Mutation Research/Mutation Research Genomics最新文献_第4页

Rapid characterization of mutations in amplified human hprt cDNA by polyacrylamide gel electrophoresis 用聚丙烯酰胺凝胶电泳快速鉴定扩增的人hprt cDNA突变

Mutation Research/Mutation Research Genomics Pub Date : 1998-11-01 DOI: 10.1016/S1383-5726(98)00008-9

Fátima Garganta, Günter Krause, Gerhard Scherer

{"title":"Rapid characterization of mutations in amplified human hprt cDNA by polyacrylamide gel electrophoresis","authors":"Fátima Garganta, Günter Krause, Gerhard Scherer","doi":"10.1016/S1383-5726(98)00008-9","DOIUrl":"10.1016/S1383-5726(98)00008-9","url":null,"abstract":"<div>Compiling hprt mutation spectra involves the isolation and analysis of numerous 6-thioguanine-resistant clones for identifying characteristic point mutations. Since cDNA amplificates are compulsary intermediates in most mutant classification protocols, we suggest their preliminary characterization by polyacrylamide gel electrophoresis for the rapid distinction of clonal and independent mutants and for streamlining mutant analysis procedures. Based on the human hprt cDNA sequence a strategy was developed for mapping missing exons by analytical digests with a small panel of restriction enzymes. In mutant classification schemes, polyacrylamide gel electrophoresis of AluI-digested cDNA amplificates increased the sensitivity for detecting RT-PCR products of reduced size, e.g., in the case of missing exon 5. Restriction analysis of cDNA amplificates from 109 independent mutant clones showed a significant increase of exon loss after NNK induction as compared to spontaneous or BaP-induced mutants. The determination of exon loss from cDNA amplificates, as carried out for 39 independent mutant PCR products, might direct towards the genomic target sequences carrying the point mutations, that caused the aberrant splicing, thus eliminating the need of laborious multiplex PCR comprising all exons. For single-strand conformation polymorphism (SSCP) analysis of five known point mutations, sub-amplificates comprising exons 7 and 8 of hprt cDNA were obtained. After a combined heat and alkali denaturation of the double-stranded PCR products, the samples were separated in pre-cast polyacrylamide gels under non-denaturing conditions. Five known nucleotide substitutions within the amplified region, including the C508T hot spot mutation, resulted in mobility shifts of single-strand bands relative to the wild type pattern.</div>","PeriodicalId":100939,"journal":{"name":"Mutation Research/Mutation Research Genomics","volume":"406 1","pages":"Pages 33-43"},"PeriodicalIF":0.0,"publicationDate":"1998-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1016/S1383-5726(98)00008-9","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"20825784","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 3

Polymorphisms in the human DNA ligase I gene (LIG1) including a complex GT repeat 人类DNA连接酶I基因(LIG1)的多态性，包括一个复杂的GT重复序列

Mutation Research/Mutation Research Genomics Pub Date : 1998-11-01 DOI: 10.1016/S1383-5726(98)00005-3

Kenneth J Livak, Wayne A Little, Sylvia L Stack, Thomas A Patterson

{"title":"Polymorphisms in the human DNA ligase I gene (LIG1) including a complex GT repeat","authors":"Kenneth J Livak, Wayne A Little, Sylvia L Stack, Thomas A Patterson","doi":"10.1016/S1383-5726(98)00005-3","DOIUrl":"10.1016/S1383-5726(98)00005-3","url":null,"abstract":"<div>Sequencing of a human DNA ligase I cDNA clone derived from HeLa cells revealed two unreported differences with the published sequence: a single base change and a three-base deletion. Both differences are in exon 6, and were analyzed by amplifying a segment containing exon 5, intron 6, and exon 6. The first finding was that intron 6 is approximately 2.6 kb in size, not the 1 kb reported in the literature. By sequence analysis of amplified segments, the single-base difference in exon 6 was shown to be polymorphic, with HeLa cells heterozygous for the A/C difference. Analysis of 60 unrelated individuals found a frequency of 0.5 for each allele. Primer extension reactions across the exon 5/exon 6 boundary were performed on cDNA obtained from HeLa cells and human thymus. The results show that the three-base deletion is due to a variation in splicing. For both HeLa and thymus, two-thirds of the transcripts are like the published cDNA sequence and one-third have the three-base deletion. Finally, sequencing of part of intron 6 revealed the presence of a complex GT repeat consisting of a 48–50 nucleotide polypurine tract followed by a variable number of GT residues. This entire unit of polypurine tract plus GTs is repeated three times. Detection of the repeated sequences required the development of specialized cloning and PCR conditions. Analysis of a pedigree showed that this complex repeat is polymorphic.</div>","PeriodicalId":100939,"journal":{"name":"Mutation Research/Mutation Research Genomics","volume":"406 1","pages":"Pages 1-8"},"PeriodicalIF":0.0,"publicationDate":"1998-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1016/S1383-5726(98)00005-3","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"20825781","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 9

Trinucleotide repeat length variation in the human ribosomal protein L14 gene (RPL14): localization to 3p21.3 and loss of heterozygosity in lung and oral cancers 人核糖体蛋白L14基因(RPL14)的三核苷酸重复长度变异:定位于3p21.3和肺癌和口腔癌的杂合性缺失

Mutation Research/Mutation Research Genomics Pub Date : 1998-11-01 DOI: 10.1016/S1383-5726(98)00006-5

Sharon P. Shriver , Mark D. Shriver , Dayna L. Tirpak , Lillian M. Bloch , Jay D. Hunt , Robert E. Ferrell , Jill M. Siegfried

{"title":"Trinucleotide repeat length variation in the human ribosomal protein L14 gene (RPL14): localization to 3p21.3 and loss of heterozygosity in lung and oral cancers","authors":"Sharon P. Shriver , Mark D. Shriver , Dayna L. Tirpak , Lillian M. Bloch , Jay D. Hunt , Robert E. Ferrell , Jill M. Siegfried","doi":"10.1016/S1383-5726(98)00006-5","DOIUrl":"10.1016/S1383-5726(98)00006-5","url":null,"abstract":"<div>Chromosome 3p is consistently deleted in lung cancer, oral squamous cell carcinoma, and renal cell carcinoma, and is believed to contain several tumor suppressor genes. We have shown a role for chromosome 3 in tumor suppression by microcell-mediated chromosome transfer experiments. We have isolated a gene that is located at 3p21.3 within the smallest region of deletion overlap in lung tumors and is the human homolog of the ribosomal protein L14 gene (RPL14). The RPL14 sequence contains a highly polymorphic trinucleotide repeat array which encodes a variable-length polyalanine tract. Genotype analysis of RPL14 shows that this locus is 68% heterozygous in the normal population, compared with 25% in non-small cell lung cancer (NSCLC) cell lines (p=0.008). Cell cultures derived from normal bronchial epithelium show a 65% level of heterozygosity, reflecting that of the normal population. Squamous cell carcinoma of the head and neck (SCCHN), which has the same risk factors as lung cancer and is hypothesized to have a similar etiology, demonstrates 54% loss of heterozygosity at the RNA level, suggesting that transcriptional loss may be a primary mechanism of RPL14 alteration in SCCHN. In addition, RPL14 shows significant differences in allele frequency distribution in ethnically-defined populations, making this sequence a useful marker for the study of ethnicity-adjusted lung cancer risk.</div>","PeriodicalId":100939,"journal":{"name":"Mutation Research/Mutation Research Genomics","volume":"406 1","pages":"Pages 9-23"},"PeriodicalIF":0.0,"publicationDate":"1998-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1016/S1383-5726(98)00006-5","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"20825782","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 38

Application of random amplified polymorphic DNA PCR for genomic analysis of HIV-1-infected individuals 随机扩增多态性DNA PCR在hiv -1感染者基因组分析中的应用

Mutation Research/Mutation Research Genomics Pub Date : 1998-11-01 DOI: 10.1016/S1383-5726(98)00007-7

Felix O Aikhionbare , Cheryl Newman , Chad Womack , William Roth , Ketan Shah , Vincent C Bond

{"title":"Application of random amplified polymorphic DNA PCR for genomic analysis of HIV-1-infected individuals","authors":"Felix O Aikhionbare , Cheryl Newman , Chad Womack , William Roth , Ketan Shah , Vincent C Bond","doi":"10.1016/S1383-5726(98)00007-7","DOIUrl":"10.1016/S1383-5726(98)00007-7","url":null,"abstract":"<div>Random amplified polymorphic DNA polymerase chain reaction (RAPD-PCR) is a DNA fingerprinting technique used to detect genomic polymorphisms. We employed sixteen different RAPD-PCR 10-mer primers to amplify DNA from the peripheral blood mononuclear cells (PBMC) of 80 HIV-1-infected individuals. These individuals were previously identified as either heterozygotes (+/Δ32) and homozygotes (+/+) for the CCR5 locus by PCR with gene specific primers. Four of the sixteen randomly selected RAPD primers produced distinguishable banding profiles between CCR5 (+/Δ32) heterozygotes and CCR5 (+/+) homozygotes. Direct sequencing of some RAPD-PCR products obtained with one of the four RAPD primers that were tested yielded clearly readable, but limited sequences, which were similar to portions of the previously published sequences for (+/+) homozygotes (98% similarity) and (+/Δ32) heterozygotes (87% similarity) of the CCR5 alleles. Thus, the RAPD-PCR technique may be useful for the identification of human molecular markers that may correlate with susceptibility to HIV-1-infection, or differences in disease progression among HIV-1-infected individuals.</div>","PeriodicalId":100939,"journal":{"name":"Mutation Research/Mutation Research Genomics","volume":"406 1","pages":"Pages 25-31"},"PeriodicalIF":0.0,"publicationDate":"1998-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1016/S1383-5726(98)00007-7","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"20825783","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 2

Chromosomal assignment of loci susceptible to replication errors by radiation hybrid mapping 辐射杂交作图对复制错误易感位点的染色体分配

Mutation Research/Mutation Research Genomics Pub Date : 1998-05-01 DOI: 10.1016/S1383-5726(97)00015-0

Maja Krajinovic , Chantal Richer , Ljiljana Lukovic , Damian Labuda , Daniel Sinnett

引用次数: 4

Genetic variation within the ovine cystic fibrosis transmembrane conductance regulator gene1 绵羊囊性纤维化跨膜电导调节基因1的遗传变异

Mutation Research/Mutation Research Genomics Pub Date : 1998-05-01 DOI: 10.1016/S1383-5726(97)00012-5

Scott J Tebbutt, Michael B Lakeman, Jane C Wilson-Wheeler, Diana F Hill

引用次数: 6

A novel missense mutation and frameshift mutations in the type II receptor of transforming growth factor-β gene in sporadic colon cancer with microsatellite instability 散发性结肠癌伴微卫星不稳定性转化生长因子-β基因II型受体的一个新的错义突变和移码突变

Mutation Research/Mutation Research Genomics Pub Date : 1998-05-01 DOI: 10.1016/S1383-5726(98)00003-X

Hideo Orimo , Miyoko Ikejima , Eiitsu Nakajima , Mitsuru Emi , Takashi Shimada

{"title":"A novel missense mutation and frameshift mutations in the type II receptor of transforming growth factor-β gene in sporadic colon cancer with microsatellite instability","authors":"Hideo Orimo , Miyoko Ikejima , Eiitsu Nakajima , Mitsuru Emi , Takashi Shimada","doi":"10.1016/S1383-5726(98)00003-X","DOIUrl":"https://doi.org/10.1016/S1383-5726(98)00003-X","url":null,"abstract":"<div>Microsatellite instability of DNA samples of 79 sporadic colon cancer patients were analyzed. These samples were also screened to search mutations in the repeat sequences in the gene for the type II receptor of transforming growth factor-β (TGF-β RII) using polymerase chain reaction (PCR), electrophoresis with urea gel, and PCR-single strand conformation polymorphism (PCR-SSCP) method. The incidence of microsatellite instability, defined as severe replication error phenotype (RER) with microsatellite alterations in more than three loci, was 6%. Deletion and insertion of an A residue in the (A)10 region, which cause frameshift mutation, were found in four samples and their incidence in the samples with microsatellite instability was 80%. A novel nucleotide substitution of T for G at 1918, which causes missense mutation of arginine to leucine at codon 528, was found in a sample with microsatellite instability. The mutation at 1918 was in highly conservative amino acid residue.</div>","PeriodicalId":100939,"journal":{"name":"Mutation Research/Mutation Research Genomics","volume":"382 3","pages":"Pages 115-120"},"PeriodicalIF":0.0,"publicationDate":"1998-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1016/S1383-5726(98)00003-X","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"72110372","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 9

Decreased mitochondrial RNA levels without accumulation of mitochondrial DNA deletions in aging Drosophila melanogaster 衰老的黑腹果蝇线粒体RNA水平降低而线粒体DNA缺失不积累

Mutation Research/Mutation Research Genomics Pub Date : 1998-05-01 DOI: 10.1016/S1383-5726(97)00013-7

Steven R Schwarze , Richard Weindruch , Judd M Aiken

{"title":"Decreased mitochondrial RNA levels without accumulation of mitochondrial DNA deletions in aging Drosophila melanogaster","authors":"Steven R Schwarze , Richard Weindruch , Judd M Aiken","doi":"10.1016/S1383-5726(97)00013-7","DOIUrl":"10.1016/S1383-5726(97)00013-7","url":null,"abstract":"<div>Declines in electron transport system (ETS) activity have been reported to occur with advancing age in Drosophila melanogaster and many other animals. It has been proposed that these changes are importantly involved in the aging process. ETS decline has been attributed to mitochondrial nucleic acid damage. We analyzed various ages of D. melanogaster (embryos to 60-day-old adults) for the presence of mutated mitochondrial DNA (mtDNA) genomes. Although mtDNA genomes with large DNA deletions (up to 5 kb) were identified, abundance was low and remained constant throughout adult life. Therefore, these mtDNA deletions do not appear to be sufficiently abundant to cause large declines in ETS activity. Next, we analyzed various ages of D. melanogaster for the abundance of four mitochondrial-encoded and two nuclear-encoded ETS transcripts. The abundance of the mitochondrial transcripts declined 5–10-fold, while the nuclear-encoded transcripts declined 2–5-fold with advancing age. Separation of flies on the basis of flight loss was used to distinguish physiologic age from chronological age. Insects capable of flight at 30 days of age were found to have a 4-fold higher abundance of cox I mitochondrial-encoded RNA compared to flightless insects. No difference, however, was apparent in the nuclear-encoded β-ATPase RNA level, suggesting only mitochondrial RNA (mtRNA) declines are associated with life expectancy.</div>","PeriodicalId":100939,"journal":{"name":"Mutation Research/Mutation Research Genomics","volume":"382 3","pages":"Pages 99-107"},"PeriodicalIF":0.0,"publicationDate":"1998-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1016/S1383-5726(97)00013-7","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"20608098","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 26

Identification of known p53 point mutations by capillary electrophoresis using unique mobility profiles in a blinded study 在一项盲法研究中，用毛细管电泳鉴定已知的p53点突变

Mutation Research/Mutation Research Genomics Pub Date : 1998-05-01 DOI: 10.1016/S1383-5726(98)00004-1

H.Michael Wenz , Srini Ramachandra , Catherine D O'Connell , Donald H Atha

{"title":"Identification of known p53 point mutations by capillary electrophoresis using unique mobility profiles in a blinded study","authors":"H.Michael Wenz , Srini Ramachandra , Catherine D O'Connell , Donald H Atha","doi":"10.1016/S1383-5726(98)00004-1","DOIUrl":"10.1016/S1383-5726(98)00004-1","url":null,"abstract":"<div>This study is part of an ongoing project at the National Institute of Standards and Technology (NIST) that generates a panel of DNA clones containing the most common mutations found in the human p53 tumor suppressor gene. This panel will be made available as a reference source for evaluation and testing for p53 mutations. Single strand conformation polymorphism (SSCP) analysis has found widespread acceptance as a tool for simply and rapidly screening for mutations, albeit with a detection rate that can be below 100%. We have begun to analyze mutations found in exon 7 of the p53 gene by SSCP using laser induced fluorescence capillary electrophoresis (LIF-CE). PCR fragments, containing single point mutations, were amplified from genomic DNA isolated from cell lines using primers labeled with two different fluorophores. This dual labeling approach allowed better traceability of mobility shifts as a function of the experimental conditions. While analyzing the clones H596, Colo320, Namalwa and wild type (reference samples) at different temperatures, ranging from 25 to 45°C, it was observed that each mutation responded in a unique way to changes in temperature both in magnitude and direction of shifts relative to the wild type sample. In a blinded study, ten p53 exon 7 samples were matched automatically, using ABI PRISM Genotyper® software, against the four reference samples. From these 10 samples, six were correctly identified as containing one of the reference mutations, two corresponded to wild type, and two were correctly identified as non-reference mutations. This approach should prove helpful in the rapid screening of target sequences that are known to bear a limited number of mutations.</div>","PeriodicalId":100939,"journal":{"name":"Mutation Research/Mutation Research Genomics","volume":"382 3","pages":"Pages 121-132"},"PeriodicalIF":0.0,"publicationDate":"1998-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1016/S1383-5726(98)00004-1","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"20608698","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 28

DNA sequence flanking the protein coding regions of the rat Hprt gene 大鼠Hprt基因蛋白编码区两侧的DNA序列

Mutation Research/Mutation Research Genomics Pub Date : 1998-05-01 DOI: 10.1016/S1383-5726(97)00014-9

Tao Chen , Roberta A. Mittelstaedt , Robert H. Heflich

引用次数: 4