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Molecular Characterization of a 70 kDa Heat Shock Protein (HSP70) Gene in Entamoeba dispar
S Rezaie,A Birami,M Rezaian
Iranian Journal of Public Health , 2006,
Abstract: Amebiasis caused by Entamoeba histolytica is still mentioned as one of the major health problems in tropical and subtropical areas. E. histolytica has recently been redescribed as two distinct species; E. histolytica and E. dispar. In the present study, we characterized the 70 kDa Heat Shock Protein (HSP70) of E. dispar at molecular level and compared it with that of E. histolytica. With these findings, we were able to distinguishe E. dispar from the infectious E. histolytica. Pairs of 21 nucleotide primers were designed from highly conserved regions of the same gene in other eukaryotic cells. Mentioned primers were utilized in PCR by using isolated genomic DNA template of E. dispar and the PCR fragments were then sequenced. By the time, 1020 nucleotides have been sequenced and characterized within open reading frame of this new gene which encode a polypeptide with 337 amino acids. Nucleotide sequence comparison in gene data banks (NCBI, NIH) for both the partial DNA and its deduced amino acid sequence revealed significant homology with members of the eukaryotic 70 kDa HSP family. Small parts of the mentioned sequences from E. dispar were about 100% identical to the sequences of 70 kDa HSP from E. histolytica other eukaryotic cells. The new partial gene fragment and its encoded protein have been submitted to the gene data banks (NCBI, NIH) and registered under the accession number of AY763790.
Distance-dependent coherent charge transport in DNA: crossover from tunneling to free propagation  [PDF]
Natallia V. Grib, Dmitry A. Ryndyk, Rafael Gutiérrez, Gianaurelio Cuniberti
Journal of Biophysical Chemistry (JBPC) , 2010, DOI: 10.4236/jbpc.2010.12010
Abstract: Using a tight-binding model, we investigate the influence of intra- and interstand coupling parameters on the charge transport properties in a G-(T)j-GGG DNA sequence and its (G:C)-(T:A)j-(G:C)3 duplex attached to four electrodes. Dependences of the transmission function and of the corresponding conductance of the system on the number of bridging sites were obtained. Simulation results of a recently proposed two-strand superexchange (tunneling) model were reproduced and extended. It is demonstrated that the crossover from strong to weak distance-dependent charge transport is elucidated by a transition from under-barrier tunneling mechanism to free over-barrier propagation in the coherent regime, controlled by temperature and coupling parameters. The role of DNA- electrode coupling has been also considered. It was found that an asymmetry in the DNA-electrode coupling has a drastic effect on the con-ductance leading to an increase in delocaliza-tion of the electronic states in the DNA duplex.
Uptake and Organ Distribution of Feed Introduced Plasmid DNA in Growing or Pregnant Rats  [PDF]
Idun M. Gr?nsberg, Lise Nordg?rd, Kristin Fenton, Beate Hegge, Kaare M. Nielsen, Susan Bardocz, Arpad Pusztai, Terje Traavik
Food and Nutrition Sciences (FNS) , 2011, DOI: 10.4236/fns.2011.24053
Abstract: Fragments of DNA present in food and feed are taken up by the gastrointestinal tract (GIT) of mammals. The extent of uptake varies according to organism, study design and DNA source. This study explores the hypothesis that actively growing, as well as pregnant rats, are more likely to take up DNA from the GIT than mature animals due to the high demand for nutrients for tissue and organ development. Plasmid DNA (pDNA) was added to standard feed for growing, and pregnant rats. The young rats received one pDNA (50 μg) containing meal by gavage. Blood, organ and tissue samples were harvested at 2 h to 3 days post feeding (p.f). The pregnant females were fed pellets containing pDNA (100 μg) daily, starting at day 5 after established pregnancy. Females and foeti were killed at days 7 and 14 of gestation, and pups at the time of weaning. Genomic DNA was analyzed by PCR followed by Southern blot and real-time PCR. A 201 bp target sequence was detected in mesenteric lymph nodes, spleen, liver and pancreas from growing rats 2 h p.f. At 6 h, target DNA was detectable in the kidneys, and at three days p.f. in the liver. Target DNA was not detected in samples from pregnant rats, their foeti or pups. In conclusion, low level of feed introduced DNA could be transiently detected in organs of young, growing rats. However, indications of increased DNA uptake levels in the GIT of growing rats were not found.
Thermodynamic contributions of 5’- and 3’-single strand dangling-ends to the stability of short duplex DNAs  [PDF]
Rebekah Dickman, Fidelis Manyanga, Greg P. Brewood, Daniel J. Fish, Cameron A. Fish, Charlie Summers, M. Todd Horne, Albert S. Benight
Journal of Biophysical Chemistry (JBPC) , 2012, DOI: 10.4236/jbpc.2012.31001
Abstract: Differential scanning calorimetry (DSC) melting analysis was performed on 27 short double stranded DNA duplexes containing 15 to 25 base pairs. Experimental duplexes were divided into two categories containing either two 5’ dangling-ends or one 5’ and one 3’ dangling-end. Duplex regions were incrementally reduced from 25 to 15 base pairs with a concurrent increase in length of dangling-ends from 1 to 10 bases. Blunt-ended duplexes from 15 to 25 base pairs served as controls. An additional set of molecules containing 21 base pair duplexes and a single four base dangling-end were also examined. DSC melting curves were measured in varying concentrations of sodium ion (Na+). From these measurements, thermodynamic parameters for 5’ and 3’ dangling-ends were evaluated as a function of dangling end length. 5’ ends were found to be slightly stabilizing but essentially constant while the 3’ ends were destabilizing with increasing length of the dangling-end. 3’ ends also display a stronger dependence on Na+ concentration. In lower Na+ environment, the 3’ ends were more destabilizing than in higher salt environment suggesting a more significant electrostatic component of the destabilizing interactions. Analysis of thermodynamic parameters of dangling ended duplexes as a function of Na+ concentration indicated the 3' dangling ends behave differently than 5' dangling ended and blunt-ended duplexes. Molecules with one 5' and one 3' dangling end showed variation in excess specific heat capacity (ΔCp) when compared to the blunt-ended molecule, while the molecules with two 5’ ends had ΔCp values that were essentially the same as blunt-ended duplexes. These observations suggested differences exist in duplexes with 3’ and 5’ dangling ends, which are interpreted in terms of composite differences in interactions with Na+, solvent, and terminal base pairs of the duplex.
Murine embryo development following cytoplasmic injection of linear and condensed DNA  [PDF]
Marya Dunlap-Brown, Stephen P. Butler, William H. Velander, Francis C. Gwazdauskas
Open Journal of Animal Sciences (OJAS) , 2012, DOI: 10.4236/ojas.2012.24034
Abstract: In 1985 Brinster et al. [1] observed that linearized DNA injected in the cytoplasm of mouse zygotes underwent spontaneous supercoiling within 24 h. This finding suggests that DNA prefers and functions best in tertiary structure. In an effort to improve the efficiency of transgenesis by cytoplasmic injection, DNA was condensed with MgCl2to form a three dimensional rod-shaped DNA prior to injection in pronuclear stage murine zygotes. The DNA used was enhanced green fluorescent protein on a cytomegalovirus promoter (CMV-EGFP) and served as a marker for gene integration and protein expression in culture conditions. The condensed CMV-EGFP construct was injected in the cytoplasm at 3 concentrations (100, n = 816; 425, n = 464; and 625 μg/ml, n = 708). For comparison linear CMV-EGFP construct was injected into the pronucleus (5 μg/ml, n = 196) and into the cytoplasm (625 μg/ml, n = 628). In all treatment groups the control and buffer injected embryos developed similarly. Among DNA treatment groups, the highest development of fluorescing embryos was observed in zygotes injected in the cytoplasm with linear CMV-EGFP (625 μg/ml); however, zygotes injected in the cytoplasm with condensed CMV-EGFP (625 μg/ml) had the highest percentage (44%) of fluorescing embryos, the highest percentage (16.7%) of fluorescing morula and blastocysts, and the lowest percentage of fluorescence mosaicism at every stage of embryo development after 4 d in culture; thereby making it the best method for generating transgenic animals.
Directly immobilized DNA sensor for label-free detection of herpes virus  [PDF]
Phuong Dinh Tam, Mai Anh Tuan, Nguyen Duc Chien
Journal of Biomedical Science and Engineering (JBiSE) , 2009, DOI: 10.4236/jbise.2009.26054
Abstract: This paper reports the direct immobilization of deoxyribonucleic acid (DNA) sequences of Herpes simplex virus (5’–AT CAC CGA CCC GGA GAG GGA C–3’) on the surface of DNA sensor by using the cyclic voltammetric method with the presence of pyrrole. The potential was scanned from –0.7 volt to + 0.6 volt, the scanning rate was at 100mV/s. This kind of DNA sensor was developed to detect Herpes virus DNA in real samples. The FTIR was applied to verify specific binding of DNA sequence and conducting polymer, the morphology of conducting polymer doped with DNA strands was investigated by using a field emission scanning electron microscope (FE-SEM). The results showed that output signal given by coimmobilized DNA/PPy membrane sensor was better than that given by APTS immobilized membrane sensors. The sensor can detect as low as 2 nM of DNA target in real samples.
Identification of DNA Photolyase Gene of a Drought Tolerance Branching Line of Kenaf (Hibiscus cannabinus L.)  [PDF]
Estri Laras Arumingtyas, Nur Basuki, Sutiman Bambang Sumitro, Sudjindro  
American Journal of Plant Sciences (AJPS) , 2012, DOI: 10.4236/ajps.2012.36098
Abstract: DNA photolyase is a photoreactivation enzyme which has a role in the mechanism of DNA repair. The existence of DNA photolyase gene could affect the success of mutation process. SM026H is a drought tolerance branching line of kenaf identified by Research Institute for Tobacco and Fibre Crops, Karangploso, Malang, Indonesia which has a potential as a source for mutation breeding. In this research, cloning and sequencing of the DNA photolyase gene of line SM026H were conducted. The DNA of SM026H leaves was isolated using DNeasy Kit, then was amplified by Polymerase Chain Reaction (PCR) using AC1 as a forward primer, paired with AC3R or AC4R as reverse primers and checked using electrophoresis on a 0.7% agarose gel. The AC1-AC3R primer pair produced a DNA fragment with a size of 750 bp, whereas the AC1-AC4R primer pair produced a DNA fragment of 1000 bp in size. Cloning of the AC1-AC3R and AC1-AC4R fragments was done prior to sequencing. Preparation for cloning was done by running and extracting the PCR products from a 0.7% Low Melting Agarose (LMA), and then ligating them to a pCR21 plasmid using an electroporation method. Two sub-fragments of each AC1-AC3R and AC1-AC4R fragments were identified. They were 600 bp and 500 bp, resulting from the AC1-AC3R fragment, and 1300 bp and 500 bp, resulting from the AC1-AC4R fragment. The sequencing result of those sub-fragments was analyzed using a Basic Local Alignment Search Tool (BLAST) program. It was shown that the sequences have a degree of homology to DNA photolyase sequence of Arabidopsis thaliana A. thaliana, Cucumis sativus, Spinacia oleracea, Stellaria longipes and Oryza sativa. These suggested that the kenaf line SM0026H possesses the DNA photolyase gene. However, further study needs to be done to ascertain that the gene was not the cryptochrome.
Forensic odontostomatology  [PDF]
Geoffrey H. Sperber
Forensic Medicine and Anatomy Research (FMAR) , 2013, DOI: 10.4236/fmar.2013.14019
Abstract: Teeth are the most durable and enduring structures of human anatomy, surviving fragmentation, partial incineration and severe decomposition. The role of teeth in identification is manifested as significant specifiers of deceased or living individuals. The characteristics of dentalmorphology are genetically-inherited and ascribable to racial or familial ancestry. Dental age identification is attributable to young individuals. The chemical composition of teeth identifies diets during life, as attritional wear patterns do. Bite marks transiently relate to the perpetrator of attacks. Dental restorations and prostheses are evidence of economic, cultural and social status of deceased individuals. Palatal rugae patterns are unique to individuals. The DNA identification of postmortem dental pulp tissue relating to a deceased individual or a living relative is the ultimate criterion of positive association of forensic recognition, as other means of identification become less effective, forensic dental identification increases in importance.
Degradation of nucleic acids and nucleotides in several conditions with perspectives of retrieval: A review  [PDF]
José Martínez Reyes, Lenin Ejecatl Medina Orozco, Melitón Estrada Jaramillo, Iván Vera Romero, Agustina Ortiz Soriano
Advances in Bioscience and Biotechnology (ABB) , 2014, DOI: 10.4236/abb.2014.51006
Abstract:

Deoxyribonucleic acid (DNA) or oligonucleotides, can be modified in several ways as chemical degradation by electrophilic reaction, attack of radicals, hydrolytic deamination or oxidative damage caused by ionizing radiation. This work discussed these degradation mechanisms, determining the effects on these biomolecules. The actual knowledge about DNA damages only permits partial enzymatic repair treatments.

Methylation Abnormalities in Mammary Carcinoma: The Methylation Suicide Hypothesis  [PDF]
Anne H. O’Donnell, John R. Edwards, Robert A. Rollins, Nathan D. Vander Kraats, Tao Su, Hanina H. Hibshoosh, Timothy H. Bestor
Journal of Cancer Therapy (JCT) , 2014, DOI: 10.4236/jct.2014.514131
Abstract: Promoter silencing by ectopic de novo methylation of tumor suppressor genes has been proposed as comparable or equivalent to inactivating mutations as a factor in carcinogenesis. However, this hypotheses had not previously been tested by high resolution, high-coverage whole-genome methylation profiling in primary carcinomas. We have determined the genomic methylation status of a series of primary mammary carcinomas and matched control tissues by examination of more than 2.7 billion CpG dinucleotides. Most of the tumors showed variable losses of DNA methylation from all sequence compartments, but increases in promoter methylation were infrequent, very small in extent, and were observed largely at CpG-poor promoters. De novo methylation at the promoters of proto-oncogenes and tumor suppressor genes occurred at approximately the same frequency. The findings indicate that tumor suppressor silencing by de novo methylation is much less common than currently believed. We put forward a hypothesis under which the demethylation commonly observed in carcinomas is a manifestation of a defensive system that kills incipient cancer cells.
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