International Nuclear Information System (INIS)

Genes encoding mutants of the thymine photodimer repair enzyme from bacteriophage T4 (T4 endonuclease V) having an amino acid substitution (T127M, W128A, W128S, Y129A, K130L, Y131A, Y132A) were constructed by use of a previously obtained synthetic gene and expressed in Escherichia coli under the control of the E. coli tryptophan promoter. An in vitro assay of partially fractionated mutant proteins for glycosylase activity was performed with chemically synthesized substrates containing a thymine photodimer. T127M and K130L showed almost the same activity as the wild-type protein. Although W128S, Y131A, and Y132A were slightly active, W128A and Y129A lost activity. The results indicated that the aromatic amino acids around position 130 may be important for the glycosylase activity. Mutant T127M was purified, and the Km value was found to be of the same order as that of the wild type (10(-8) M). In vivo activities for all mutants were characterized with UV-sensitive ...

International Nuclear Information System (INIS)

January, 1992, to October, 1995, sixty-four patients with advanced head and neck cancer underwent head and neck reconstructive surgery using myocutaneous or revascularized flaps; in the same period, all patients were consecutively examined with CT and MRI. Myocutaneous flaps were used in 26 patients: 12 flaps were tubular and 14 linear. Revascularized flaps were used in 38 patients: to repair a large defect in 26 patients (14 latissimus dorsi flaps and 12 temporal muscle flaps) and to repair an oral damage in 12 patients (5 revascularized radial and 7 jejunal flaps). CT and MR images of myocutaneous flaps showed the flaps as fatty areas, repairing large surgical defects, hypodense at CT and hyperintense at MRI, with no post-contrast enhancement. The appearance of revascularized ...

UK PubMed Central (United Kingdom)

Human apurinic/apyrimidinic endonuclease (hApe1) encodes two important functional activities: an essential base excision repair (BER) activity and a redox activity that regulates expression...Full Text Available

International Nuclear Information System (INIS)

... [2] Hee Taek Chae, et al., "On the Natural Convection Cooling in HANARO;

International Nuclear Information System (INIS)

The addition of paraquat (methyl viologen) to a growing culture of Escherichia coli K-12 led within 1 hr to a 10- to 20-fold increase in the level of endonuclease IV, a DNase for apurinic/apyrimidinic sites. The induction was blocked by chloramphenicol. Increases of 3-fold or more were also seen with plumbagin, menadione, and phenazine methosulfate. H_2O_2 produced no more than a 2-fold increase in endonuclease IV activity. The following agents had no significant effect: streptonigrin, nitrofurantoin, tert-butyl hydroperoxide, #gamma# rays, 260-nm UV radiation, methyl methanesulfonate, mitomycin C, and ascorbate. Paraquat, plumbagin, menadione, and phenazine methosulfate are known to generate superoxide radical anions via redox cycling in vivo. A mutant lacking superoxide dismutase was unusually sensitive to induction by paraquat. In addition, endonuclease IV could be induced by merely growing the mutant in pure O_2. The ...

Science.gov (United States)

Recent work on the aerodynamics of flapping flight reveals fundamental differences in the mechanisms of aerodynamic force generation between fixed and flapping wings. When fixed wings translate at high angles of attack, they periodically generate and shed leading and trailing edge vortices as reflected in their fluctuating aerodynamic force traces and associated flow visualization. In contrast, wings flapping at high angles of attack generate stable leading edge vorticity, which persists throughout the duration of the stroke and enhances mean aerodynamic forces. Here, we show that aerodynamic forces can be controlled by altering the trailing edge flexibility of a flapping wing. We used a dynamically scaled mechanical model of flapping flight (Re approximately 2000) to measure the aerodynamic forces on flapping wings of variable flexural stiffness (EI). For low ...

British Library Electronic Table of Contents (United Kingdom)

Abstract TLR9 recognizes unmethylated CpG-containing DNA commonly found in bacteria. Synthetic oligonucleotides containing CpG-motifs (CpG ODNs) recapitulate the activation of TLR9 by microbial DNA, whereas inversion of the CG dinucleotide within the CpG motif to GC (GpC ODNs) renders such ODNs inactive. This difference cannot be attributed to binding of ODNs to the full-length TLR9 ectodomain, as both CpG and GpC ODNs bind comparably. Activation of murine TLR9 requires cleavage into an active C-terminal fragment, which binds CpG robustly. We therefore compared the ability of CpG and GpC ODNs to bind to full-length and C-terminal TLR9, and their impact on the cleavage of TLR9. We found that CpG binds better to C-terminal TLR9 when compared with GpC, despite comparably low binding of both O...

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Synaptobrevin is a synaptic vesicle protein that has an essential role in exocytosis and forms the SNARE complex with syntaxin and SNAP-25. We have analyzed the structure of isolated synaptobrevin and its binary interaction with syntaxin using NMR spectroscopy. Our results demonstrate that isolated synaptobrevin is largely unfolded in solution. The entire SNARE motif of synaptobrevin is capable of interacting with the isolated C-terminal SNARE motif of syntaxin but only a few residues bind to the full-length cytoplasmic region of syntaxin. This result suggests an interaction between the N- and C-terminal regions of syntaxin that competes with core complex assembly.

UK PubMed Central (United Kingdom)

The integration sites for viral DNA in cells infected with Moloney murine leukemia virus (M-MuLV) were studied by restriction endonuclease cleavage of cellular DNA followed by electrophoresis in agarose...Full Text Available

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Glioblastoma multiforme is the most aggressive form of human brain tumor, which has no effective cure. Previously, we have demonstrated that overexpression of the C-terminal fragment of the human telomerase reverse transcriptase (hTERTC27) inhibits the growth and tumorigenicity of human cervical cancer HeLa cells. In this study, the therapeutic effect and molecular mechanisms of hTERTC27-mediated cancer gene therapy were further explored in vivo in established human glioblastoma xenografts in nude mice. We showed that intratumoral injection of adeno-associated virus carrying hTERTC27 (rAAV-hTERTC27) is highly effective in reducing the growth of the subcutaneously transplanted glioblastoma tumors. Histological analyses showed that rAAV-hTERTC27 treatment leads to profound necrosis, apoptosi...

UK PubMed Central (United Kingdom)

A duplication of the polypurine tract (PPT) at the center of the human immunodeficiency virus type 1 (HIV-1) genome (the cPPT) has been shown to prime a separate plus-strand initiation and to result...Full Text Available

UK PubMed Central (United Kingdom)

RIN proteins serve as guanine nucleotide exchange factors for Rab5a. They are characterized by the presence of a RIN homology domain and a C-terminal Vps9 domain. Currently three family members have...Full Text Available

UK PubMed Central (United Kingdom)

Hepatitis C virus (HCV) nonstructural protein 5B (NS5B), the viral RNA-dependent RNA polymerase (RdRp), is a tail-anchored protein with a highly conserved C-terminal transmembrane domain (TMD) that...Full Text Available

Science.gov (United States)

Excision repair in uv irradiated human fibroblasts has been examined in portions of DNA replicating after irradiation versus those remaining unreplicated. Two approaches, one using a uv-endonuclease to estimate pyrimidine dimers remaining in DNA, the other using density labeling to measure excision resynthesis, indicate that the extent of repair is the same for both replicated and unreplicated DNA.

Science.gov (United States)

In a 47-year-old male patient a bronchopleural fistula was apparent 22 days after extended right-sided diaphragma-pericardio-pleuro-pneumonectomy for pleuramesothelioma. The thoracic cavity was infected. Rethoracotomy was performed and the fistula was closed using an omental pedicle flap. The bronchial stump became tight and the cavity fluid became sterile. No abdominal complications were seen. The patient died 8 months later from malignant pericardial infiltration. The ability of greater omentum to revascularize ischemic tissue, to absorb fluid and to resist local infection is proved and used in several subspecialities of surgery. Nevertheless the use of the greater omentum in the management of bronchopleural fistula has been rarely published. The reported case shows that the closure of a large bronchopleural fistula is possible by using the attributes of the omental tissue. PMID:2264043

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The effect of high lift airfoils on the cost effectiveness of HAWT and VAWT (horizontal and vertical axis wind turbine) machines is studied. The scope involved first studying modern two dimensional airfoils, and developing a generalized formulation for their performance in terms of lift, drag, and thickness at appropriate Reynolds numbers. Single element, multi-element, symmetrical, extra thick airfoils and jet flap airfoils were analyzed. The jet flap airfoils were considered to be unacceptable because of excessive power requirements. Then the effect of using the above airfoils on the rotors of a variety of wind turbines was made. Qualitative representation of the type of airfoils studied is given.

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We have constructed a 27-kDa hTERT C-terminal polypeptide (hTERTC27) devoid of domains required for telomerase activity and demonstrated that it is capable of nuclear translocation/telomere-end targeting. Here we showed that expression of a low level of hTERTC27 renders hTERT positive HeLa cells sensitive to H(2)O(2)-induced oxidative stress and subsequent cell senescence. The senescence-associated gene, the cyclin/cdk inhibitor p21(Waf1), was up-regulated. This occurs without changing the expression of endogenous hTERT, causing significant telomere shortening or inhibiting telomerase activity. Results from this study suggest for the first time that in addition to telomerase activity, the C-terminus of hTERT also plays a role in hTERT-mediated cellular resistance to oxidative stress. PMID:12565825

British Library Electronic Table of Contents (United Kingdom)

SR proteins are essential splicing factors whose function is controlled by multi-site phosphorylation of a C-terminal domain rich in arginine-serine repeats (RS domain). The protein kinase SRPK1 has been shown to polyphosphorylate the N-terminal portion of the RS domain (RS1) of the SR protein ASF/SF2, a modification that promotes nuclear entry of this splicing factor and engagement in splicing function. Later, dephosphorylation is required for maturation of the spliceosome and other RNA processing steps. While phosphates are attached to RS1 in a sequential manner by SRPK1, little is known about how they are removed. To investigate factors that control dephosphorylation, we monitored region-specific mapping of phosphorylation sites in ASF/SF2 as a function of the protein phosphatase PP1. W...

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Abstract Objectives: The purpose of this study was to evaluate the osseointegration of the dental implants placed into the mandible augmented with different techniques in pigs. Material and methods: Four adult domestic pigs were used. Horizontal augmentation of the mandible was performed in animals by using vascularized femur flap (VFF), non-vascularized femur graft (NVFG) and monocortical mandibular block graft (MG). After 5 months of healing 10 dental implants were placed into each augmented site. The pigs were sacrificed after 3 months of healing. Undecalcified sections were prepared for histomorphometric analysis. Results: Mean bone-implant contact (BIC) values for implants placed into MG, NVFG and VFF were 57.38 11.97%, 76.5 7.88%, 76.53 8.15%, respectively. The BIC values of NVFG and...

International Nuclear Information System (INIS)

Nop25 is a putative RNA binding nucleolar protein associated with rRNA transcription. The present study was undertaken to determine the mechanism of Nop25 localization in the nucleolus. Deletion experiments of Nop25 amino acid sequence showed Nop25 to contain a nuclear targeting sequence in the N-terminal and a nucleolar targeting sequence in the C-terminal. By expressing derivative peptides from the C-terminal as GFP-fusion proteins in the cells, a lysine and arginine residue-enriched peptide (KRKHPRRAQDSTKKPPSATRTSKTQRRRR) allowed a GFP-fusion protein to be transported and fully retained in the nucleolus. When the peptide was fused with cMyc epitope and expressed in the cells, a cMyc epitope was then detected in the nucleolus. Nop25 did not localize in the nucleolus by deletion of the peptide from Nop25. Furthermore, deletion of a subdomain (KRKHPRRAQ) in the peptide or amino acid substitution of lysine and arginine residues in the subdomain ...

Science.gov (United States)

The infection of cultured human cells with baboon endogenous virus (BEV) frequently leads to an association of viral DNA with a specific genetic locus (termed BEVI, for baboon endogenous virus infection) on chromosome 6. Restriction endonuclease digestion of DNA from BEV-infected human cells and their derived somatic cell clones frequently revealed a common cellular DNA sequence in the proximity of one of the junctions between cellular DNA and the integrated virus. We propose that a short cellular DNA sequence, repeated on chromosome 6 and separated by unique DNA sequences, presents a high-affinity target for the integration of BEV in human cells. PMID:6401843

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The aim of this study was to evaluate the feasibility of splitting each head of the gastrocnemius muscle into two segments. This prospective study was conducted on 35 fresh cadavers with injection of radio-opaque contrast material in the popliteal artery. Seventy medial gastrocnemius and 70 lateral gastrocnemius muscle specimens were obtained. Gastrocnemius intramuscular arterial anatomy was analysed by using the digital X-ray technique. The most common vascular pattern found in this study was both bellies supplied by individual sural arteries, which shows distal bifurcation. The medial sural artery always showed bifurcation in the medial gastrocnemius muscle in both lower limbs, whereas this bifurcation of the lateral sural artery was present in only 87% cases and 13% of cases showed a si...

British Library Electronic Table of Contents (United Kingdom)

Purpose The aim of this study was to evaluate the role of host periosteum and recipient bed perforation on the healing of an onlay graft to the mandible based on histologic and immunohistochemical analysis. Materials and methods Each of the 12 adult sheep used in the study received four iliac corticocancellous onlay bone grafts on the lateral surface of the mandible. In experiment 1, the block graft was placed in direct contact with the recipient bed and fixed with micro-screws, and in experiment 2, the recipient cortical bed was perforated before graft placement. The host periosteum around the graft was excised before flap replacement in experiment 3, and in experiment 4, a sheet of silicone membrane was placed between the graft and the recipient bed. The animals were euthanised at 4, 8, ...

British Library Electronic Table of Contents (United Kingdom)

Sinus lifting and reconstruction of localized alveolar defects are often required after closure of a large oroantral fistula (OAF) to allow for subsequent implant installation. This study describes a combined surgical technique that involves sinus lifting, bony closure, and reconstruction of the alveolar defect at the site of an OAF. The sinus membrane was reconstructed as a continuous layer by combining the residual sinus membrane with a rotated part of oral mucosa around the OAF. Autogenous bone from the chin and/or ramus was grafted into the prepared sinus space and alveolar defect, and the graft was covered by a buccal advancement flap. This technique was used to treat 8 patients who had large OAFs in the posterior maxillary region. The treatment was successful in all cases, and the te...

International Nuclear Information System (INIS)

A radioimmunoassay is described for the molluscan neuropeptide, Phe-Met-Arg-Phe-NH"2(FMRFamide). The antibody used is C-terminal-specific and shows slight but significant (1-2%) cross-reactivity with chicken pancreatic polypeptide (APP). The assay has been used to identify in rat brain extracts a pair of molecules that may represent mammalian counterparts of FMRFamide. Their concentrations were highest in spinal cord and hypothalamus (>10 pmol.g"-_1) and lowest in cerebellum and striatum (<3.5 pmol.g"-_1). The two immunoreactive peptides were separated on CM ion-exchange chromatography where they appeared to be less basic than FMRFamide. On Sephadex G50 gel filtration one eluted in a similar position to FMRFamide and the other slightly earlier suggesting it may be of higher molecular weight. The rat immunoreactive components do not correspond to previously described neuropeptides or hormones, and may be members of a new group of mammalian neuropeptides with ...

International Nuclear Information System (INIS)

The production of molecularly imprinted polymers (MIPs) for the recognition of C-terminal cholecystokinin pentapeptide (CCK-5) in the presence of metal ion is reported. The MIPs were produced under the same molar ratio of template to monomers (acrylamide, N,N'-methylene bisacrylamide) in the presence or absence of nitrilotriacetic acid-nickel (Ni-NTA) complex. Scanning electron microscopy images of MIPs were obtained in an attempt to correlate the adsorption characteristics with polymer's morphology. Subsequently Ni2+ was removed and substituted by other divalent ions such as Mg2+, Fe2+, Zn2+, Co2+ and Cu2+. It was found that polymers containing the metal ion complex with the order Fe-NTA, Ni-NTA and Cu-NTA presented lower dissociation constant values than the rest thus exhibiting stronger guest binding activity. The percentage of theoretical maximum binding sites Bmax was almost the same for these ions, indicating that the ion-template coordination is responsible ...

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The overall objective of this project has been to develop immunochemical methods to quantitate unique DNA base damages in order to facilitate studies on radiation-induced damage production and repair. Specifically, we have been using antibodies raised to damaged bases to quantitate unique lesions in model systems in order to evaluate their potential biological consequences. Our approach has been to synthesize modified nucleotides or nucleosides, conjugate them to protein carriers, and use the conjugates as immunogens in rabbits or to prepare monoclonal antibodies. We have been studying damages that are stable radiolysis products found in X-irradiated DNA and thus of potential biological consequence. Our aim is to build an in vitro and in vivo data base on the interactions between model DNA lesions and such cellular enzymes as DNA polymerases and repair endonucleases. Initial studies have focused on pyrimidine ring saturation products (thymine glycol.and ...

Science.gov (United States)

The unsteady aerodynamic forces of a model fruit fly wing in flapping motion were investigated by numerically solving the Navier-Stokes equations. The flapping motion consisted of translation and rotation [the translation velocity (u(t)) varied according to the simple harmonic function (SHF), and the rotation was confined to a short period around stroke reversal]. First, it was shown that for a wing of given geometry with u(t) varying as the SHF, the aerodynamic force coefficients depended only on five non-dimensional parameters, i.e. Reynolds number (Re), stroke amplitude (Phi), mid-stroke angle of attack (alpha(m)), non-dimensional duration of wing rotation (Delta tau(r)) and rotation timing [the mean translation velocity at radius of the second moment of wing area (U), the mean chord length (c) and c/U were used as reference velocity, length and time, respectively]. Next, the force coefficients were investigated for a case in which typical ...

Science.gov (United States)

A patient with a totally edentulous maxilla and a seVere Class III intermaxillary relationship in the anterior region was treated by implants. In the mandible, there were 10 teeth between the second premolars. The inclination and width of the maxillary anterior residual bone were measured on cephalometric X-ray film obtained before treatment. The results of cephalometric analysis did not support clockwise rotation of the mandible or lingual angling of the maxillary anterior teeth by use of prosthesis to improve the Class III relationship. Ten implants were simultaneously placed in the maxilla. Then, a maxillary temporary full bridge was seated after reduction of the crown lengths of the mandibular anterior teeth. An apically positioned flap operation was performed to eliminate periodontal pockets and to obtain clinically suitable crown lengths of the mandibular anterior teeth. A noncemented, screw-retained maxillary full bridge and a conventional mandibular full ...

International Nuclear Information System (INIS)

As part of the Probabilistic Safety Analysis of the Multi Purpose Reactor, MPR, the list of Postulated Initiating Events was analyzed and one of these PIEs corresponds to the Loss of Coolant Flow. It is well known that during the operation life of a research reactor a LOFA could eventually occur and, once this event takes place, in time detection and automatic actions, thanks to the engineering safety features of the system, will mitigate the incident evolution. The postulated event corresponds to a loss of flow due to a total loss of power supply. The goal of the present work is to provide a general description and the engineering safety features of the MPR, as well as describe the sequence of scenarios during a LOFA. Temporal evolution of main parameters is presented, also. During Stage A of the Commissioning Program measurements of the core cooling system pump coast-down were performed in order to validate previous simulation results, as well as, flap valves ...

Science.gov (United States)

A computational fluid dynamics (CFD) is coupled with a computational structural dynamics (CSD) to simulate the unsteady rotor flow with aeroelasticity effects. An unstructured upwind Navier-Stokes solver was developed for this simulation, with 2nd order time-accurate dual-time stepping method for temporal discretization and low Mach number preconditioning method. For turbulent flows, both the Spalart-Allmaras and Menter's SST model are available. Mesh deformation is achieved through a fast dynamic grid method called Delaunay graph map method for unsteady flow simulation. The rotor blades are modeled as Hodges & Dowell's nonlinear beams coupled flap-lag-torsion. The rotorcraft computational structural dynamics code employs the 15-dof beam finite element formulation for modeling. The structure code was validated by comparing the natural frequencies of a rotor model with UMARC. The flow and structure codes are coupled tightly with information exchange several ...

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Trihydroxynaphthalene reductase catalyzes two intermediate steps in the fungal melanin biosynthetic pathway. The enzyme, a typical short-chain dehydrogenase, is the biochemical target of three commercial fungicides. The fungicides bind preferentially to the NADPH form of the enzyme. Three X-ray structures of the Magnaporthe grisea enzyme complexed with NADPH and two commercial and one experimental fungicide were determined at 1.7 {angstrom} (pyroquilon), 2.0 {angstrom} (2,3-dihydro-4-nitro-1H-inden-1-one, 1), and 2.1 {angstrom} (phthalide) resolutions. The chemically distinct inhibitors occupy similar space within the enzyme's active site. The three inhibitors share hydrogen bonds with the side chain hydroxyls of Ser-164 and Tyr-178 via a carbonyl oxygen (pyroquilon and 1) or via a carbonyl oxygen and a ring oxygen (phthalide). Active site residues occupy similar positions among the three structures. A buried water molecule that is hydrogen bonded to the NZ nitrogen of Lys-182 ...

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NMR spectroscopy combined with paramagnetic relaxation agents was used to study the positioning of the 40-residue Alzheimer Amyloid {beta}-peptide A{beta}(1-40) in SDS micelles. 5-Doxyl stearic acid incorporated into the micelle or Mn{sup 2+} ions in the aqueous solvent were used to determine the position of the peptide relative to the micelle geometry. In SDS solvent, the two {alpha}-helices induced in A{beta}(1-40), comprising residues 15-24, and 29-35, respectively, are surrounded by flexible unstructured regions. NMR signals from these unstructured regions are strongly attenuated in the presence of Mn{sup 2+} showing that these regions are positioned mostly outside the micelle. The central helix (residues 15-24) is significantly affected by 5-doxyl stearic acid however somewhat less for residues 16, 20, 22 and 23. This {alpha}-helix therefore resides in the SDS headgroup region with the face with residues 16, 20, 22 and 23 directed away from the hydrophobic interior of the micelle. ...

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Amino acids other than those that serve as ligands have been found to influence the chemical properties of transferrin iron. The catalytic ability of pyrophosphate to mediate transferrin iron release to a terminal acceptor is largely quenched by modification non-liganded histine groups on the protein. The first order rate constants of iron release for several partially histidine modified protein samples were measured. A statistical method was employed to establish that one non-liganded histidine per metal binding domain was responsible for the reduction in rate constant. These results imply that the iron mediated chelator, pyrophosphate, binds directly to a histidine residue on the protein during the iron release process. EPR spectroscopic results are consistent with this interpretation. Kinetic and amino acid sequence studies of ovotransferrin and lactoferrin, in addition to human serum transferrin, have allowed the tentative assignment of His-207 in the N-terminal domain and His-535 ...

International Nuclear Information System (INIS)

The helicase action of the Escherichia coli UvrAB complex on a covalently closed circular DNA template was monitored using bacterial DNA topoisomerase I, which specifically removes negative supercoils. In the presence of E. coli DNA topoisomerase I and ATP, the UvrAB complex gradually introduced positive supercoils into the input relaxed plasmid DNA template. Positive supercoils were not produced when E. coli DNA topoisomerase I was replaced by eukaryotic DNA topoisomerase I or when both E. coli and eukaryotic DNA topoisomerases I were added simultaneously. These results suggest that like other DNA helix-tracking processes, the ATP-dependent action of the UvrAM complex on duplex DNA simultaneously generates both positive and negative supercoils, which are not constrained by protein binding but are torsionally strained. The supercoiling activity of UvrAB on UV-damaged DNA was also studied using UV-damaged plasmid DNA and a mutant UvrA protein that lacks the 40 ...

Science.gov (United States)

High molecular weight DNA from pleroceroid larvae of the tapeworm Spirometra mansonoides was purified from isolated nuclei by conventional techniques. The DNA so isolated has a melting temperature (Tm) of 87 degrees C and a guanine plus cytosine (G/C) content of 44%. 5-Methyl cytosine could not be detected in plerocercoid DNA by HPLC analysis of DNA hydrolysates, by radiolabeling 5'-termini of MspI digests with polynucleotide kinase, or by comparing restriction patterns generated by MspI and HpaII. Renaturation kinetics demonstrated that the genome of S. mansonoides contains repetitive as well as single copy sequences and has a genome size estimated at approx. 1.6 X 10(9) bp. Hybridization was carried out between plerocercoid DNA and cDNAs for human beta-actin, alpha-tubulin and growth hormone (hGH). Rationale for this analysis was based on known homologies among actin and tubulin genes in numerous species and on apparent similarities between hGH and a plerocercoid growth factor that ...

International Nuclear Information System (INIS)

We have compared the kinetics for repair of UV-induced cyclobutane pyrimidine dimers in the DNA of keratinocytes in human skin and in cell culture. A small area of the buttocks of volunteers was exposed to UVB-irradiation and biopsies were taken at various time intervals. Epidermal keratinocytes in culture from the same subjects were exposed to UVC with doses chosen to elicit comparable yields of dimers in cellular DNA. The initial density of pyrimidine dimers and the kinetics of their removal were assessed utilizing the dimer-specific T4 endonuclease V and sedimentation of the unlabeled DNA through alkaline sucrose gradients. The position of DNA in the gradients was determined using a monoclonal antibody against random sequences of single-stranded DNA in a sensitive immunoassay. The initial dimer frequency was 3.9-6.7 per 10(8) daltons DNA. About 40% of the dimers were removed within 1 h, 70% in 6 h, and 90% in 24 h for both in vivo and in vitro samples. The early ...

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IntroductionBreast cancer is the most diagnosed and second leading cause of cancer deaths in the U.S. female population. An estimated 5 to 10 percent of all breast cancers are inherited, caused by mutations in the breast cancer susceptibility genes (BRCA1/2). As many as 90% of all mutations are nonsense mutations, causing a truncated polypeptide product. A popular and low cost method of mutation detection has been the protein truncation test (PTT), where target regions of BRCA1/2 are PCR amplified, transcribed/translated in a cell-free protein synthesis system and analyzed for truncated polypeptides by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and autoradiography. We previously reported a novel High Throughput Solid-Phase PTT (HTS-PTT) based on an enzyme-linked immunosorbent assay (ELISA) format that eliminates the need for radioactivity, SDS-PAGE and subjective interpretation of the results. Here, we report the next generation HTS-PTT using ...

International Nuclear Information System (INIS)

The detection of foot-and-mouth disease (FMD) virus from various kinds of field samples (tissue extract and cell culture isolate) was studied using the polymerase chain reaction (PCR) technique. The gene selected for diagnosis was the polymerase gene and an amplification target product of 454 bp in length was produced using AP5/AP6 primer sets. The PCR product was further examined by NcoI endonuclease digestion. The presence of the internal restriction site was confirmed by demonstration of two small fragments of 330 bp and 124 bp in length. Forty-nine samples that gave positive and negative results by ELISA typing and were positive by the PCR test were tested by NcoI digestion to confirm the results. About 10% of PCR products could not be confirmed by the method. Furthermore the FMD RNA polymerase gene could be detected by the PCR method in samples negative in both ELISA typing and the virus isolation test. A total of 23 samples were examined and compared after ...

International Nuclear Information System (INIS)

Purpose/Objective: The role of programmed cell death (apoptosis) as a cellular response to cancer therapy such as radiation or chemotherapy is the subject of much study, and manipulation of the apoptotic response in tumor cells may be valuable in the treatment of a variety of cancers. Both p53 dependent and independent apoptotic pathways have been identified; p53 is mutated in at least 50 % of human cancers and a majority of radiation resistant tumors contain p53 mutations. This study is designed to examine the induction of programmed cell death in a human colon carcinoma cell line that possesses two mutated p53 alleles. Ionizing radiation alone, or in combination with the chemotherapeutic drug 5-fluorouracil (5-FU), were used to elicit the apoptotic response. This study will focus on whether these treatments can induce a significant apoptotic response in cells that have mutated p53 alleles. Materials and Methods: HT-29 cells were assessed for clonogenic survival after being plated at ...