing two different fluorescent stains, the ProQ-Diamond, which stains only phosphorylated proteins, and Sypro Ruby, which stains the entire subset of proteins. This workflow can be applied to gain insights into the regulatory mechanisms induced by signaling molecules such as cyclic nucleotides through the quantification and subsequent identification of responsive phospho- and total proteins. © Springer Science+Business Media New York 2013.

Unbiased stereologic techniques for practical use in diagnostic histopathology

DEFF Research Database (Denmark)

Sørensen, Flemming Brandt

1995-01-01

Grading of malignancy by the examination of morphologic and cytologic details in histologic sections from malignant neoplasms is based exclusively on qualitative features, associated with significant subjectivity, and thus rather poor reproducibility. The traditional way of malignancy grading may...... by introducing quantitative techniques in the histopathologic discipline of malignancy grading. Unbiased stereologic methods, especially based on measurements of nuclear three-dimensional mean size, have during the last decade proved their value in this regard. In this survey, the methods are reviewed regarding...... the basic technique involved, sampling, efficiency, and reproducibility. Various types of cancers, where stereologic grading of malignancy has been used, are reviewed and discussed with regard to the development of a new objective and reproducible basis for carrying out prognosis-related malignancy grading...

Protein profile analysis of Malaysian snake venoms by two-dimensional gel electrophoresis

Directory of Open Access Journals (Sweden)

J Vejayan

2010-01-01

Full Text Available Snake venoms comprise a highly complex mixture of proteins, which requires for their characterization the use of versatile two-dimensional electrophoresis techniques. In the present study, venoms obtained from eight snakes (Ophiophagus hannah, Naja kaouthia, Naja sumatrana, Bungarus fasciatus, Trimeresurus sumatranus, Tropidolaemus wagleri, Enhydrina schistosa and Calloselasma rhodostoma commonly found in Malaysia were separated based on two independent properties, isoelectric point (pI and molecular weight (MW. Many differences in snake venoms at the inter-family, inter-subfamily, inter-genus and inter-species levels were revealed. Notably, proteins from individuals of the Viperidae family - Trimeresurus sumatranus, Tropidolaemus wagleri and Calloselasma rhodostoma - were found to be numerous and scattered by the two-dimensional gel electrophoresis (2DE specifically in regions between 37 and 100 kDa compared to the Elapidae venom proteins. The latter were clustered at the basic and lower molecular mass region (less than 20 kDa. Trains of spots were commonly observed, indicating that these proteins may be derived from post-translational modifications. Ophiophagus hannah (Elapidae revealed a great amount of protein spots in the higher molecular mass range when compared to Enhydrina schistosa, Naja kaouthia, Naja sumatrana and Bungarus fasciatus. Overall 2DE showed large differences in the venom profile of each species, which might be employed as an ancillary tool to the identification of venomous snake species.

Sol-gel Technology and Advanced Electrochemical Energy Storage Materials

Science.gov (United States)

Chu, Chung-tse; Zheng, Haixing

1996-01-01

Advanced materials play an important role in the development of electrochemical energy devices such as batteries, fuel cells, and electrochemical capacitors. The sol-gel process is a versatile solution for use in the fabrication of ceramic materials with tailored stoichiometry, microstructure, and properties. This processing technique is particularly useful in producing porous materials with high surface area and low density, two of the most desirable characteristics for electrode materials. In addition,the porous surface of gels can be modified chemically to create tailored surface properties, and inorganic/organic micro-composites can be prepared for improved material performance device fabrication. Applications of several sol-gel derived electrode materials in different energy storage devices are illustrated in this paper. V2O5 gels are shown to be a promising cathode material for solid state lithium batteries. Carbon aerogels, amorphous RuO2 gels and sol-gel derived hafnium compounds have been studied as electrode materials for high energy density and high power density electrochemical capacitors.

Isolation of an insulin-like growth factor II cDNA with a unique 5' untranslated region from human placenta

International Nuclear Information System (INIS)

Shen, Shujane; Daimon, Makoto; Wang, Chunyeh; Ilan, J.; Jansen, M.

1988-01-01

Human insulin-like growth factor II (IGF-II) cDNA from a placental library was isolated and sequenced. The 5' untranslated region (5'-UTR) sequence of this cDNA differs completely from that of adult human liver and has considerable base sequence identity to the same region of an IGF-II cDNA of a rat liver cell line, BRL-3A. Human placental poly(A) + RNA was probed with either the 5'-UTR of the isolated human placental IGF-II cDNA or the 5'-UTR of the IGF-II cDNA obtained from adult human liver. No transcripts were detected by using the 5'-UTR of the adult liver IGF-II as the probe. In contrast, three transcripts of 6.0, 3.2, and 2.2 kilobases were detected by using the 5'-UTR of the placental IGF-II cDNA as the probe or the probe from the coding sequence. A fourth IGF-II transcript of 4.9 kilobases presumably containing a 5'-UTR consisting of a base sequence dissimilar to that of either IGF-II 5'-UTR was apparent. Therefore, IGF-II transcripts detected may be products of alternative splicing as their 5'-UTR sequence is contained within the human IGF-II gene or they may be a consequence of alternative promoter utilization in placenta

Structural organization of the human glucocorticoid receptor determined by one- and two-dimensional gel electrophoresis of proteolytic receptor fragments

International Nuclear Information System (INIS)

Smith, A.C.; Harmon, J.M.

1987-01-01

The structural organization of the steroid-binding protein of the IM-9 cell glucocorticoid receptor was investigated by using one- and two-dimensional gel electrophoresis of proteolytic receptor fragments. One-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) of receptor fragments isolated after trypsin digestion of immunopurified [ 3 H]dexamethasone 21-mesylate ([ 3 H]DM-) labeled receptor revealed the presence of a stable 26.5-kilodalton (kDa) steroid-containing non-DNA-binding fragment, derived from a larger, less stable, 29-kDa fragment. The 26.5-kDa tryptic fragment appeared to be completely contained within a 41-kDa, steroid-containing, DNA-binding species isolated after chymotrypsin digestion of the intact protein. Two-dimensional electrophoretic analysis of the [ 3 H]DM-labeled tryptic fragments resolved two 26.5-kDa and two 29-kDa components. This was the same number of isoforms seen in the intact protein, indicating that the charge heterogeneity of the steroid-binding protein is the result of modification within the steroid-containing, non-DNA-binding, 26.5-kDa tryptic fragment. Two-dimensional analysis of the 41-kDa [ 3 H]DM-labeled chymotryptic species revealed a pattern of isoforms more complex than that seen either in the intact protein or in the steroid-containing tryptic fragments. These results suggest that the 41-kDa [ 3 H]DM-labeled species resolved by one-dimensional SDS-PAGE after chymotrypsin digestion may be composed of several distinct proteolytic fragments

Effects of gel composition on the radiation induced density change in PAG polymer gel dosimeters: a model and experimental investigations

International Nuclear Information System (INIS)

Hilts, M; Jirasek, A; Duzenli, C

2004-01-01

Due to a density change that occurs in irradiated polyacrylamide gel (PAG), x-ray computed tomography (CT) has emerged as a feasible method of performing polymer gel dosimetry. However, applicability of the technique is currently limited by low sensitivity of the density change to dose. This work investigates the effect of PAG composition on the radiation induced density change and provides direction for future work in improving the sensitivity of CT polymer gel dosimetry. A model is developed that describes the PAG density change (Δρ gel ) as a function of both polymer yield (%P) and an intrinsic density change, per unit polymer yield, that occurs on conversion of monomer to polymer (Δρ polymer ). %P is a function of the fraction of monomer consumed and the weight fraction of monomer in the unirradiated gel (%T). Applying the model to experimental CT and Raman spectroscopic data, two important fundamental properties of the response of PAG density to dose (Δρ gel dose response) are discovered. The first property is that Δρ polymer depends on PAG %C (cross-linking fraction of total monomer) such that low and high %C PAGs exhibit a higher Δρ polymer than do more intermediate %C PAGs. This relationship is opposite to the relationship of polymer yield to %C and is explained by the effect of %C on the type of polymer formed. The second property is that the Δρ gel dose response is linearly dependent on %T. From the model, the inference is that, at least for %T≤12%, monomer consumption and Δρ polymer depend solely on %C. In terms of optimizing CT polymer gel dosimetry for high sensitivity, these results indicate that Δρ polymer can be expected to vary with each polymer gel system and thus should be considered when choosing a polymer gel for CT gel dosimetry. However, Δρ polymer and %P cannot be maximized simultaneously and maximizing %P, by choosing gels with intermediate %C and high %T, is found to have the greatest impact on increasing the

Benchmarking of the Oxford Nanopore MinION sequencing for quantitative and qualitative assessment of cDNA populations.

Science.gov (United States)

Oikonomopoulos, Spyros; Wang, Yu Chang; Djambazian, Haig; Badescu, Dunarel; Ragoussis, Jiannis

2016-08-24

To assess the performance of the Oxford Nanopore Technologies MinION sequencing platform, cDNAs from the External RNA Controls Consortium (ERCC) RNA Spike-In mix were sequenced. This mix mimics mammalian mRNA species and consists of 92 polyadenylated transcripts with known concentration. cDNA libraries were generated using a template switching protocol to facilitate the direct comparison between different sequencing platforms. The MinION performance was assessed for its ability to sequence the cDNAs directly with good accuracy in terms of abundance and full length. The abundance of the ERCC cDNA molecules sequenced by MinION agreed with their expected concentration. No length or GC content bias was observed. The majority of cDNAs were sequenced as full length. Additionally, a complex cDNA population derived from a human HEK-293 cell line was sequenced on an Illumina HiSeq 2500, PacBio RS II and ONT MinION platforms. We observed that there was a good agreement in the measured cDNA abundance between PacBio RS II and ONT MinION (rpearson = 0.82, isoforms with length more than 700bp) and between Illumina HiSeq 2500 and ONT MinION (rpearson = 0.75). This indicates that the ONT MinION can sequence quantitatively both long and short full length cDNA molecules.

A linear concatenation strategy to construct 5'-enriched amplified cDNA libraries using multiple displacement amplification.

Science.gov (United States)

Gadkar, Vijay J; Filion, Martin

2013-06-01

In various experimental systems, limiting available amounts of RNA may prevent a researcher from performing large-scale analyses of gene transcripts. One way to circumvent this is to 'pre-amplify' the starting RNA/cDNA, so that sufficient amounts are available for any downstream analysis. In the present study, we report the development of a novel protocol for constructing amplified cDNA libraries using the Phi29 DNA polymerase based multiple displacement amplification (MDA) system. Using as little as 200 ng of total RNA, we developed a linear concatenation strategy to make the single-stranded cDNA template amenable for MDA. The concatenation, made possible by the template switching property of the reverse transcriptase enzyme, resulted in the amplified cDNA library with intact 5' ends. MDA generated micrograms of template, allowing large-scale polymerase chain reaction analyses or other large-scale downstream applications. As the amplified cDNA library contains intact 5' ends, it is also compatible with 5' RACE analyses of specific gene transcripts. Empirical validation of this protocol is demonstrated on a highly characterized (tomato) and an uncharacterized (corn gromwell) experimental system.

Identification of a cryptic prokaryotic promoter within the cDNA encoding the 5' end of dengue virus RNA genome.

Directory of Open Access Journals (Sweden)

Dongsheng Li

Full Text Available Infectious cDNA clones of RNA viruses are important research tools, but flavivirus cDNA clones have proven difficult to assemble and propagate in bacteria. This has been attributed to genetic instability and/or host cell toxicity, however the mechanism leading to these difficulties has not been fully elucidated. Here we identify and characterize an efficient cryptic bacterial promoter in the cDNA encoding the dengue virus (DENV 5' UTR. Following cryptic transcription in E. coli, protein expression initiated at a conserved in-frame AUG that is downstream from the authentic DENV initiation codon, yielding a DENV polyprotein fragment that was truncated at the N-terminus. A more complete understanding of constitutive viral protein expression in E. coli might help explain the cloning and propagation difficulties generally observed with flavivirus cDNA.

Isolation of CYP3A5P cDNA from human liver: a reflection of a novel cytochrome P-450 pseudogene.

Science.gov (United States)

Schuetz, J D; Guzelian, P S

1995-03-14

We have isolated, from a human liver cDNA library, a 1627 bp CYP3A5 cDNA variant (CYP3A5P) that contains several large insertions, deletions, and in-frame termination codons. By comparison with the genomic structure of other CYP3A genes, the major insertions in CYP3A5P cDNA demarcate the inferred sites of several CYP3A5 exons. The segments inserted in CYP3A5P have no homology with splice donor acceptor sites. It is unlikely that CYP3A5P cDNA represents an artifact of the cloning procedures since Southern blot analysis of human genomic DNA disclosed that CYP3A5P cDNA hybridized with a DNA fragment distinct from fragments that hybridized with either CYP3A5, CYP3A3 or CYP3A4. Moreover, analysis of adult human liver RNA on Northern blots hybridized with a CYP3A5P cDNA fragment revealed the presence of an mRNA with the predicted size of CYP3A5P. We conclude that CYP3A5P cDNA was derived from a separate gene, CYP3A5P, most likely a pseudogene evolved from CYP3A5.

Functional study on two artificial liver bioreactors with collagen gel

Directory of Open Access Journals (Sweden)

XU Bing

2014-10-01

Full Text Available ObjectiveTo improve the hollow fiber bioreactor of artificial liver. MethodsRat hepatocytes mixed with collagen solution were injected into the external cavity of a hollow fiber reactor to construct a bioreactor of hepatocytes suspended in collagen gel (group Ⅰ. Other rat hepatocytes suspended in solution were injected into the external cavity of a hollow fiber reactor with a layer of collagen on the wall of the external cavity to construct a bioreactor of collagen layer and hepatocytes (group Ⅱ. For each group, the culture solution circulated through the internal cavity of the hollow fiber bioreactor; the bioreactor was put in a culture box for 9 d, and the culture solution in the internal cavity was exchanged for new one every 24 h; the concentrations of albumin (Alb, urea, and lactate dehydrogenase (LDH in the culture solution samples were measured to examine the hepatocyte function of the bioreactor. Statistical analysis was performed using SPSS 130. Continuous data were expressed as mean±SD, and comparison between groups was made by paired t test. ResultsFor groups Ⅰ and Ⅱ, Alb levels reached peak values on day 3 of culture (1.41±0.08 g/L and 0.65±0.05 g/L; from day 3 to 9, group I had a significantly higher Alb level than group Ⅱ (t＞7.572, P＜0.01. For groups Ⅰ and Ⅱ, urea levels reached peak values on days 3 and 5 of culture (1.73±0.14 mmol/L and 1.56±0.18 mmol/L; from days 5 to 9, group I had a significantly higher urea level than group Ⅱ (t＞8.418, P＜0.01. For groups Ⅰ and Ⅱ, LDH levels reached peak values on day 9 of culture (32.03±9.13 U/L and 70.17±25.28 U/L; from days 1 to 9, group I had a significantly lower LDH level than group Ⅱ(t＞5.633, P＜0.01. Therefore, the bioreactor of hepatocytes suspended in collagen gel (group Ⅰ showed a better hepatocyte function and less hepatic enzyme leakage compared with the bioreactor of collagen layer and hepatocytes (group Ⅱ. Conclusion

Background-free, high sensitivity staining of proteins in one- and two-dimensional sodium dodecyl sulfate-polyacrylamide gels using a luminescent ruthenium complex.

Science.gov (United States)

Berggren, K; Chernokalskaya, E; Steinberg, T H; Kemper, C; Lopez, M F; Diwu, Z; Haugland, R P; Patton, W F

2000-07-01

SYPRO Ruby dye is a permanent stain comprised of ruthenium as part of an organic complex that interacts noncovalently with proteins. SYPRO Ruby Protein Gel Stain provides a sensitive, gentle, fluorescence-based method for detecting proteins in one-dimensional and two-dimensional sodium dodecyl sulfate-polyacrylamide gels. Proteins are fixed, stained from 3h to overnight and then rinsed in deionized water or dilute methanol/acetic acid solution for 30 min. The stain can be visualized using a wide range of excitation sources commonly used in image analysis systems including a 302 nm UV-B transilluminator, 473 nm second harmonic generation (SHG) laser, 488 nm argon-ion laser, 532 nm yttrium-aluminum-garnet (YAG) laser, xenon arc lamp, blue fluorescent light bulb or blue light-emitting diode (LED). The sensitivity of SYPRO Ruby Protein Gel Stain is superior to colloidal Coomassie Brilliant Blue (CBB) stain or monobromobimane labeling and comparable with the highest sensitivity silver or zinc-imidazole staining procedures available. The linear dynamic range of SYPRO Ruby Protein Gel stain extends over three orders of magnitude, which is vastly superior to silver, zinc-imidazole, monobromobimane and CBB stain. The fluorescent stain does not contain superfluous chemicals (formaldehyde, glutaraldehyde, Tween-20) that frequently interfere with peptide identification in mass spectrometry. While peptide mass profiles are severely altered in protein samples prelabeled with monobromobimane, successful identification of proteins by peptide mass profiling using matrix-assisted laser desorption/ionization mass spectrometry was easily performed after protein detection with SYPRO Ruby Protein Gel stain.

Cloning and expression of a cDNA coding for a human monocyte-derived plasminogen activator inhibitor

International Nuclear Information System (INIS)

Antalis, T.M.; Clark, M.A.; Barnes, T.; Lehrbach, P.R.; Devine, P.L.; Schevzov, G.; Goss, N.H.; Stephens, R.W.; Tolstoshev, P.

1988-01-01

Human monocyte-derived plasminogen activator inhibitor (mPAI-2) was purified to homogeneity from the U937 cell line and partially sequenced. Oligonucleotide probes derived from this sequence were used to screen a cDNA library prepared from U937 cells. One positive clone was sequenced and contained most of the coding sequence as well as a long incomplete 3' untranslated region (1112 base pairs). This cDNA sequence was shown to encode mPAI-2 by hybrid-select translation. A cDNA clone encoding the remainder of the mPAI-2 mRNA was obtained by primer extension of U937 poly(A) + RNA using a probe complementary to the mPAI-2 coding region. The coding sequence for mPAI-2 was placed under the control of the λ P/sub L/ promoter, and the protein expressed in Escherichia coli formed a complex with urokinase that could be detected immunologically. By nucleotide sequence analysis, mPAI-2 cDNA encodes a protein containing 415 amino acids with a predicted unglycosylated M/sub r/ of 46,543. The predicted amino acid sequence of mPAI-2 is very similar to placental PAI-2 and shows extensive homology with members of the serine protease inhibitor (serpin) superfamily. mPAI-2 was found to be more homologous to ovalbumin (37%) than the endothelial plasminogen activator inhibitor, PAI-1 (26%). The 3' untranslated region of the mPAI-2 cDNA contains a putative regulatory sequence that has been associated with the inflammatory mediators

Gel versus capillary electrophoresis genotyping for categorizing treatment outcomes in two anti-malarial trials in Uganda

OpenAIRE

Hubbard Alan E; Dorsey Grant; Gupta Vinay; Rosenthal Philip J; Greenhouse Bryan

2010-01-01

Abstract Background Molecular genotyping is performed in anti-malarial trials to determine whether recurrent parasitaemia after therapy represents a recrudescence (treatment failure) or new infection. The use of capillary instead of agarose gel electrophoresis for genotyping offers technical advantages, but it is unclear whether capillary electrophoresis will result in improved classification of anti-malarial treatment outcomes. Methods Samples were genotyped using both gel and capillary elec...

Detection of reverse transcriptase termination sites using cDNA ligation and massive parallel sequencing

DEFF Research Database (Denmark)

Kielpinski, Lukasz J; Boyd, Mette; Sandelin, Albin

2013-01-01

Detection of reverse transcriptase termination sites is important in many different applications, such as structural probing of RNAs, rapid amplification of cDNA 5' ends (5' RACE), cap analysis of gene expression, and detection of RNA modifications and protein-RNA cross-links. The throughput...... of these methods can be increased by applying massive parallel sequencing technologies.Here, we describe a versatile method for detection of reverse transcriptase termination sites based on ligation of an adapter to the 3' end of cDNA with bacteriophage TS2126 RNA ligase (CircLigase™). In the following PCR...

An Example of an Improvable Rao-Blackwell Improvement, Inefficient Maximum Likelihood Estimator, and Unbiased Generalized Bayes Estimator.

Science.gov (United States)

Galili, Tal; Meilijson, Isaac

2016-01-02

The Rao-Blackwell theorem offers a procedure for converting a crude unbiased estimator of a parameter θ into a "better" one, in fact unique and optimal if the improvement is based on a minimal sufficient statistic that is complete. In contrast, behind every minimal sufficient statistic that is not complete, there is an improvable Rao-Blackwell improvement. This is illustrated via a simple example based on the uniform distribution, in which a rather natural Rao-Blackwell improvement is uniformly improvable. Furthermore, in this example the maximum likelihood estimator is inefficient, and an unbiased generalized Bayes estimator performs exceptionally well. Counterexamples of this sort can be useful didactic tools for explaining the true nature of a methodology and possible consequences when some of the assumptions are violated. [Received December 2014. Revised September 2015.].

Methods to determine the transcriptomes of trypanosomes in mixtures with mammalian cells: the effects of parasite purification and selective cDNA amplification.

Directory of Open Access Journals (Sweden)

Julius Mulindwa

2014-04-01

Full Text Available Patterns of gene expression in cultured Trypanosoma brucei bloodstream and procyclic forms have been extensively characterized, and some comparisons have been made with trypanosomes grown to high parasitaemias in laboratory rodents. We do not know, however, to what extent these transcriptomes resemble those in infected Tsetse flies - or in humans or cattle, where parasitaemias are substantially lower. For clinical and field samples it is difficult to characterize parasite gene expression because of the large excess of host cell RNA. We have here examined two potential solutions to this problem for bloodstream form trypanosomes, assaying transcriptomes by high throughput cDNA sequencing (RNASeq. We first purified the parasites from blood of infected rats. We found that a red blood cell lysis procedure affected the transcriptome substantially more than purification using a DEAE cellulose column, but that too introduced significant distortions and variability. As an alternative, we specifically amplified parasite sequences from a mixture containing a 1000-fold excess of human RNA. We first purified polyadenylated RNA, then made trypanosome-specific cDNA by priming with a spliced leader primer. Finally, the cDNA was amplified using nested primers. The amplification procedure was able to produce samples in which 20% of sequence reads mapped to the trypanosome genome. Synthesis of the second cDNA strand with a spliced leader primer, followed by amplification, is sufficiently reproducible to allow comparison of different samples so long as they are all treated in the same way. However, SL priming distorted the abundances of the cDNA products and definitely cannot be used, by itself, to measure absolute mRNA levels. The amplification method might be suitable for clinical samples with low parasitaemias, and could also be adapted for other Kinetoplastids and to samples from infected vectors.

Enhanced specificity in immunoscreening of expression cDNA clones using radiolabeled antigen overlay

International Nuclear Information System (INIS)

Chao, S.; Chao, L.; Chao, J.

1989-01-01

A highly sensitive and specific method has been developed for immunoscreening clones from an expression cDNA library. The procedures utilize a radiolabeled antigen detection method described originally for the immunoblotting of plasma proteins. Screening of rat alpha 1-antitrypsin clones was used. Comparison between Western blots of alpha 1-antitrypsin using both labeled antigen and protein A detection methods showed that the former yielded lower background and greater sensitivity than the latter. Further, this technique was shown to have a lower detection limit of less than 20 ng through Western blot analysis of varying concentrations of alpha 1-antitrypsin. The procedures are based on the expression of the protein by cDNA clones containing the DNA inserts in the correct reading frame. Following the transfer of phage proteins to nitrocellulose membranes, the bivalent antibodies bind monovalently to both nitrocellulose-bound-antigen in the phage lysates and radiolabeled antigen. The radiolabeled antigen overlay method is superior to the protein A detection method in sensitivity, specificity and reproducibility. This improved method can be applied in general for screening expression cDNA libraries, provided that the specific antiserum and radiolabeled antigen are available

NMR mechanisms in gel dosimetry

International Nuclear Information System (INIS)

Schreiner, L J

2009-01-01

Nuclear magnetic resonance was critical to the development of gel dosimetry, as it established the potential for three dimensional dosimetry with chemical dosimeter systems through magnetic resonance imaging [1]. In the last two decades MRI has served as the gold standard for imaging, while NMR relaxometry has played an important role in the development and understanding of the behaviour of new gel dosimetry systems. Therefore, an appreciation of the relaxation mechanisms determining the NMR behaviour of irradiated gel dosimeters is important for a full comprehension of a considerable component of the literature on gel dosimetry. A number of excellent papers have presented this important theory, this brief review will highlight some of the salient points made previously [1-5]. The spin relaxation of gel dosimeters (which determines the dose dependence in most conventional MR imaging) is determined principally by the protons on water molecules in the system. These water protons exist in different environments, or groups (see Figure 1): on bulk water, on water hydrating the chemical species that are being modified under irradiation, and on water hydrating the gel matrix used to spatially stabilize the dosimeter (e.g., gelatin, agarose, etc). The spin relaxation depends on the inherent relaxation rate of each spin group, that is, on the relaxation rate which would be observed for the specific group if it were isolated. Also, the different water environments are not isolated from each other, and the observed relaxation rate also depends on the rate of exchange of magnetization between the groups, and on the fraction of protons in each group. In fact, the water exchanges quickly between the environments, so that relaxation is in what is usually termed the fast exchange regime. In the limit of fast exchange, the relaxation of the water protons is well characterized by a single exponential and hence by a single apparent relaxation rate. In irradiated gel dosimeters this

Effects of interferon gamma on Chlamydia trachomatis serovar A and L2 protein expression investigated by two-dimensional gel electrophoresis

DEFF Research Database (Denmark)

Shaw, A; Christiansen, Gunna; Birkelund, Svend

1999-01-01

]methionine and two-dimensional gel electrophoresis with immobilized pH gradients in order to investigate changes in the protein expression of C. trachomatis serovar A and L2 caused by treatment with IFN-gamma. In contrast to what was observed in C. trachomatis L2, our results showed that, in C. trachomatis A, down...

Removing water from gels

International Nuclear Information System (INIS)

Lane, E.S.; Winter, J.A.

1982-01-01

Water is removed from a gel material by contacting the gel material with an organic liquid and contacting the organic liquid with a gas such that water is taken up by the gas. The invention, in one embodiment, may be used to dry gel materials whilst maintaining an open porous network therein. In one example, the invention is applied to gel precipitated spheres containing uranium and plutonium. (author)

Isolation and characterization of human glycophorin A cDNA clones by a synthetic oligonucleotide approach: nucleotide sequence and mRNA structure

International Nuclear Information System (INIS)

Siebert, P.D.; Fukuda, M.

1986-01-01

In an effort to understand the relationships among and the regulation of human glycophorins, the authors have isolated and characterized several glycophorin A-specific cDNA clones obtained from a human erythroleukemic K562 cell cDNA library. This was accomplished by using mixed synthetic oligonucleotides, corresponding to various regions of the known amino acid sequence, to prime the synthesis of the cDNA as well as to screen the cDNA library. They also used synthetic oligonucleotides to sequence the largest of the glycophorin cDNAs. The nucleotide sequence obtained suggests the presence of a potential leader peptide, consistent with the membrane localization of this glycoprotein. Examination of the structure of glycophorin mRNA by blot hybridization revealed the existence of several electrophoretically distinct mRNAs numbering three or four, depending on the size of the glycophorin cDNA used as a hybridization probe. The smaller cDNA hybridized to three mRNAs of approximately 2.8, 1.7, and 1.0 kilobases. In contrast, the larger cDNA hybridized to an additional mRNA of approximately 0.6 kilobases. Further examination of the relationships between these multiple mRNAs by blot hybridization was conducted with the use of exact-sequence oligonucleotide probes constructed from various regions of the cDNA representing portions of the amino acid sequence of glycophorin A with or without known homology with glycophorin B. In total, the results obtained are consistent with the hypothesis that the three larger mRNAs represent glycophorin A gene transcripts and that the smallest (0.6 kilobase) mRNA may be specific for glycophorin B

Numerical modelling and experimental study of liquid evaporation during gel formation

Science.gov (United States)

Pokusaev, B. G.; Khramtsov, D. P.

2017-11-01

Gels are promising materials in biotechnology and medicine as a medium for storing cells for bioprinting applications. Gel is a two-phase system consisting of solid medium and liquid phase. Understanding of a gel structure evolution and gel aging during liquid evaporation is a crucial step in developing new additive bioprinting technologies. A numerical and experimental study of liquid evaporation was performed. In experimental study an evaporation process of an agarose gel layer located on Petri dish was observed and mass difference was detected using electronic scales. Numerical model was based on a smoothed particle hydrodynamics method. Gel in a model was represented as a solid-liquid system and liquid evaporation was modelled due to capillary forces and heat transfer. Comparison of experimental data and numerical results demonstrated that model can adequately represent evaporation process in agarose gel.

Characterization of cDNA encoding molt-inhibiting hormone of the crab, Cancer pagurus; expression of MIH in non-X-organ tissues.

Science.gov (United States)

Lu, W; Wainwright, G; Olohan, L A; Webster, S G; Rees, H H; Turner, P C

2001-10-31

Synthesis of ecdysteroids (molting hormones) by crustacean Y-organs is regulated by a neuropeptide, molt-inhibiting hormone (MIH), produced in eyestalk neural ganglia. We report here the molecular cloning of a cDNA encoding MIH of the edible crab, Cancer pagurus. Full-length MIH cDNA was obtained by using reverse transcription-polymerase chain reaction (RT-PCR) with degenerate oligonucleotides based upon the amino acid sequence of MIH, in conjunction with 5'- and 3'-RACE. Full-length clones of MIH cDNA were obtained that encoded a 35 amino acid putative signal peptide and the mature 78 amino acid peptide. Of various tissues examined by Northern blot analysis, the X-organ was the sole major site of expression of the MIH gene. However, a nested-PCR approach using non-degenerate MIH-specific primers indicated the presence of MIH transcripts in other tissues. Southern blot analysis indicated a simple gene arrangement with at least two copies of the MIH gene in the genome of C. pagurus. Additional Southern blotting experiments detected MIH-hybridizing bands in another Cancer species, Cancer antennarius and another crab species, Carcinus maenas.

Effects of Shear Fracture on In-depth Profile Modification of Weak Gels

Institute of Scientific and Technical Information of China (English)

Li Xianjie; Song Xinwang; Yue Xiang'an; Hou Jirui; Fang Lichun; Zhang Huazhen

2007-01-01

Two sand packs were filled with fine glass beads and quartz sand respectively. The characteristics of crosslinked polymer flowing through the sand packs as well as the influence of shear fracture of porous media on the in-depth profile modification of the weak gel generated from the crosslinked polymer were investigated. The results indicated that under the dynamic condition crosslinking reaction happened in both sand packs,and the weak gels in these two cases became small gel particles after water flooding. The differences were:the dynamic gelation time in the quartz sand pack was longer than that in the glass bead pack. Residual resistance factor (FRR) caused by the weak gel in the quartz sand pack was smaller than that in the glass bead pack. The weak gel became gel particles after being scoured by subsequent flood water. A weak gel with uniform apparent viscosity and sealing characteristics was generated in every part of the glass bead pack,which could not only move deeply into the sand pack but also seal the high capacity channels again when it reached the deep part. The weak gel performed in-depth profile modification in the glass bead pack,while in the quartz sand pack,the weak gel was concentrated with 100 cm from the entrance of the sand pack. When propelled by the subsequent flood water,the weak gel could move towards the deep part of the sand pack but then became tiny gel particles and could not effectively seal the high capacity channels there. The in-depth profile modification of the weak gel was very weak in the quartz sand pack. It was the shear fracture of porous media that mainly affected the properties and weakened the in-depth profile modification of the weak gel.

An analysis of expressed sequence tags of developing castor endosperm using a full-length cDNA library

Directory of Open Access Journals (Sweden)

Wallis James G

2007-07-01

Full Text Available Abstract Background Castor seeds are a major source for ricinoleate, an important industrial raw material. Genomics studies of castor plant will provide critical information for understanding seed metabolism, for effectively engineering ricinoleate production in transgenic oilseeds, or for genetically improving castor plants by eliminating toxic and allergic proteins in seeds. Results Full-length cDNAs are useful resources in annotating genes and in providing functional analysis of genes and their products. We constructed a full-length cDNA library from developing castor endosperm, and obtained 4,720 ESTs from 5'-ends of the cDNA clones representing 1,908 unique sequences. The most abundant transcripts are genes encoding storage proteins, ricin, agglutinin and oleosins. Several other sequences are also very numerous, including two acidic triacylglycerol lipases, and the oleate hydroxylase (FAH12 gene that is responsible for ricinoleate biosynthesis. The role(s of the lipases in developing castor seeds are not clear, and co-expressing of a lipase and the FAH12 did not result in significant changes in hydroxy fatty acid accumulation in transgenic Arabidopsis seeds. Only one oleate desaturase (FAD2 gene was identified in our cDNA sequences. Sequence and functional analyses of the castor FAD2 were carried out since it had not been characterized previously. Overexpression of castor FAD2 in a FAH12-expressing Arabidopsis line resulted in decreased accumulation of hydroxy fatty acids in transgenic seeds. Conclusion Our results suggest that transcriptional regulation of FAD2 and FAH12 genes maybe one of the mechanisms that contribute to a high level of ricinoleate accumulation in castor endosperm. The full-length cDNA library will be used to search for additional genes that affect ricinoleate accumulation in seed oils. Our EST sequences will also be useful to annotate the castor genome, which whole sequence is being generated by shotgun sequencing at

Infectious Maize rayado fino virus from Cloned cDNA.

Science.gov (United States)

Edwards, Michael C; Weiland, John J; Todd, Jane; Stewart, Lucy R

2015-06-01

A full-length cDNA clone was produced from a U.S. isolate of Maize rayado fino virus (MRFV), the type member of the genus Marafivirus within the family Tymoviridae. Infectivity of transcripts derived from cDNA clones was demonstrated by infection of maize plants and protoplasts, as well as by transmission via the known leafhopper vectors Dalbulus maidis and Graminella nigrifrons that transmit the virus in a persistent-propagative manner. Infection of maize plants through vascular puncture inoculation of seed with transcript RNA resulted in the induction of fine stipple stripe symptoms typical of those produced by wild-type MRFV and a frequency of infection comparable with that of the wild type. Northern and Western blotting confirmed the production of MRFV-specific RNAs and proteins in infected plants and protoplasts. An unanticipated increase in subgenomic RNA synthesis over levels in infected plants was observed in protoplasts infected with either wild-type or cloned virus. A conserved cleavage site motif previously demonstrated to function in both Oat blue dwarf virus capsid protein and tymoviral nonstructural protein processing was identified near the amino terminus of the MRFV replicase polyprotein, suggesting that cleavage at this site also may occur.

Automation of cDNA Synthesis and Labelling Improves Reproducibility

Directory of Open Access Journals (Sweden)

Daniel Klevebring

2009-01-01

Full Text Available Background. Several technologies, such as in-depth sequencing and microarrays, enable large-scale interrogation of genomes and transcriptomes. In this study, we asses reproducibility and throughput by moving all laboratory procedures to a robotic workstation, capable of handling superparamagnetic beads. Here, we describe a fully automated procedure for cDNA synthesis and labelling for microarrays, where the purification steps prior to and after labelling are based on precipitation of DNA on carboxylic acid-coated paramagnetic beads. Results. The fully automated procedure allows for samples arrayed on a microtiter plate to be processed in parallel without manual intervention and ensuring high reproducibility. We compare our results to a manual sample preparation procedure and, in addition, use a comprehensive reference dataset to show that the protocol described performs better than similar manual procedures. Conclusions. We demonstrate, in an automated gene expression microarray experiment, a reduced variance between replicates, resulting in an increase in the statistical power to detect differentially expressed genes, thus allowing smaller differences between samples to be identified. This protocol can with minor modifications be used to create cDNA libraries for other applications such as in-depth analysis using next-generation sequencing technologies.

A theory for fracture of polymeric gels

Science.gov (United States)

Mao, Yunwei; Anand, Lallit

2018-06-01

A polymeric gel is a cross-linked polymer network swollen with a solvent. If the concentration of the solvent or the deformation is increased to substantial levels, especially in the presence of flaws, then the gel may rupture. Although various theoretical aspects of coupling of fluid permeation with large deformation of polymeric gels are reasonably well-understood and modeled in the literature, the understanding and modeling of the effects of fluid diffusion on the damage and fracture of polymeric gels is still in its infancy. In this paper we formulate a thermodynamically-consistent theory for fracture of polymeric gels - a theory which accounts for the coupled effects of fluid diffusion, large deformations, damage, and also the gradient effects of damage. The particular constitutive equations for fracture of a gel proposed in our paper, contain two essential new ingredients: (i) Our constitutive equation for the change in free energy of a polymer network accounts for not only changes in the entropy, but also changes in the internal energy due the stretching of the Kuhn segments of the polymer chains in the network. (ii) The damage and failure of the polymer network is taken to occur by chain-scission, a process which is driven by the changes in the internal energy of the stretched polymer chains in the network, and not directly by changes in the configurational entropy of the polymer chains. The theory developed in this paper is numerically implemented in an open-source finite element code MOOSE, by writing our own application. Using this simulation capability we report on our study of the fracture of a polymeric gel, and some interesting phenomena which show the importance of the diffusion of the fluid on fracture response of the gel are highlighted.

Spot quantification in two dimensional gel electrophoresis image analysis: comparison of different approaches and presentation of a novel compound fitting algorithm

Science.gov (United States)

2014-01-01

Background Various computer-based methods exist for the detection and quantification of protein spots in two dimensional gel electrophoresis images. Area-based methods are commonly used for spot quantification: an area is assigned to each spot and the sum of the pixel intensities in that area, the so-called volume, is used a measure for spot signal. Other methods use the optical density, i.e. the intensity of the most intense pixel of a spot, or calculate the volume from the parameters of a fitted function. Results In this study we compare the performance of different spot quantification methods using synthetic and real data. We propose a ready-to-use algorithm for spot detection and quantification that uses fitting of two dimensional Gaussian function curves for the extraction of data from two dimensional gel electrophoresis (2-DE) images. The algorithm implements fitting using logical compounds and is computationally efficient. The applicability of the compound fitting algorithm was evaluated for various simulated data and compared with other quantification approaches. We provide evidence that even if an incorrect bell-shaped function is used, the fitting method is superior to other approaches, especially when spots overlap. Finally, we validated the method with experimental data of urea-based 2-DE of Aβ peptides andre-analyzed published data sets. Our methods showed higher precision and accuracy than other approaches when applied to exposure time series and standard gels. Conclusion Compound fitting as a quantification method for 2-DE spots shows several advantages over other approaches and could be combined with various spot detection methods. The algorithm was scripted in MATLAB (Mathworks) and is available as a supplemental file. PMID:24915860

Pseudomonas aeruginosa endophthalmitis after penetrating keratoplasty transmitted from the same donor to two recipients confirmed by pulsed-field gel electrophoresis.

Science.gov (United States)

Oguido, Ana Paula Miyagusko Taba; Casella, Antonio Marcelo Barbante; Hofling-Lima, Ana Luisa; Pacheco, Sergio Arruda; Bispo, Paulo José Martins; Marques, Fernanda

2011-09-01

Two devastating cases of multidrug-resistant Pseudomonas aeruginosa endophthalmitis after keratoplasty as the result of transmission from the same donor were confirmed by pulsed-field gel electrophoresis. Strategies for preventing donor-to-host transmission, such as the use of antimicrobial agents of greater efficacy and better methods for detecting microorganisms in preservation medium, could minimize this type of transmission.

Two-dimensional gel electrophoresis data in support of leaf comparative proteomics of two citrus species differing in boron-tolerance

Directory of Open Access Journals (Sweden)

Wen Sang

2015-09-01

Full Text Available Here, we provide the data from a comparative proteomics approach used to investigate the response of boron (B-tolerant ‘Xuegan’ (Citrus sinensis and B-intolerant ‘Sour pummelo’ (Citrus grandis leaves to B-toxicity. Using two-dimensional gel electrophoresis (2-DE technique, we identified 50 and 45 protein species with a fold change of more than 1.5 and a P-value of less than 0.05 from B-toxic C. sinensis and C. grandis leaves. These B-toxicity-responsive protein species were mainly involved in carbohydrate and energy metabolism, antioxidation and detoxification, stress responses, coenzyme biosynthesis, protein and amino acid metabolism, signal transduction, cell transport, cytoskeleton, nucleotide metabolism, and cell cycle and DNA processing. A detailed analysis of this data may be obtained from Sang et al. (J. Proteomics 114 (2015[1].

An Unbiased Unscented Transform Based Kalman Filter for 3D Radar

Institute of Scientific and Technical Information of China (English)

WANGGuohong; XIUJianjuan; HEYou

2004-01-01

As a derivative-free alternative to the Extended Kalman filter (EKF) in the framework of state estimation, the Unscented Kalman filter (UKF) has potential applications in nonlinear filtering. By noting the fact that the unscented transform is generally biased when converting the radar measurements from spherical coordinates into Cartesian coordinates, a new filtering algorithm for 3D radar, called Unbiased unscented Kalman filter (UUKF), is proposed. The new algorithm is validated by Monte Carlo simulation runs. Simulation results show that the UUKF is more effective than the UKF, EKF and the Converted measurement Kalman filter (CMKF).

Cloning of low dose radiation induced gene RIG1 by RACE based on non-cloned cDNA library

International Nuclear Information System (INIS)

Luo Ying; Sui Jianli; Tie Yi; Zhang Yuanping; Zhou Pingkun; Sun Zhixian

2001-01-01

Objective: To obtain full-length cDNA of radiation induced new gene RIG1 based on its EST fragment. Methods: Based on non-cloned cDNA library, enhanced nested RACE PCR and biotin-avidin labelled probe for magnetic bead purification was used to obtain full-length cDNA of RIG1. Results: About 1 kb of 3' end of RIG1 gene was successfully cloned by this set of methods and cloning of RIG1 5' end is proceeding well. Conclusion: The result is consistent with the design of experiment. This set of protocol is useful for cloning of full-length gene based on EST fragment

Ionic and viscoelastic mechanisms of a bucky-gel actuator

Science.gov (United States)

Kruusamäe, Karl; Sugino, Takushi; Asaka, Kinji

2015-07-01

Ionic electromechanically active polymers (IEAPs) are considered attractive candidates for soft, miniature, and lightweight actuators. The bucky-gel actuator is a carbonaceous subtype of IEAP that due to its structure (i.e. two highly porous electrodes sandwiching a thin ion-permeable electrolyte layer) and composition (i.e. being composed of soft porous polymer, carbon nanotubes, and ionic liquid) is very similar to an electric double-layer capacitor. In response to the voltage applied between the electrodes of a bucky-gel actuator, the laminar structure bends. The time domain behavior exhibits, however, a phenomenon called the back-relaxation, i.e., after some time the direction of bending is reversed even though voltage remains constant. In spite of the working mechanism of IEAP actuators being generally attributed to the transport of ions within the soft multilayer system, the specific details remain unclear. A so-called two-carrier model proposes that the bending and subsequent back-relaxation are caused by the relocation of two ionic species having different mobilities as they enter and exit the electrode layers. By adopting the two-carrier model for bucky-gel actuators, we see very good agreement between the mathematical representation and the experimental data of the electromechanical behavior. Furthermore, since the bucky-gel actuator is viscoelastic, we propose to use the time domain response of a blocking force as the key parameter related to the inner ionic mechanism. We also introduce a method to estimate the viscoelastic creep compliance function from the time domain responses for curvature and blocking force. This analysis includes four types of bucky-gel actuators of varying composition and structure.

EXPRESSION PROFILING OF ESTROGENIC COMPOUNDS USING A SHEEPSHEAD MINNOW CDNA MACROARRAY

Science.gov (United States)

Larkin, Patrick, Leroy C. Folmar, Michael J. Hemmer, Arianna J. Poston and Nancy D. Denslow. 2003. Expression Profiling of Estrogenic Compounds Using a Sheepshead Minnow cDNA Macroarray. Environ. Health Perspect. 111(6):839-846. (ERL,GB 1171). A variety of anthropogenic c...

Cloning and expression of a cDNA coding for a human monocyte-derived plasminogen activator inhibitor.

Science.gov (United States)

Antalis, T M; Clark, M A; Barnes, T; Lehrbach, P R; Devine, P L; Schevzov, G; Goss, N H; Stephens, R W; Tolstoshev, P

1988-02-01

Human monocyte-derived plasminogen activator inhibitor (mPAI-2) was purified to homogeneity from the U937 cell line and partially sequenced. Oligonucleotide probes derived from this sequence were used to screen a cDNA library prepared from U937 cells. One positive clone was sequenced and contained most of the coding sequence as well as a long incomplete 3' untranslated region (1112 base pairs). This cDNA sequence was shown to encode mPAI-2 by hybrid-select translation. A cDNA clone encoding the remainder of the mPAI-2 mRNA was obtained by primer extension of U937 poly(A)+ RNA using a probe complementary to the mPAI-2 coding region. The coding sequence for mPAI-2 was placed under the control of the lambda PL promoter, and the protein expressed in Escherichia coli formed a complex with urokinase that could be detected immunologically. By nucleotide sequence analysis, mPAI-2 cDNA encodes a protein containing 415 amino acids with a predicted unglycosylated Mr of 46,543. The predicted amino acid sequence of mPAI-2 is very similar to placental PAI-2 (3 amino acid differences) and shows extensive homology with members of the serine protease inhibitor (serpin) superfamily. mPAI-2 was found to be more homologous to ovalbumin (37%) than the endothelial plasminogen activator inhibitor, PAI-1 (26%). Like ovalbumin, mPAI-2 appears to have no typical amino-terminal signal sequence. The 3' untranslated region of the mPAI-2 cDNA contains a putative regulatory sequence that has been associated with the inflammatory mediators.

In Situ Formation of Steroidal Supramolecular Gels Designed for Drug Release

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Hana Bunzen

2013-03-01

Full Text Available In this work, a steroidal gelator containing an imine bond was synthesized, and its gelation behavior as well as a sensitivity of its gels towards acids was investigated. It was shown that the gels were acid-responsive, and that the gelator molecules could be prepared either by a conventional synthesis or directly in situ during the gel forming process. The gels prepared by both methods were studied and it was found that they had very similar macro- and microscopic properties. Furthermore, the possibility to use the gels as carriers for aromatic drugs such as 5-chloro-8-hydroxyquinoline, pyrazinecarboxamide, and antipyrine was investigated and the prepared two-component gels were studied with regard to their potential applications in drug delivery, particularly in a pH-controlled drug release.

Reproducibility and signal response linearity of Alanine gel dosimeter

International Nuclear Information System (INIS)

Silva, Cleber Feijo Silva; Campos, Leticia Lucente

2008-01-01

Gel Dosimetry has been studied mainly for medical applications, because it presents signal response in the dose range used in radiotherapy treatments and it can be applied for three dimensional dosimetry. Alanine gel dosimeter is a new gel material developed at IPEN that presents significant improvement on previous alanine systems developed by Costa (1994). The DL-Alanine (C 3 H 7 NO 2 ) is an amino acid tissue equivalent that improves the production of ferric ions in the solution. These ferric ions concentration can be measured by spectrophotometry technique. This work aims to study the reproducibility of the alanine gel solutions and the signal response as a function of gamma radiation dose, considering that these two properties are very important for characterizing and standardizing any dosimeter. (author)

Metal-silica sol-gel materials

Science.gov (United States)

Stiegman, Albert E. (Inventor)

2002-01-01

The present invention relates to a single phase metal-silica sol-gel glass formed by the co-condensation of a transition metal with silicon atoms where the metal atoms are uniformly distributed within the sol-gel glass as individual metal centers. Any transition metal may be used in the sol-gel glasses. The present invention also relates to sensor materials where the sensor material is formed using the single phase metal-silica sol-gel glasses. The sensor materials may be in the form of a thin film or may be attached to an optical fiber. The present invention also relates to a method of sensing chemicals using the chemical sensors by monitoring the chromatic change of the metal-silica sol-gel glass when the chemical binds to the sensor. The present invention also relates to oxidation catalysts where a metal-silica sol-gel glass catalyzes the reaction. The present invention also relates to a method of performing oxidation reactions using the metal-silica sol-gel glasses. The present invention also relates to organopolymer metal-silica sol-gel composites where the pores of the metal-silica sol-gel glasses are filled with an organic polymer polymerized by the sol-gel glass.

Study of nuclear glasses alteration gel and synthesis of some model gels; Etude du gel d`alteration des verres nucleaires et synthese de gels modeles

Energy Technology Data Exchange (ETDEWEB)

Ricol, S

1995-10-05

This work deals with the general problem of alteration of the reference nuclear glass R7T7. Attention is paid particularly to the altered layer formed at the glass surface during alteration process. In opposition to previous works, related essentially to glass dissolution kinetics based on chemical analyses of lixiviated elements, this thesis deals with alteration problems through structural studies of the reference glass and derived gel. This approach allows the determination of mechanisms for the gel formation and a better understanding of the behaviour of glasses towards lixiviation. Both approaches appeared complementary. Based on several spectroscopic techniques, this work showed the particular role of cations such as calcium, zirconium and iron. Studies of silica-based synthetic gels showed the synergic effect of formers cation and of one highly coordinated cation. The variation of the wavenumber related to Si-O-Si asymmetric stretching vibration can be correlated to O/Si ratio for ternary systems Si/Na/Zr. On the contrary, the Si losses of the materials depend on the number of non-bridging oxygen atoms. In the perspective of long-term behaviour, the alteration gel presents better characteristics than initial glass. It is therefore a highly stable material in static conditions. In the same way, synthetic gels are materials with very low solubilities (much lower than the alteration gel) and could be used as confining matrices. (authors). refs., 71 figs., 37 tabs.

Three human alcohol dehydrogenase subunits: cDNA structure and molecular and evolutionary divergence

International Nuclear Information System (INIS)

Ikuta, T.; Szeto, S.; Yoshida, A.

1986-01-01

Class I human alcohol dehydrogenase (ADH; alcohol:NAD + oxidoreductase, EC 1.1.1.1) consists of several homo- and heterodimers of α, β, and γ subunits that are governed by the ADH1, ADH2, and ADH3 loci. The authors previously cloned a full length of cDNA for the β subunit, and the complete sequence of 374 amino acid residues was established. cDNAs for the α and γ subunits were cloned and characterized. A human liver cDNA library, constructed in phage λgt11, was screened by using a synthetic oligonucleotide probe that was matched to the γ but not to the β sequence. Clone pUCADHγ21 and clone pUCADHα15L differed from β cDNA with respect to restriction sites and hybridization with the nucleotide probe. Clone pUCADHγ21 contained an insertion of 1.5 kilobase pairs (kbp) and encodes 374 amino acid residues compatible with the reported amino acid sequence of the γ subunit. Clone pUCADHα15L contained an insertion of 2.4 kbp and included nucleotide sequences that encode 374 amino acid residues for another subunit, the γ subunit. In addition, this clone contained the sequences that encode the COOH-terminal part of the β subunit at its extended 5' region. The amino acid sequences and coding regions of the cDNAs of the three subunits are very similar. A high degree of resemblance is observed also in their 3' noncoding regions. However, distinctive differences exist in the vicinity of the Zn-binding cysteine residue at position 46. Based on the cDNA sequences and the deduced amino acid sequences of the three subunits, their structural and evolutionary relationships are discussed

Thoria sol-gel processes

International Nuclear Information System (INIS)

Matthews, R.B.

1978-10-01

Alternate fuel fabrication techniques are being developed at WNRE as part of the thorium fuel cycle program. The sol-gel techniques are attractive and this report assembles and summarizes information relating to thoria sol-gel fuels. Some background information on the behaviour and advantages of sol-gel fuel forms is presented, followed by a review of relevant colloid chemistry and an explanation of the fundamental steps of sol-gel processes. Finally, several variants to the basic process are reviewed and evaluated. (author)

Review of Fricke gel dosimeters

International Nuclear Information System (INIS)

Schreiner, L J

2004-01-01

The innovation of adding a gel matrix to the traditional Fricke dosimeter to stabilize geometric information established the field of gel dosimetry for radiation therapy. A discussion of Fricke gels provides an overview of the issues that determine the dose response of all gel dosimeters in general. In this paper we review some of the features of Fricke systems to illustrate these issues and, in addition, to motivate renewed clinical interest in Fricke gels

Unbiased group-wise image registration: applications in brain fiber tract atlas construction and functional connectivity analysis.

Science.gov (United States)

Geng, Xiujuan; Gu, Hong; Shin, Wanyong; Ross, Thomas J; Yang, Yihong

2011-10-01

We propose an unbiased implicit-reference group-wise (IRG) image registration method and demonstrate its applications in the construction of a brain white matter fiber tract atlas and the analysis of resting-state functional MRI (fMRI) connectivity. Most image registration techniques pair-wise align images to a selected reference image and group analyses are performed in the reference space, which may produce bias. The proposed method jointly estimates transformations, with an elastic deformation model, registering all images to an implicit reference corresponding to the group average. The unbiased registration is applied to build a fiber tract atlas by registering a group of diffusion tensor images. Compared to reference-based registration, the IRG registration improves the fiber track overlap within the group. After applying the method in the fMRI connectivity analysis, results suggest a general improvement in functional connectivity maps at a group level in terms of larger cluster size and higher average t-scores.

Gel nano-particulates against radioactivity; Des nanoparticules en gel contre la radioactivite

Energy Technology Data Exchange (ETDEWEB)

Deroin, Ph

2004-11-01

The Argonne research center (USA) has developed a 'super-gel' compound, a polymer close to those used in baby's diapers, which can reach a 90% efficiency in the radioactive decontamination of porous materials, like bricks or concrete. The contaminated materials are sprayed with a mixture of polymer gel and wetting agent with nano-particulates in suspension. Under the action of the wetting agent, radioactivity migrates from the pores to the gel and is trapped by the nano-particulates. The drying and recycling of the gel allows to reduce the volume of radioactive wastes. Short paper. (J.S.)

SiO2 sol-gel films after ammonia and heat two-step treatments

International Nuclear Information System (INIS)

Zhang Chunlai; Wang Biyi; Tian Dongbin; Yin Wei; Jiang Xiaodong; Yuan Xiaodong; Yan Lianghong; Zhang Hongliang; Zhao Songnan; Lv Haibing

2008-01-01

SiO 2 thin films were deposited using tetraethoxylsilane as precursor, ammonia as catalyst on K9 glass by sol-gel method. These films were post-treated by ammonia and heat. The properties of the coatings were characterized by ellipsometer, UV-vis spectrophotometry, FTIR-spectroscopy, scanning probe microscope and contact angle measurement apparatus. The resuits indicate that the thickness of the films with ammonia and heat treatment tend to decrease. Both the refractive index and water contact angle increase after ammonia treatment. However, they both decrease after heat treatment. The former increases by 0.236 for the first step, then decreases by 0.202 for the second. The latter increases to 58.92 degree, then decreases to 38.07 degree. The transmittance of the coatings turn to be better and continuously shift to short wave by UV-vis spectrophotometry. The surface becomes smoother by AFM after the two-step treatment. (authors)

Construction and characterization of a cDNA library from human ...

African Journals Online (AJOL)

The tumor-suppressor gene p53 and its downstream genes consist of a complicated gene network, and the challenge to understand the network is to identify p53 downstream genes. In order to isolate and identify new p53 regulated genes, we constructed and characterized a normalized cDNA library from human brain ...

Nucleotide sequence of cloned cDNA for human sphingolipid activator protein 1 precursor

International Nuclear Information System (INIS)

Dewji, N.N.; Wenger, D.A.; O'Brien, J.S.

1987-01-01

Two cDNA clones encoding prepro-sphingolipid activator protein 1 (SAP-1) were isolated from a λ gt11 human hepatoma expression library using polyclonal antibodies. These had inserts of ≅ 2 kilobases (λ-S-1.2 and λ-S-1.3) and both were both homologous with a previously isolated clone (λ-S-1.1) for mature SAP-1. The authors report here the nucleotide sequence of the longer two EcoRI fragments of S-1.2 and S-1.3 that were not the same and the derived amino acid sequences of mature SAP-1 and its prepro form. The open reading frame encodes 19 amino acids, which are colinear with the amino-terminal sequence of mature SAP-1, and extends far beyond the predicted carboxyl terminus of mature SAP-1, indicating extensive carboxyl-terminal processing. The nucleotide sequence of cDNA encoding prepro-SAP-1 includes 1449 bases from the assigned initiation codon ATG at base-pair 472 to the stop codon TGA at base-pair 1921. The first 23 amino acids coded after the initiation ATG are characteristic of a signal peptide. The calculated molecular mass for a polypeptide encoded by 1449 bases is ≅ 53 kDa, in keeping with the reported value for pro-SAP-1. The data indicate that after removal of the signal peptide mature SAP-1 is generated by removing an additional 7 amino acids from the amino terminus and ≅ 373 amino acids from the carboxyl terminus. One potential glycosylation site was previously found in mature SAP-1. Three additional potential glycosylation sites are present in the processed carboxyl-terminal polypeptide, which they designate as P-2

From the Macro to the Micro: Gel Mapping to Differentiate between Sporozoites of Two Immunologically Distinct Strains of Eimeria maxima (Strains M6 and Guelph).

Science.gov (United States)

El-Ashram, Saeed; Yin, Qing; Liu, Hongbin; Al Nasr, Ibrahim; Liu, Xianyong; Suo, Xun; Barta, John

2015-01-01

Two immunologically distinct strains of E. maxima were examined in this study: the M6 strain and the Guelph strain. The differential expression between the sporozoites of the two strains of E. maxima was determined by image analysis of 100 μg of protein from each strain separated by standard one- and conventional two-dimensional polyacrylamide gel electrophoresis. In addition to differences in both molecular weight and the electrophoretic mobility, differences in the intensity of polypeptide bands for example, GS 136.4 and M6 169 were explored. Pooled gels were prepared from each strain. A representative 2D-PAGE gel spanning a non-linear pH range of 3-10 of E. maxima strain M6 consisted of approximately 694 polypeptide spots with about 67 (9.6%) of the polypeptide spots being unique relative to the other strain. E. maxima strain GS had about 696 discernable polypeptide spots with 69 spots (9.9%) that differed from those of the M6 strain. In-depth characterization of the variable polypeptide spots; unique polypeptide spots (absence or presence) and shared polypeptide spots with modifications may lead to novel vaccine target in the form of multi-component, multi-stage, multi-immunovariant strains, multi-species subunit vaccine, and diagnostic probe for E. maxima.

Automatic and quantitative measurement of collagen gel contraction using model-guided segmentation

Science.gov (United States)

Chen, Hsin-Chen; Yang, Tai-Hua; Thoreson, Andrew R.; Zhao, Chunfeng; Amadio, Peter C.; Sun, Yung-Nien; Su, Fong-Chin; An, Kai-Nan

2013-08-01

Quantitative measurement of collagen gel contraction plays a critical role in the field of tissue engineering because it provides spatial-temporal assessment (e.g., changes of gel area and diameter during the contraction process) reflecting the cell behavior and tissue material properties. So far the assessment of collagen gels relies on manual segmentation, which is time-consuming and suffers from serious intra- and inter-observer variability. In this study, we propose an automatic method combining various image processing techniques to resolve these problems. The proposed method first detects the maximal feasible contraction range of circular references (e.g., culture dish) and avoids the interference of irrelevant objects in the given image. Then, a three-step color conversion strategy is applied to normalize and enhance the contrast between the gel and background. We subsequently introduce a deformable circular model which utilizes regional intensity contrast and circular shape constraint to locate the gel boundary. An adaptive weighting scheme was employed to coordinate the model behavior, so that the proposed system can overcome variations of gel boundary appearances at different contraction stages. Two measurements of collagen gels (i.e., area and diameter) can readily be obtained based on the segmentation results. Experimental results, including 120 gel images for accuracy validation, showed high agreement between the proposed method and manual segmentation with an average dice similarity coefficient larger than 0.95. The results also demonstrated obvious improvement in gel contours obtained by the proposed method over two popular, generic segmentation methods.

Cloning of the cDNA for murine von Willebrand factor and identification of orthologous genes reveals the extent of conservation among diverse species.

Science.gov (United States)

Chitta, Mohan S; Duhé, Roy J; Kermode, John C

2007-05-01

Interaction of von Willebrand factor (VWF) with circulating platelets promotes hemostasis when a blood vessel is injured. The A1 domain of VWF is responsible for the initial interaction with platelets and is well conserved among species. Knowledge of the cDNA and genomic DNA sequences for human VWF allowed us to predict the cDNA sequence for murine VWF in silico and amplify its entire coding region by RT-PCR. The murine VWF cDNA has an open reading frame of 8,442 bp, encoding a protein of 2,813 amino acid residues with 83% identity to human pre-pro-VWF. The same strategy was used to predict in silico the cDNA sequence for the ortholog of VWF in a further six species. Many of these predictions diverged substantially from the putative Reference Sequences derived by ab initio methods. Our predicted sequences indicated that the VWF gene has a conserved structure of 52 exons in all seven mammalian species examined, as well as in the chicken. There is a minor structural variation in the pufferfish Takifugu rubripes insofar as the VWF gene in this species has 53 exons. Comparison of the translated amino acid sequences also revealed a high degree of conservation. In particular, the cysteine residues are conserved precisely throughout both the pro-peptide and the mature VWF sequence in all species, with a minor exception in the pufferfish VWF ortholog where two adjacent cysteine residues are omitted. The marked conservation of cysteine residues emphasizes the importance of the intricate pattern of disulfide bonds in governing the structure of pro-VWF and regulating the function of the mature VWF protein. It should also be emphasized that many of the conserved features of the VWF gene and protein were obscured when the comparison among species was based on the putative Reference Sequences instead of our predicted cDNA sequences.

Large strain deformation behavior of polymeric gels in shear- and cavitation rheology

Science.gov (United States)

Hashemnejad, Seyed Meysam; Kundu, Santanu

Polymeric gels are used in many applications including in biomedical and in food industries. Investigation of mechanical responses of swollen polymer gels and linking that to the polymer chain dynamics are of significant interest. Here, large strain deformation behavior of two different gel systems and with different network architecture will be presented. We consider biologically relevant polysaccharide hydrogels, formed through ionic and covalent crosslinking, and physically associating triblock copolymer gels in a midblock selective solvent. Gels with similar low-strain shear modulus display distinctly different non-linear rheological behavior in large strain shear deformation. Both these gels display strain-stiffening behavior in shear-deformation prior to macroscopic fracture of the network, however, only the alginate gels display negative normal stress. The cavitation rheology data show that the critical pressure for cavitation is higher for alginate gels than that observed for triblock gels. These distinctly different large-strain deformation behavior has been related to the gel network structure, as alginate chains are much stiffer than the triblock polymer chains.

Two dimensional gel electrophoresis using narrow pH 3-5.6 immobilised pH gradient strips identifies potential novel disease biomarkers in plasma or serum

OpenAIRE

sprotocols

2015-01-01

Authors: Bevin Gangadharan & Nicole Zitzmann ### Abstract Two-dimensional gel electrophoresis (2-DE) is a protein separation technique often used to separate plasma or serum proteins in an attempt to identify novel biomarkers. This protocol describes how to run 2-DE gels using narrow pH 3-5.6 immobilised pH gradient strips to separate 2 mg of serum proteins. pH 3-6 ampholytes are used to enhance the solubility of proteins in this pH range before the serum proteins are separated in the...

Design of Autonomous Gel Actuators

Directory of Open Access Journals (Sweden)

Shuji Hashimoto

2011-01-01

Full Text Available In this paper, we introduce autonomous gel actuators driven by chemical energy. The polymer gels prepared here have cyclic chemical reaction networks. With a cyclic reaction, the polymer gels generate periodical motion. The periodic motion of the gel is produced by the chemical energy of the oscillatory Belouzov-Zhabotinsky (BZ reaction. We have succeeded in making synthetic polymer gel move autonomously like a living organism. This experimental fact represents the great possibility of the chemical robot.

Molecular characterization of a Leishmania donovani cDNA clone with similarity to human 20S proteasome a-type subunit

DEFF Research Database (Denmark)

Christensen, C B; Jørgensen, L; Jensen, A T

2000-01-01

Using plasma from patients infected or previously infected with Leishmania donovanii, we isolated a L. donovanii cDNA clone with similarity to the proteasome a-type subunit from humans and other eukaryotes. The cDNA clone, designated LePa, was DNA sequenced and Northern blot analysis of L....... donovanii poly(A(+))mRNA indicated the isolation of a full length cDNA clone with a transcript size of 1.9 kb. The expressed recombinant LePa fusion protein induced proliferation of peripheral blood mononuclear cells in one out of seven patients who had suffered from visceral leishmaniasis. Plasma from 16...

Characterization of alkali silica reaction gels using Raman spectroscopy

International Nuclear Information System (INIS)

Balachandran, C.; Muñoz, J.F.; Arnold, T.

2017-01-01

The ability of Raman spectroscopy to characterize amorphous materials makes this technique ideal to study alkali silica reaction (ASR) gels. The structure of several synthetic ASR gels was thoroughly characterized using Raman Spectroscopy. The results were validated with additional techniques such as Fourier transmission infrared spectroscopy, X-ray powder diffraction and thermogravimetric analysis. The Raman spectra were found to have two broad bands in the 800 to 1200 cm −1 range and the 400 to 700 cm −1 range indicating the amorphous nature of the gel. Important information regarding the silicate polymerization was deduced from both of these spectral regions. An increase in alkali content of the gels caused a depolymerization in the silicate framework which manifested in the Raman spectra as a gradual shift of predominant peaks in both regions. The trends in silicate depolymerization were in agreement with results from a NMR spectroscopy study on similar synthetic ASR gels.

Optimal processing for gel electrophoresis images: Applying Monte Carlo Tree Search in GelApp.

Science.gov (United States)

Nguyen, Phi-Vu; Ghezal, Ali; Hsueh, Ya-Chih; Boudier, Thomas; Gan, Samuel Ken-En; Lee, Hwee Kuan

2016-08-01

In biomedical research, gel band size estimation in electrophoresis analysis is a routine process. To facilitate and automate this process, numerous software have been released, notably the GelApp mobile app. However, the band detection accuracy is limited due to a band detection algorithm that cannot adapt to the variations in input images. To address this, we used the Monte Carlo Tree Search with Upper Confidence Bound (MCTS-UCB) method to efficiently search for optimal image processing pipelines for the band detection task, thereby improving the segmentation algorithm. Incorporating this into GelApp, we report a significant enhancement of gel band detection accuracy by 55.9 ± 2.0% for protein polyacrylamide gels, and 35.9 ± 2.5% for DNA SYBR green agarose gels. This implementation is a proof-of-concept in demonstrating MCTS-UCB as a strategy to optimize general image segmentation. The improved version of GelApp-GelApp 2.0-is freely available on both Google Play Store (for Android platform), and Apple App Store (for iOS platform). © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Sol-gel encapsulation for controlled drug release and biosensing

Science.gov (United States)

Fang, Jonathan

The main focus of this dissertation is to investigate the use of sol-gel encapsulation of biomolecules for controlled drug release and biosensing. Controlled drug release has advantages over conventional therapies in that it maintains a constant, therapeutic drug level in the body for prolonged periods of time. The anti-hypertensive drug Captopril was encapsulated in sol-gel materials of various forms, such as silica xerogels and nanoparticles. The primary objective was to show that sol-gel silica materials are promising drug carriers for controlled release by releasing Captopril at a release rate that is within a therapeutic range. We were able to demonstrate desired release for over a week from Captopril-doped silica xerogels and overall release from Captopril-doped silica nanoparticles. As an aside, the antibiotic Vancomycin was also encapsulated in these porous silica nanoparticles and desired release was obtained for several days in-vitro. The second part of the dissertation focuses on immobilizing antibodies and proteins in sol-gel to detect various analytes, such as hormones and amino acids. Sol-gel competitive immunoassays on antibody-doped silica xerogels were used for hormone detection. Calibration for insulin and C-peptide in standard solutions was obtained in the nM range. In addition, NASA-Ames is also interested in developing a reagentless biosensor using bacterial periplasmic binding proteins (bPBPs) to detect specific biomarkers, such as amino acids and phosphate. These bPBPs were doubly labeled with two different fluorophores and encapsulated in silica xerogels. Ligand-binding experiments were performed on the bPBPs in solution and in sol-gel. Ligand-binding was monitored by fluorescence resonance energy transfer (FRET) between the two fluorophores on the bPBP. Titration data show that one bPBP has retained its ligand-binding properties in sol-gel.

cDNA, genomic sequence cloning and analysis of the ribosomal ...

African Journals Online (AJOL)

Ribosomal protein L37A (RPL37A) is a component of 60S large ribosomal subunit encoded by the RPL37A gene, which belongs to the family of ribosomal L37AE proteins, located in the cytoplasm. The complementary deoxyribonucleic acid (cDNA) and the genomic sequence of RPL37A were cloned successfully from giant ...

Observation of intermittency in gene expression on cDNA microarrays

CERN Document Server

Peterson, L E

2002-01-01

We used scaled factorial moments to search for intermittency in the log expression ratios (LERs) for thousands of genes spotted on cDNA microarrays (gene chips). Results indicate varying levels of intermittency in gene expression. The observation of intermittency in the data analyzed provides a complimentary handle on moderately expressed genes, generally not tackled by conventional techniques.

cDNA sequence and tissue expression analysis of glucokinase from ...

African Journals Online (AJOL)

Yomi

2012-01-10

Jan 10, 2012 ... distribution of GK mRNA in brain, mesenteric adipose tissue, spleen, white muscle and liver of grass ... expression profile of GK mRNA in liver normalized with β-actin level was 31, 454 and 649-fold compared .... Primers and expected products used for GK gene cDNA RT-PCR, RACE and real-time PCR.

Disiloxanes and Functionalized Silica Gels: One Route, Two Complementary Outcomes-Guanidinium and Pyridinium Ion-Exchangers.

Directory of Open Access Journals (Sweden)

Łukasz Tabisz

Full Text Available Five novel disiloxane compounds comprising guanidinium and pyridinium moieties were obtained with high yields and purity. The verified synthetic pathways were then applied for modification of pre-functionalized silica gel, producing materials with the analogous organic side-chains. These halide-containing compounds and materials were then compared as to their ion-exchange properties: two disiloxanes proved to be effective in leaching different anions (nitrate, benzoate and ascorbate from solid to organic phase, and pyridinium-functionalized silica gels showed selectivity towards perchlorate ion, removing it from methanolic solutions with preference to other singly charged anions. The results presented demonstrate that both compounds and materials containing silicon-carbon bonds can be produced using the same methodology, but offer strikingly different application opportunities. Comparison of their properties provides additional insight into the binding mode of different anions and hints at how the transition from a flexible siloxane bridge to immobilization on solid surface influences anion-binding selectivity. Additionally, one of the siloxane dipodands was found to form a crystalline and poorly soluble nitrate salt (1.316 g/L, water, although it was miscible with a wide range of solvents as a hydrochloride. A possible explanation is given with the help of semi-empirical calculations. A simple, time- and cost-efficient automated potentiometric titration methodology was used as a viable analytical tool for studying ion-exchange processes for both compounds and materials, in addition to standard NMR, FT-IR and ESI-MS methods.

Two-dimensional gel electrophoresis and FTIR spectroscopy reveal both forms of yeast plasma membrane H(+)-ATPase in activated and basal-level enzyme preparations

Czech Academy of Sciences Publication Activity Database

Lapathitis, Georgios; Tanfani, F.; Kotyk, Arnošt; Bertoli, E.

2001-01-01

Roč. 505, č. 1 (2001), s. 155-158 ISSN 0014-5793 R&D Projects: GA ČR GA204/98/0474 Keywords : H+-ATPase * plasma membrane * two-dimensional gel electrophoresis Subject RIV: CE - Biochemistry Impact factor: 3.644, year: 2001

Comparative two-dimensional gel analysis and microsequencing identifies gelsolin as one of the most prominent downregulated markers of transformed human fibroblast and epithelial cells

DEFF Research Database (Denmark)

Vandekerckhove, J; Bauw, G; Vancompernolle, K

1990-01-01

A systematic comparison of the protein synthesis patterns of cultured normal and transformed human fibroblasts and epithelial cells, using two-dimensional gel protein analysis combined with computerized imaging and data acquisition, identified a 90-kD protein (SSP 5714) as one of the most striking...... downregulated markers typical of the transformed state. Using the information stored in the comprehensive human cellular protein database, we found this protein strongly expressed in several fetal tissues and one of them, epidermis, served as a source for preparative two-dimensional gel electrophoresis. Partial...... and by coelectrophoresis with purified human gelsolin. These results suggest that an important regulatory protein of the microfilament system may play a role in defining the phenotype of transformed human fibroblast and epithelial cells in culture. Udgivelsesdato: 1990-Jul...

Comparative two-dimensional gel analysis and microsequencing identifies gelsolin as one of the most prominent downregulated markers of transformed human fibroblast and epithelial cells

DEFF Research Database (Denmark)

Vandekerckhove, J; Bauw, G; Vancompernolle, K

1990-01-01

downregulated markers typical of the transformed state. Using the information stored in the comprehensive human cellular protein database, we found this protein strongly expressed in several fetal tissues and one of them, epidermis, served as a source for preparative two-dimensional gel electrophoresis. Partial......A systematic comparison of the protein synthesis patterns of cultured normal and transformed human fibroblasts and epithelial cells, using two-dimensional gel protein analysis combined with computerized imaging and data acquisition, identified a 90-kD protein (SSP 5714) as one of the most striking...... and by coelectrophoresis with purified human gelsolin. These results suggest that an important regulatory protein of the microfilament system may play a role in defining the phenotype of transformed human fibroblast and epithelial cells in culture. Udgivelsesdato: 1990-Jul...

Some properties and cDNA cloning of proteinaceous toxins from two species of lionfish (Pterois antennata and Pterois volitans).

Science.gov (United States)

Kiriake, Aya; Shiomi, Kazuo

2011-11-01

Lionfish, members of the genera Pterois, Parapterois and Dendrochirus, are well known to be venomous, having venomous glandular tissues in dorsal, pelvic and anal spines. The lionfish toxins have been shown to cross-react with the stonefish toxins by neutralization tests using the commercial stonefish antivenom, although their chemical properties including structures have been little characterized. In this study, an antiserum against neoverrucotoxin, the stonefish Synanceia verrucosa toxin, was first raised in a guinea pig and used in immunoblotting and inhibition immunoblotting to confirm that two species of Pterois lionfish (P. antennata and P. volitans) contain a 75kDa protein (corresponding to the toxin subunit) cross-reacting with neoverrucotoxin. Then, the amino acid sequences of the P. antennata and P. volitans toxins were successfully determined by cDNA cloning using primers designed from the highly conserved sequences of the stonefish toxins. Notably, either α-subunits (699 amino acid residues) or β-subunits (698 amino acid residues) of the P. antennata and P. volitans toxins share as high as 99% sequence identity with each other. Furthermore, both α- and β-subunits of the lionfish toxins exhibit high sequence identity (70-80% identity) with each other and also with the β-subunits of the stonefish toxins. As reported for the stonefish toxins, the lionfish toxins also contain a B30.2/SPRY domain (comprising nearly 200 amino acid residues) in the C-terminal region of each subunit. Copyright © 2011 Elsevier Ltd. All rights reserved.

Isolation, nucleotide sequence and expression of a cDNA encoding feline granulocyte colony-stimulating factor.

Science.gov (United States)

Dunham, S P; Onions, D E

2001-06-21

A cDNA encoding feline granulocyte colony stimulating factor (fG-CSF) was cloned from alveolar macrophages using the reverse transcriptase-polymerase chain reaction. The cDNA is 949 bp in length and encodes a predicted mature protein of 174 amino acids. Recombinant fG-CSF was expressed as a glutathione S-transferase fusion and purified by affinity chromatography. Biological activity of the recombinant protein was demonstrated using the murine myeloblastic cell line GNFS-60, which showed an ED50 for fG-CSF of approximately 2 ng/ml. Copyright 2001 Academic Press.

cDNA cloning of human DNA topoisomerase I. Catalytic activity of a 67.7-kDa carboxyl-terminal fragment

International Nuclear Information System (INIS)

D'Arpa, P.; Machlin, P.S.; Ratrie, H. III; Rothfield, N.F.; Cleveland, D.W.; Earnshaw, W.C.

1988-01-01

cDNA clones encoding human topoisomerase I were isolated from an expression vector library (λgt11) screened with autoimmune anti-topoisomerase I serum. One of these clones has been expressed as a fusion protein comprised of a 32-kDa fragment of the bacterial TrpE protein linked to 67.7 kDa of protein encoded by the cDNA. Three lines of evidence indicate that the cloned cDNA encodes topoisomerase I. (i) Proteolysis maps of the fusion protein and human nuclear topoisomerase I are essentially identical. (ii) The fusion protein relaxes supercoiled DNA, an activity that can be immunoprecipitated by anti-topoisomerase I serum. (iii) Sequence analysis has revealed that the longest cDNA clone (3645 base pairs) encodes a protein of 765 amino acids that shares 42% identity with Saccharomyces cerevisiae topoisomerase I. The sequence data also show that the catalytically active 67.7-kDa fragment is comprised of the carboxyl terminus

Characterization of cDNA encoding human placental anticoagulant protein (PP4): Homology with the lipocortin family

International Nuclear Information System (INIS)

Grundmann, U.; Abel, K.J.; Bohn, H.; Loebermann, H.; Lottspeich, F.; Kuepper, H.

1988-01-01

A cDNA library prepared from human placenta was screened for sequences encoding the placental protein 4 (PP4). PP4 is an anticoagulant protein that acts as an indirect inhibitor of the thromboplastin-specific complex, which is involved in the blood coagulation cascade. Partial amino acid sequence information from PP4-derived cyanogen bromide fragments was used to design three oligonucleotide probes for screening the library. From 10 6 independent recombinants, 18 clones were identified that hybridized to all three probes. These 18 recombinants contained cDNA inserts encoding a protein of 320 amino acid residues. In addition to the PP4 cDNA the authors identified 9 other recombinants encoding a protein with considerable similarity (74%) to PP4, which was termed PP4-X. PP4 and PP4-X belong to the lipocortin family, as judged by their homology to lipocortin I and calpactin I

Nucleotide sequence of a human cDNA encoding a ras-related protein (rap1B)

Energy Technology Data Exchange (ETDEWEB)

Pizon, V; Lerosey, I; Chardin, P; Tavitian, A [INSERM, Paris (France)

1988-08-11

The authors have previously characterized two human ras-related genes rap1 and rap2. Using the rap1 clone as probe they isolated and sequenced a new rap cDNA encoding the 184aa rap1B protein. The rap1B protein is 95% identical to rap1 and shares several properties with the ras protein suggesting that it could bind GTP/GDP and have a membrane location. As for rap1, the structural characteristics of rap1B suggest that the rap and ras proteins might interact on the same effector.

Characterization of a human MSX-2 cDNA and its fragment isolated as a transformation suppressor gene against v-Ki-ras oncogene.

Science.gov (United States)

Takahashi, C; Akiyama, N; Matsuzaki, T; Takai, S; Kitayama, H; Noda, M

1996-05-16

A cDNA (termed CT124) encoding a carboxyl-terminal fragment of the human homeobox protein MSX-2 was found to induce flat reversion when expressed in v-Ki-ras-transformed NIH3T3 cells. Although the expression of endogenous MSX-2 gene is low in most of the normal adult tissues examined, it is frequently activated in carcinoma-derived cell lines. Likewise, the gene is inactive in NIH3T3 cells but is transcriptionally activated after transformation by v-Ki-ras oncogene, suggesting that the intact MSX-2 may play a positive, rather than suppressive, role in cell transformation. To test this possibility, we isolated a near full-length human MSX-2 cDNA and tested its activities in two cell systems, i.e. fibroblast and myoblast. In NIH3T3 fibroblasts, although the gene by itself failed to confer a transformed phenotype, antisense MSX-2 cDNA as well as truncated CT124 cDNA interfered with the transforming activities of v-Ki-ras oncogene. In C2C12 myoblasts, MSX-2 was found to suppress MyoD gene expression, as do activated ras oncogenes, under certain culture conditions, and CT124 was found to inhibit the activities of both MSX-2 and ras in this system as well. Our findings not only suggest that CT124 may act as a dominant suppressor of MSX-2 but also raise the possibility that MSX-2 gene may be an important downstream target for the Ras signaling pathways.

cDNA cloning and characterization of mouse DTEF-1 and ETF, members of the TEA/ATTS family of transcription factors.

Science.gov (United States)

Yockey, C E; Shimizu, N

1998-02-01

Members of the TEA/ATTS family of transcription factors have been found in most representative eukaryotic organisms. In vertebrates, the TEA family contains at least four members, which share overlapping DNA-binding specificity and have similar transcriptional activation properties. In this article, we describe the cDNA cloning and characterization of the murine TEA proteins DTEF-1 (mDTEF-1) and ETF. Using in situ hybridization analysis of mouse embryos, we found that mDTEF-1 and ETF transcript distributions substantially overlap. ETF is expressed throughout the embryo except in the myocardium early in development, whereas late in development, it is enriched in lung and neuroectoderm. Mouse DTEF-1 is expressed at a much lower level throughout development and is substantially enriched in ectoderm and skin, as well as in the developing pituitary at midgestation. Northern blot analysis of adult mouse tissue total RNA showed that both ETF and mDTEF-1 are abundant in uterus and lung relative to other tissues. Using gel mobility shift assays and GAL4-fusion protein analysis, we demonstrated that the full coding sequences of ETF and mDTEF-1 encode M-CAT/GT-IIC-binding proteins containing activation domains.

Polymer gels and networks

National Research Council Canada - National Science Library

Osada, Yoshihito; Khokhlov, A. R

2002-01-01

... or magnetic field, etc.). It was realized that not only can polymer gels absorb and hold a considerable volume of liquids, but they can also be forced to expel the absorbed liquid in a controlled manner. Of particular interest are hydrogels, i.e., polymer gels, which swell extensively in water. The most common hydrogels are polyelectrolyte gels: ...

Amplification of a transcriptionally active DNA sequence in the human brain

International Nuclear Information System (INIS)

Yakovlev, A.G.; Sazonov, A.E.; Spunde, A.Ya.; Gindilis, V.M.

1986-01-01

The authors present their findings of tissue-specific amplification of a DNA fragment actively transcribed in the human brain. This genome fragment was found in the library complement of cDNA of the human brain and evidently belongs to a new class of moderate repetitions of DNA with an unstable copying capacity in the human genome. The authors isolated total cell RNA from various human tissues (brain, placenta), and rat tissues (brain, liver), by the method of hot phenol extraction with guanidine thiocynate. The poly(A + ) RNA fraction was isolated by chromatography. Synthesis of cDNA was done on a matrix of poly(A + ) RNA of human brain. The cDNA obtained was cloned in plasmid pBR322 for the PstI site using (dC/dG) sequences synthesized on the 3' ends of the vector molecule and cDNA respectively. In cloning 75 ng cDNA, the authors obtained approximately 10 5 recombinant. This library was analyzed by the hybridization method on columns with two radioactive ( 32 P) probes: the total cDNA preparation and the total nuclear DNA from the human brain. The number of copies of the cloned DNA fragment in the genome was determined by dot hybridization. Restricting fragments of human and rat DNA genomes homologous to the cloned cDNA were identified on radio-autographs. In each case, 10 micrograms of EcoRI DNA hydrolyzate was fractionated in 1% agarose gel. The probe was also readied with RNA samples fractionated in agarose gel with formaldehyde and transferred to a nitrocellulose filter under weak vacuum. The filter was hybridized with 0.1 micrograms DNA pAG 02, labeled with ( 32 P) to a specific activity of 0.5-1 x 10 9 counts/min x microgram. The autograph was exposed with amplifying screens at -70 0 C for 2 days

Preparation of continuous alumina gel fibres by aqueous sol–gel ...

Indian Academy of Sciences (India)

Abstract. Continuous alumina gel fibres were prepared by sol–gel method. The spinning sol was prepared by mixing aluminum nitrate, lactic acid and polyvinylpyrrolidone with a mass ratio of 10:3:1·5. Thermogravimetry– differential scanning calorimetry (TG–DSC), Fourier transform infrared (FTIR) spectra, X-ray diffraction ...

cDNA cloning, mRNA distribution and heterogeneity, chromosomal location, and RFLP analysis of human osteopontin (OPN)

DEFF Research Database (Denmark)

Young, M F; Kerr, J M; Termine, J D

1990-01-01

A human osteopontin (OP) cDNA was isolated from a library made from primary cultures of human bone cells. The distribution of osteopontin mRNA in human tissues was investigated by Northern analysis and showed that the human message was predominant in cultures of bone cells and in decidua cells...... osteopontin cDNA indicated that the gene is a single copy with an approximate length of 5.4-8.2 kb....

Diffusion measurement in ferrous infused gel dosimeters

International Nuclear Information System (INIS)

Zahmatkesh, M. H.; Healy, B. J.

2003-01-01

Background: The compositions of Ferrous sulphate, Agarose and Xylenol orange dye and Ferrous sulphate, Gelatin and Xylenol orange dye in solution of distilled water and sulphuric acid are two tissue-equivalent gel dosimeters. Ionizing radiation causes oxidation of Fe 2+ ion to Fe 3+ ions which diffuse through the gel matrix and blur the image of absorbed dose over a period of hours after irradiation. Materials and methods: 25 m M sulphuric acid, 0.4 mm ferrous ammonium sulphate, 0.2 mm xylenol orange dye and 1% by weight agarose in distilled water named Agarose and Xylenol orange dye and 0.1 mm ferrous ammonium sulphate, 0.1 mm xylenol orange dye, 50 mm sulphuric acid and 5% by weight gelatin in distilled water named Gelatin and Xylenol orange dye are used as two gel dosimeters. All chemicals were supplied by Sigma Ald ridge Company, Germany. The gels were poured in Perspex casts and were irradiated to a beam of X ray from linear accelerators or X ray machine. Results: In this study diffusion coefficients of Agarose and Xylenol orange dye and Gelatin and Xylenol orange dye dosimeters have been measured through a computer program for different temperature. The ferric ion diffusion coefficient (D) for the Agarose and Xylenol orange dye and Gelatin and Xylenol orange dye dosimeters were measured as (1.19.±0.03) x 10 -2 cm 2 .hr -1 and (0.83±0.03) x 10 -2 cm 2 .hr -1 respectively at room temperature. Conclusion: For both dosimeters the diffusion coefficients decreased with gel storage temperatures down to 6 d ig C . Gelatin and Xylenol orange dye dosimeters have advantage of lower diffusion coefficient for a specified temperature

Isolation of stress responsive Psb A gene from rice (Oryza sativa l.) using differential display.

Science.gov (United States)

Tyagi, Aruna; Chandra, Arti

2006-08-01

Differential display (DD) experiments were performed on drought-tolerant rice (Oryza sativa L.) genotype N22 to identify both upregulated and downregulated partial cDNAs with respect to moisture stress. DNA polymorphism was detected between drought-stressed and control leaf tissues on the DD gels. A partial cDNA showing differential expression, with respect to moisture stress was isolated from the gel. Northern blotting analysis was performed using this cDNA as a probe and it was observed that mRNA corresponding to this transcript was accumulated to high level in rice leaves under water deficit stress. At the DNA sequence level, the partial cDNA showed homology with psb A gene encoding for Dl protein.

Development and evaluation of exemestane-loaded lyotropic liquid crystalline gel formulations

OpenAIRE

Musa, Muhammad Nuh; David, Sheba Rani; Zulkipli, Ihsan Nazurah; Mahadi, Abdul Hanif; Chakravarthi, Srikumar; Rajabalaya, Rajan

2017-01-01

Introduction: The use of liquid crystalline (LC) gel formulations for drug delivery has considerably improved the current delivery methods in terms of bioavailability and efficacy. The purpose of this study was to develop and evaluate LC gel formulations to deliver the anti-cancer drug exemestane through transdermal route. Methods: Two LC gel formulations were prepared by phase separation coacervation method using glyceryl monooleate (GMO), Tween 80 and Pluronic® F127 (F127). The formulations...

Sealing wells with gel

Energy Technology Data Exchange (ETDEWEB)

Lopez, E C

1967-10-01

A new system is being used in Mexico to temporarily plug producing wells. The temporary seal is a gel with a catalyst. The use of this temporary plug allows gas-lift wells to be taken off production in order to carry out emergency repairs. The gel solidifies by the action of the catalyst to a high temperature (70 - 150/sup 0/C). By locating the bottom of the tubing at the top of the production interval, the gel material will go into the permeable formation, and immediately set. When the gel has solidified, it seals off the horizon that must not be stimulated, and leaves the others exposed to the acid action. When the treatment is finished, the gel, by action of the catalyst, is liquefied and removed from the formation, being produced with the oil.

Modelling of erosion of bentonite gel by gel/sol flow

Energy Technology Data Exchange (ETDEWEB)

Moreno, Luis; Neretnieks, Ivars; Longcheng Liu (Chemical Engineering and Technology, School of Chemical Science and Engineering, Royal Institute of Technology, Stockholm (Sweden))

2010-11-15

Bentonite intrusion into a fracture intersecting the canister deposition hole is modelled. The model describes the expansion of the bentonite within the fracture. It accounts for the repulsive electrostatic double-layer forces, the attractive van der Waals forces and friction forces between the particles and the water. The model also takes into account the diffusion of the colloid particles in the smectite sol. Diffusion of a counterion, sodium, is accounted for as this strongly influences the double layer force and the viscosity of the gel/sol. The gel/sol is considered to be a fluid with a varying viscosity that is strongly dependent on the bentonite volume fraction in the gel and the sodium concentration in the water. Two different geometries were modelled; a rectangular and a cylindrical. The rectangular geometry was used to gain experience with the processes and mechanisms and how they interact since the cylindrical geometry was somewhat less stable numerically and more time consuming. In the rectangular geometry a fracture 1 metre long in the flow direction was modelled. In both geometries the fracture size was selected sufficiently large to ensure that the water velocity, near the distant border was nearly the same as the approaching water velocity and that the smectite concentration there was vanishingly small. It was found that the velocity of the fluid drops considerably where the bentonite volume fraction is larger than 1-2%. This is due to the strong increase in viscosity with increasing bentonite volume fraction. The loss of smectite by the slowly flowing fluid was found to be proportional to the square root of the seeping water velocity for the rectangular geometry. For the cylindrical geometry, the dependence is somewhat lower (exponent about 0.4) since the length of the gel/water interface decreases with increasing water flow rate. The penetration depth of the gel/water interface decreases with increasing water flow rate. For water velocity of the

Modelling of erosion of bentonite gel by gel/sol flow

International Nuclear Information System (INIS)

Moreno, Luis; Neretnieks, Ivars; Longcheng Liu

2010-11-01

Bentonite intrusion into a fracture intersecting the canister deposition hole is modelled. The model describes the expansion of the bentonite within the fracture. It accounts for the repulsive electrostatic double-layer forces, the attractive van der Waals forces and friction forces between the particles and the water. The model also takes into account the diffusion of the colloid particles in the smectite sol. Diffusion of a counterion, sodium, is accounted for as this strongly influences the double layer force and the viscosity of the gel/sol. The gel/sol is considered to be a fluid with a varying viscosity that is strongly dependent on the bentonite volume fraction in the gel and the sodium concentration in the water. Two different geometries were modelled; a rectangular and a cylindrical. The rectangular geometry was used to gain experience with the processes and mechanisms and how they interact since the cylindrical geometry was somewhat less stable numerically and more time consuming. In the rectangular geometry a fracture 1 metre long in the flow direction was modelled. In both geometries the fracture size was selected sufficiently large to ensure that the water velocity, near the distant border was nearly the same as the approaching water velocity and that the smectite concentration there was vanishingly small. It was found that the velocity of the fluid drops considerably where the bentonite volume fraction is larger than 1-2%. This is due to the strong increase in viscosity with increasing bentonite volume fraction. The loss of smectite by the slowly flowing fluid was found to be proportional to the square root of the seeping water velocity for the rectangular geometry. For the cylindrical geometry, the dependence is somewhat lower (exponent about 0.4) since the length of the gel/water interface decreases with increasing water flow rate. The penetration depth of the gel/water interface decreases with increasing water flow rate. For water velocity of the

Gel-eletroforese no diagnóstico da varíola Gel-electrophoresis in the smallpox diagnosis

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Julio A. Mesquita

1972-01-01

Full Text Available O emprego de gel-eletroforese no diagnóstico da varíola, demonstrou ser ao menos trinta vezes (30X mais sensível que o teste de agar-gel, nas condições descritas (tabela I. Doze (12 espécimes, cujos testes convencionais de inoculação em ovos embrionados e de agar-gel resultaram positivos, foram testados em suas diluições originais congeladas por mais de um ano, sendo seis deles revelados por gel-eletroforese enquanto nenhum o foi por agar-gel (tabela II. Trinta e três (33 amostras isoladas no laboratório, foram testadas com material colhido de membrana cório-alantóica da primeira inoculação para o diagnóstico, conservado em glicerina 50%, resultando 15 positivas em gel-eletroforese e apenas 3 em agar-gel (tabela II. Os últimos 60 espécimes recebidos para diagnóstico, através a Campanha de Erradicação da Varíola, também resultaram negativos em gel-eletroforese, que não mostrou falsos-positivos nas condições descritas.The test of gel-electrophoresis applied to the pox virus group showed to be at least thirth times (30X more sensitive than agar-gel test on the described conditions (Table I. Twelve specimens, which were positives form Smallpox in the conventional tests of egg inoculation and agar-gel difusion test, have been screened in their original dilutions frozen for more than 1 year and six of them were still detectable by gel-eletrophoresis, while by agar-gel test any of them was positive (Table II. Thirty three Smallpox isolates have been tested with material from first egg inoculation (chorioallantoic membranes which have been stored in glycerin 50%, at - 15ºC. Fifteen of them were still positive by gel-electrophoresis and only 3 by agar-gel (Table II. The last 60 specimens received for diagnosis from Smallpox Erradication Campaign (CEV, were negatives by both tests. The gel-electrophoresis, did not show false-positives on described conditions.

Efficacy and Tolerability of Fitostimoline in Two Different Forms (Soaked Gauzes and Cream and Citrizan Gel in the Topical Treatment of Second-Degree Superficial Cutaneous Burns

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Patrizia Martini

2011-01-01

Full Text Available A total of 227 patients (mean age 41.3 years, 52% females with at least one second-degree superficial cutaneous burn of thermal origin of a smallest transverse diameter ≥20 mm and a largest transverse diameter ≤90 mm were randomised to receive the topical application of aqueous extract of Triticum vulgare (Fitostimoline in two different forms (soaked gauzes and cream or catalase of horse origin in form of gel (Citrizan Gel, given up to healing or to a maximum of 20 days. The rate of lesion healing at end of study was significantly higher in patients treated with Fitostimoline (gauzes 97.3%, cream 91.5% than in those receiving catalase (84.5%. The pooled Fitostimoline groups were also significantly more effective than catalase gel in reducing total symptoms score, pain at medication, pain at rest, and burning at end of study. Both formulations of Fitostimoline and catalase gel were well tolerated in terms of adverse effects in the site of application.

Unbiased minimum variance estimator of a matrix exponential function. Application to Boltzmann/Bateman coupled equations solving

International Nuclear Information System (INIS)

Dumonteil, E.; Diop, C. M.

2009-01-01

This paper derives an unbiased minimum variance estimator (UMVE) of a matrix exponential function of a normal wean. The result is then used to propose a reference scheme to solve Boltzmann/Bateman coupled equations, thanks to Monte Carlo transport codes. The last section will present numerical results on a simple example. (authors)

Instrumental texture profile analysis of gelatin gel extracted from grouper skin and commercial (bovine and porcine) gelatin gels.

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Rahman, Mohammad Shafiur; Al-Mahrouqi, Abdullah Issa

2009-01-01

Mechanical compression was used to study the gelling characteristics of gelatin gels. Texture profile analysis (TPA) showed that the hardness of fish and mammalian gelatin increased significantly as the concentrations of gels increased. TPA attributes of 10% fish skin gel showed significant differences from those obtained from 20% and 30% gels. In bovine and porcine cases, such generic trends were not observed. Mechanical characteristics of 10% gels of gelatin from fish skin, determined from one cycle compression, were significantly lower than other sources of gelatin gels, while bovine and porcine gels did not show any significant differences. In the case of TPA, hardness of bovine gelatin gel was highest at 41 N for 10% gel, followed by porcine (30 N) then fish skin (5 N) gelatin gels. The gels prepared from different sources did not show any generic trends when all other mechanical attributes were considered.

A new approach for cloning hLIF cDNA from genomic DNA isolated from the oral mucous membrane.

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Cui, Y H; Zhu, G Q; Chen, Q J; Wang, Y F; Yang, M M; Song, Y X; Wang, J G; Cao, B Y

2011-11-25

Complementary DNA (cDNA) is valuable for investigating protein structure and function in the study of life science, but it is difficult to obtain by traditional reverse transcription. We employed a novel strategy to clone human leukemia inhibitory factor (hLIF) gene cDNA from genomic DNA, which was directly isolated from the mucous membrane of mouth. The hLIF sequence, which is 609 bp long and is composed of three exons, can be acquired within a few hours by amplifying each exon and splicing all of them using overlap-PCR. This new approach developed is simple, time- and cost-effective, without RNA preparation or cDNA synthesis, and is not limited to the specific tissues for a particular gene and the expression level of the gene.

APPLICATION OF CDNA MICROARRAY TO THE STUDY OF ARSENIC TOXICOLOGY AND CARCINOGENESIS

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Arsenic (As) is a common environmental toxicant and known human carcinogen. Epidemiological studies link As exposure to various disorders and cancers. However, the molecular mechanisms for As toxicity and carcinogenicity are not completely known. The cDNA microarray, a high-th...

Cloning and expression of a human kidney cDNA for an α2-adrenergic receptor subtype

International Nuclear Information System (INIS)

Regan, J.W.; Kobilka, T.S.; Yang-Feng, T.L.; Caron, M.G.; Lefkowitz, R.J.; Kobilka, B.K.

1988-01-01

An α 2 -adrenergic receptor subtype has been cloned from a human kidney cDNA library using the gene for the human platelet α 2 -adrenergic receptor as a probe. The deduced amino acid sequence resembles the human platelet α 2 -adrenergic receptor and is consistent with the structure of other members of he family of guanine nucleotide-binding protein-coupled receptors. The cDNA was expressed in a mammalian cell line (COS-7), and the α 2 -adrenergic ligand [ 3 H]rauwolscine was bound. Competition curve analysis with a variety of adrenergic ligands suggests that this cDNA clone represents the α 2 B-adrenergic receptor. The gene for this receptor is on human chromosome 4, whereas the gene for the human platelet α 2 -adrenergic receptor (α 2 A) lies on chromosome 10. This ability to express the receptor in mammalian cells, free of other adrenergic receptor subtypes, should help in developing more selective α-adrenergic ligands

Compatibility between weak gel and microorganisms in weak gel-assisted microbial enhanced oil recovery.

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Qi, Yi-Bin; Zheng, Cheng-Gang; Lv, Cheng-Yuan; Lun, Zeng-Min; Ma, Tao

2018-03-20

To investigate weak gel-assisted microbial flooding in Block Wang Long Zhuang in the Jiangsu Oilfield, the compatibility of weak gel and microbe was evaluated using laboratory experiments. Bacillus sp. W5 was isolated from the formation water in Block Wang Long Zhuang. The rate of oil degradation reached 178 mg/day, and the rate of viscosity reduction reached 75.3%. Strain W5 could produce lipopeptide with a yield of 1254 mg/L. Emulsified crude oil was dispersed in the microbial degradation system, and the average diameter of the emulsified oil particles was 18.54 μm. Bacillus sp. W5 did not affect the rheological properties of the weak gel, and the presence of the weak gel did not significantly affect bacterial reproduction (as indicated by an unchanged microbial biomass), emulsification (surface tension is 35.56 mN/m and average oil particles size is 21.38 μm), oil degradation (162 mg/day) and oil viscosity reduction (72.7%). Core-flooding experiments indicated oil recovery of 23.6% when both weak gel and Bacillus sp. W5 were injected into the system, 14.76% when only the weak gel was injected, and 9.78% with strain W5 was injected without the weak gel. The results demonstrate good compatibility between strains W5 and the weak gel and highlight the application potential of weak gel-assisted microbial flooding. Copyright © 2018 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

Molecular cloning and sequence analysis of growth hormone cDNA of Neotropical freshwater fish Pacu (Piaractus mesopotamicus

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Janeth Silva Pinheiro

2008-01-01

Full Text Available RT-PCR was used for amplifying Piaractus mesopotamicus growth hormone (GH cDNA obtained from mRNA extracted from pituitary cells. The amplified fragment was cloned and the complete cDNA sequence was determined. The cloned cDNA encompassed a sequence of 543 nucleotides that encoded a polypeptide of 178 amino acids corresponding to mature P. mesopotamicus GH. Comparison with other GH sequences showed a gap of 10 amino acids localized in the N terminus of the putative polypeptide of P. mesopotamicus. This same gap was also observed in other members of the family. Neighbor-joining tree analysis with GH sequences from fishes belonging to different taxonomic groups placed the P. mesopotamicus GH within the Otophysi group. To our knowledge, this is the first GH sequence of a Neotropical characiform fish deposited in GenBank.

Production of continuous mullite fiber via sol-gel processing

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Tucker, Dennis S.; Sparks, J. Scott; Esker, David C.

1990-01-01

The development of a continuous ceramic fiber which could be used in rocket engine and rocket boosters applications was investigated at the Marshall Space Flight Center. Methods of ceramic fiber production such as melt spinning, chemical vapor deposition, and precursor polymeric fiber decomposition are discussed and compared with sol-gel processing. The production of ceramics via the sol-gel method consists of two steps, hydrolysis and polycondensation, to form the preceramic, followed by consolidation into the glass or ceramic structure. The advantages of the sol-gel method include better homogeneity and purity, lower preparation temperature, and the ability to form unique compositions. The disadvantages are the high cost of raw materials, large shrinkage during drying and firing which can lead to cracks, and long processing times. Preparation procedures for aluminosilicate sol-gel and for continuous mullite fibers are described.