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Complete Sequencing (complete + sequencing)
Selected AbstractsCover Picture: Electrophoresis 17'09ELECTROPHORESIS, Issue 17 2009Article first published online: 26 AUG 200 Issue no. 17 is an Emphasis Issue with 10 articles on various aspects of "Proteins and Proteomics" while the remaining 10 articles are arranged into 3 different parts including "Chip Technology", "Nucleic Acids", and "Methodologies, Assays and Applications." Selected articles are: Proteomic analysis of Oenococcus oeni freeze-dried culture to assess the importance of cell acclimation to conduct malolactic fermentation in wine ((10.1002/elps.200900228)) Complete sequencing and oxidative modification of Mn-SOD (SOD2) in medulloblastoma cells ((10.1002/elps.200900168)) Differentiation of Staphylococcus aureus strains by capillary electrophoresis, zeta potential and coagulase gene polymorphism ((10.1002/elps.200900186)) [source] m.6267G>A: a recurrent mutation in the human mitochondrial DNA that reduces cytochrome c oxidase activity and is associated with tumors,HUMAN MUTATION, Issue 6 2006M. Esther Gallardo Abstract Complete sequencing of the mitochondrial genome of 13 cell lines derived from a variety of human cancers revealed nine novel mitochondrial DNA (mtDNA) variations. One of them, m.6267G>A, is a recurrent mutation that introduces the Ala122Thr substitution in the mitochondrially encoded cytochrome c oxidase I (MT-CO1): p.MT-CO1: Ala122Thr (GenBank: NP_536845.1). Biochemical analysis of the original cell lines and the transmitochondrial cybrids generated by transferring mitochondrial DNAs to a common nuclear background, indicate that cytochrome c oxidase (COX) activity, respiration, and growth in galactose are impaired by the m.6267G>A mutation. This mutation, found twice in the cancer cell lines included in this study, has been also encountered in one out of 63 breast cancer samples, one out of 64 colon cancer samples, one out of 260 prostate cancer samples, and in one out of 15 pancreatic cancer cell lines. In all instances the m.6267G>A mutation was associated to different mtDNA haplogroups. These findings, contrast with the extremely low frequency of the m.6267G>A mutation in the normal population (1:2264) and its apparent absence in other pathologies, strongly suggesting that the m.6267G>A missense mutation is a recurrent mutation specifically associated with cancer. Hum Mutat 27(6), 575,582, 2006. © 2006 Wiley-Liss, Inc. [source] Sequencing, expression, and characterization of cDNA expressed flavin-containing monooxygenase 2 from mouseJOURNAL OF BIOCHEMICAL AND MOLECULAR TOXICOLOGY, Issue 6 2001Edward D. Karoly Abstract The cDNA clone of mouse flavin-containing monooxygenase 2 (FMO2) was obtained as an expressed sequence tag (EST) isolated from a female mouse kidney cDNA library from the I.M.A.G.E. consortium (I.M.A.G.E. CloneID 1432164). Complete sequencing of the EST derived a nucleotide sequence for mouse FMO2, which contains 112 bases of 5, flanking region, 1607 bases of coding region, and 309 bases of 3, flanking region. This FMO2 sequence encodes a protein of 535 amino acids including two putative pyrophosphate binding sequences (GxGxxG/A) beginning at positions 9 and 191. Additionally, this mouse FMO protein sequence shows 87 and 86% homology to rabbit and human FMO2 respectively. The mouse FMO2 sequence was subcloned into the expression vector pJL-2, a derivative of pKK233-2 and used to transform XL1-Blue Escherichia coli. FMO activity in particulate fractions isolated from isopropyl-,-D-thiogalactopyanoside (IPTG) induced cells was heat stable (45°C for 5 min) and demonstrated optimal activity at a relatively high pH of 10.5. The expressed FMO2 enzyme showed catalytic activity towards the FMO substrate methimazole and further analysis of E. coli fractions utilizing NADPH oxidation demonstrated that the mouse FMO2 enzyme also exhibits catalytic activity towards thiourea, trimethylamine, and the insecticide phorate. © 2001 John Wiley & Sons, Inc. J Biochem Mol Toxicol 15:300,308, 2001 [source] An optimized microchip electrophoresis system for mutation detection by tandem SSCP and heteroduplex analysis for p53,gene exons,5,9ELECTROPHORESIS, Issue 19 2006Christa N. Hestekin Abstract With the complete sequencing of the human genome, there is a growing need for rapid, highly sensitive genetic mutation detection methods suitable for clinical implementation. DNA-based diagnostics such as single-strand conformational polymorphism (SSCP) and heteroduplex analysis (HA) are commonly used in research laboratories to screen for mutations, but the slab gel electrophoresis (SGE) format is ill-suited for routine clinical use. The translation of these assays from SGE to microfluidic chips offers significant speed, cost, and sensitivity advantages; however, numerous parameters must be optimized to provide highly sensitive mutation detection. Here we present a methodical study of system parameters including polymer matrix, wall coating, analysis temperature, and electric field strengths on the effectiveness of mutation detection by tandem SSCP/HA for DNA samples from exons,5,9 of the p53 gene. The effects of polymer matrix concentration and average molar mass were studied for linear polyacrylamide (LPA) solutions. We determined that a matrix of 8%,w/v 600,kDa LPA provides the most reliable SSCP/HA mutation detection on chips. The inclusion of a small amount of the dynamic wall-coating polymer poly- N -hydroxyethylacrylamide in the matrix substantially improves the resolution of SSCP conformers and extends the coating lifetime. We investigated electrophoresis temperatures between 17 and 35°C and found that the lowest temperature accessible on our chip electrophoresis system gives the best condition for high sensitivity of the tandem SSCP/HA method, especially for the SSCP conformers. Finally, the use of electrical fields between 350 and 450,V/cm provided rapid separations (<10,min) with well-resolved DNA peaks for both SSCP and HA. [source] Assessment of the rind microbial diversity in a farmhouse-produced vs a pasteurized industrially produced soft red-smear cheese using both cultivation and rDNA-based methodsJOURNAL OF APPLIED MICROBIOLOGY, Issue 3 2004C. Feurer Abstract Aims:, The diversity of the surface flora of two French red-smear soft cheeses was examined by cultivation-dependent and cultivation-independent methods to assess their composition and to evaluate the accuracy of both approaches. Methods and Results:, Culture-independent methods used involved 16S ribosomal DNA gene cloning and sequencing and single-strand conformation polymorphism analysis (SSCP). The culture-dependent method used involved direct culture and macroscopic observation, polymerase chain reaction of the 16S rRNA gene from DNA extracted from single colonies followed by complete sequencing of the gene. Only few species were recovered by both approaches either in the pasteurized and the farmer cheese. A large diversity of isolates or 16S rDNA sequences related to marine bacteria was identified at the surface of both cheeses. Conclusions:, The results indicated that all three techniques were informative and complementary to allow a more accurate representativeness of the cheese surface biodiversity. Significance and Impact of the Study:, Cultivation and molecular methods have to be combined in order to obtain an extended view of the bacterial populations of complex ecosystems. [source] Characterization of xylanolytic bacteria present in the bract phyllosphere of the date palm Phoenix dactyliferaLETTERS IN APPLIED MICROBIOLOGY, Issue 2 2007R. Rivas Abstract Aims:, Despite the interest of phyllosphere microbiology, no studies have addressed the bacteria present in bract phyllosphere, an ecosystem that has special characteristics in palm trees because the dry bracts remain on the plant until pruning and may contain polymer-degrading bacteria involved in plant degradation. Therefore, the aim of this work was to characterize xylanolytic bacteria isolated from palm bract phyllosphere. Methods and Results:, Twelve xylanolytic strains were isolated and characterized by phenotypic features and complete sequencing of 16S rRNA gene. The results showed that the isolates were phenotypically and genotypically diverse. Gram-positive isolates were classified into genus Paenibacillus some of them belonging to hitherto undescribed species of this genus. Gram-negative isolates were classified into genera Pseudomonas and Acinetobacter. Conclusions:, The results of this work confirm the complexity of the bacterial populations present in phyllospheric ecosystems and suggest that bacteria involved in plant degradation are present at the early degradation steps of this process in dry palm tree bracts. Significance and Impact of the Study:, This is the first study on bract phyllospheric bacteria able to hydrolyse vegetal polymers and offers a new perspective in the search of unexplored sources of xylanase-producing strains. [source] The few virus-like genes of Cotesia congregata bracovirus ,ARCHIVES OF INSECT BIOCHEMISTRY AND PHYSIOLOGY (ELECTRONIC), Issue 3 2006J-M. Drezen Abstract The origin of the symbiotic association between parasitoid wasps and bracoviruses is still unknown. From phylogenetic analyses, bracovirus-associated wasp species constitute a monophyletic group, the microgastroid complex. Thus all wasp,bracovirus associations could have originated from the integration of an ancestral virus in the genome of the ancestor of the microgastroids. In an effort to identify a set of virus genes that would give clues on the nature of the ancestral virus, we have recently performed the complete sequencing of the genome of CcBV, the bracovirus of the wasp Cotesia congregata. We describe here the putative proteins encoded by CcBV genome having significant similarities with sequences from known viruses and mobile elements. The analysis of CcBV gene content does not lend support to the hypothesis that bracoviruses originated from a baculovirus. Moreover, no consistent homology was found between CcBV genes and any set of genes constituting the core genome of a known free-living virus. We discuss the significance of the scarce homology found between proteins from CcBV and other viruses or mobile elements. Arch. Insect Biochem. Physiol. 61:110,122, 2006. © 2006 Wiley-Liss, Inc. 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