Mutation Research/Mutation Research Genomics最新文献

{"title":"The sensitivity of denaturing gradient gel electrophoresis: a blinded analysis","authors":"Pablo V Gejman ,&nbsp;Qiuhe Cao ,&nbsp;Françoise Guedj ,&nbsp;Steve Sommer","doi":"10.1016/S1383-5726(98)00002-8","DOIUrl":"10.1016/S1383-5726(98)00002-8","url":null,"abstract":"<div><p><span>Denaturing gradient gel electrophoresis<span> (DGGE) is considered one of the most sensitive and specific of the mutational scanning techniques, yet blinded analyses have not been reported. We report the results of a blinded study of the efficiency of DGGE to detect mutations in the Human Coagulation Factor IX. Two overlapping </span></span>genomic DNA sequences from exon 8 of Factor IX (290 bp and 539 bp length) with an unknown number of mutations were amplified with a 40 bp GC-clamp and tested blindly by DGGE. DGGE detected all mutations in the 290 bp genomic DNA segment. DGGE detected all but one mutation in the 539 bp genomic segment after experimental conditions were fully optimized but missed multiple mutations in an initial blinded experiment. These results demonstrate the utility of blinded analyses and confirm the exquisite power of DGGE for detecting mutations.</p></div>","PeriodicalId":100939,"journal":{"name":"Mutation Research/Mutation Research Genomics","volume":null,"pages":null},"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)00002-8","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"20608099","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}

{"title":"Enzymatic mutation detection. Phosphate ions increase incision efficiency of endonuclease VII at a variety of damage sites in DNA","authors":"Stefan Golz,&nbsp;Beate Greger,&nbsp;Börries Kemper","doi":"10.1016/S1383-5726(97)00011-3","DOIUrl":"10.1016/S1383-5726(97)00011-3","url":null,"abstract":"<div><p><span>The ability of endonuclease VII (endo VII) to cleave at mispairings in double-stranded DNA has recently been used for enzymatic mutation detection (EMD) [R. Youil, B.W. Kemper, R.G.H. Cotton, Proc. Natl. Acad. Sci. USA 92 (1995) 87–91]. The method is based on mapping cleavages in </span>heteroduplex DNAs obtained from mutant and wildtype sequences. Despite the capability of endo VII to cleave at all possible mispairings, relative cleavage efficiencies vary considerably for individual mismatches and may escape detection if located in an unfavorable sequence surrounding. We report here improved reaction conditions which can increase the selectivity of the enzyme for mismatches up to 500-fold, as demonstrated with a mutation in a 247 nt long fragment from exon 7 of human gene p53. The new conditions involve replacement of Tris/HCl buffer by phosphate buffer and change from pH 8.0 to 6.5. Various concentrations of phosphate ions should be tried in the assay to meet individual requirements of the substrate.</p></div>","PeriodicalId":100939,"journal":{"name":"Mutation Research/Mutation Research Genomics","volume":null,"pages":null},"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)00011-3","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"20608096","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}

{"title":"The design of a new mutation model for active genes: expression of the Escherichia coli lac operon in mammalian cells","authors":"Petra P.H Van Sloun,&nbsp;Paul H.M Lohman,&nbsp;Harry Vrieling","doi":"10.1016/S1383-5726(97)00006-X","DOIUrl":"10.1016/S1383-5726(97)00006-X","url":null,"abstract":"<div><p><span>The design of a novel transgenic mouse model is described that should allow analysis of mutations at a single cell level in all tissues of a model animal. The model is based on the correct regulation of the </span><em>Escherichia coli lac</em><span> operon in mammalian cells. Induction of a mutation in the </span><em>lacI</em> gene will result in the loss of transcriptional repression of the <em>lacZ</em> gene in mutated cells. Expression of β-galactosidase can subsequently be detected at the single cell level. The model was first tested in vitro using transfection of mouse LTK⁻ cells. <em>LacZ</em> expression was very heterogeneous in most of the stable transfectants and seemed to be subject to epigenetic inactivation. One clone (IIB1) was isolated that stably expressed <em>lacZ</em> in more than 99% of its cells. Subsequent introduction of the <em>lacI</em> gene into IIB1 cells resulted in correct transcriptional repression of the <em>lacZ</em> gene that could be alleviated by IPTG, an allosteric inducer of lacI repression. However, in time the extent of β-galactosidase induction gradually declined suggesting that the prolonged repressed transcriptional state triggers epigenetic inactivation. Variegated expression of the <em>lacZ</em> gene was not confined to cultured cells since several transgenic lines also did not express the <em>lacZ</em> transgene. This study shows that while the susceptibility of the <em>lacZ</em><span> gene to inactivation processes poses a fundamental problem, correct regulation of the expression of a reporter gene by the lacI repressor protein is feasible in mammalian cells when assayed at the single cell level. Thus, the model can in principle be used for the detection of mutagenic events at the </span><em>lacI</em> locus. Targeting of the <em>lacZ</em><span> gene to an endogenous housekeeping gene might prevent epigenetic inactivation. Alternatively, with the use of another reporter gene in the mutation detection system the proposed transgenic mouse model could be realized.</span></p></div>","PeriodicalId":100939,"journal":{"name":"Mutation Research/Mutation Research Genomics","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"1997-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1016/S1383-5726(97)00006-X","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"20291422","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}

{"title":"Search for DNA sequence variations using a MutS-based technology","authors":"Christine Bellanné-Chantelot,&nbsp;Sandrine Beaufils,&nbsp;Véronique Hourdel,&nbsp;Suzanne Lesage,&nbsp;Valérie Morel,&nbsp;Nathalie Dessinais,&nbsp;Isabelle Le Gall,&nbsp;Daniel Cohen,&nbsp;Jean Dausset","doi":"10.1016/S1383-5726(97)00007-1","DOIUrl":"10.1016/S1383-5726(97)00007-1","url":null,"abstract":"<div><p><span>The search for DNA sequence<span> variations (DSV) is emphasized with genetic studies of a large number of multifactorial diseases<span><span>. Saturation of regions of interest with diallelic polymorphisms will be an essential step to pinpoint, through association studies, predisposing genes. We have developed a solid-phase method based on the ability of mismatch binding protein MutS to recognize single nucleotide mismatches. This approach was applied to the study of 83 sequence-tagged sites (STSs) extracted from an eight </span>centimorgans (cM) </span></span></span>chromosome 21<span> region. One-third of tested STSs were found to be polymorphic leading to a frequency of one DSV every 822 base pairs (bp). Sequencing of analyzed STSs showed the high reliability of the MutS-based technology for mismatches up to 2 bp in DNA fragments<span> ranging in size from 200 bp to 1 kilobase (kb). The entire assay which is performed in a solid-phase format without the need of electrophoresis or sequencing, will provide an efficient tool for new polymorphism detection.</span></span></p></div>","PeriodicalId":100939,"journal":{"name":"Mutation Research/Mutation Research Genomics","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"1997-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1016/S1383-5726(97)00007-1","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"20291423","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}

{"title":"A nucleotide polymorphism in ERCC1 in human ovarian cancer cell lines and tumor tissues","authors":"Jing Jie Yu ,&nbsp;Chuanjie Mu ,&nbsp;Kang Bo Lee ,&nbsp;Aikou Okamoto ,&nbsp;Edward L Reed ,&nbsp;Frieda Bostick-Bruton ,&nbsp;Kimberly C Mitchell ,&nbsp;Eddie Reed","doi":"10.1016/S1383-5726(97)00004-6","DOIUrl":"10.1016/S1383-5726(97)00004-6","url":null,"abstract":"<div><p><span><span>We studied the DNA sequence<span> of the entire coding region of ERCC1 gene, in five cell lines established from human </span></span>ovarian cancer<span> (A2780, A2780/CP70, MCAS, OVCAR-3, SK-OV-3), 29 human ovarian cancer tumor tissue specimens, one human T-lymphocyte cell line (H9), and non-malignant human ovary tissue (NHO). Samples were assayed by PCR–SSCP and DNA sequence analyses. A silent mutation at codon 118 (site for </span></span>restriction endonuclease<span> MaeII) in exon 4 of the gene was detected in MCAS, OVCAR-3 and SK-OV-3 cells, and NHO. This mutation was a C→T transition, that codes for the same amino acid: asparagine. This transition converts a common codon usage (AAC) to an infrequent codon usage (AAT), whereas frequency of use is reduced two-fold. This base change was associated with a detectable band shift on SSCP analysis. For the 29 ovarian cancer specimens, the same base change was observed in 15 tumor samples and was associated with the same band shift in exon 4. Cells and tumor tissue specimens that did not contain the C→T transition, did not show the band shift in exon 4. Our data suggest that this alteration at codon 118 within the ERCC1 gene, may exist in platinum-sensitive and platinum-resistant ovarian cancer tissues.</span></p></div>","PeriodicalId":100939,"journal":{"name":"Mutation Research/Mutation Research Genomics","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"1997-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1016/S1383-5726(97)00004-6","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"20291421","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}

{"title":"Genomic analysis of single cells from human basal cell cancer using laser-assisted capture microscopy","authors":"Fredrik Pontén ,&nbsp;Cecilia Williams ,&nbsp;Gao Ling ,&nbsp;Afshin Ahmadian ,&nbsp;Monica Nistér ,&nbsp;Joakim Lundeberg ,&nbsp;Jan Pontén ,&nbsp;Mathias Uhlén","doi":"10.1016/S1383-5726(97)00008-3","DOIUrl":"10.1016/S1383-5726(97)00008-3","url":null,"abstract":"<div><p>In this study, we show that direct mutational analysis of genomic DNA<span><span> can be performed on single somatic cells extracted from a frozen, immunohistochemically stained tissue section using laser-assisted capture microscopy. Eighty-nine single tumor cells were separately dissected from one case of human basal cell cancer (BCC) and p53 mutations were analyzed by direct semi-automated sequencing of PCR fragments. Amplification was obtained for at least one of the two analyzed exons from approximately 50% of the single tumor cells. Identical p53 mutations were found in widely spread areas of the tumor, suggesting a clonal proliferation originating from one cell. Interestingly, comparison between results of immunohistochemistry and genetic analysis of the single cells revealed the same p53 mutations irrespective of the p53 </span>immunoreactivity. We propose that this approach has a great potential to allow investigation of genotypic differences in single cells and more specifically to resolve important and fundamental questions determining cancer heterogeneity.</span></p></div>","PeriodicalId":100939,"journal":{"name":"Mutation Research/Mutation Research Genomics","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"1997-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1016/S1383-5726(97)00008-3","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"20291424","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}

{"title":"G to C transversion at a splice acceptor site causes exon skipping in the cystatin B gene","authors":"Irina N Bespalova ,&nbsp;Michael Pranzatelli ,&nbsp;Margit Burmeister","doi":"10.1016/S1383-5726(97)00010-1","DOIUrl":"10.1016/S1383-5726(97)00010-1","url":null,"abstract":"<div><p>Several mutations have been described in the proteinase inhibitor cystatin B<span><span> gene from individuals affected with progressive myoclonus epilepsy of the Unverricht–Lundborg type (EPM1). One of these mutations, a 1925G→C transition at the 3′-splice acceptor site of the intron 1, was postulated to lead to inappropriate splicing of a </span>primary transcript of the cystatin B gene in EPM1 patients. In an effort to understand the expression of the 1925G→C mutation, the sequence of cystatin B mRNA transcripts from lymphoblastoid cell lines of heterozygous patients carrying the mutation were analyzed. RT-PCR of total mRNA showed two main products: the apparently normal transcript and an aberrant, 102 bp shorter transcript. Direct PCR sequencing showed that the aberrant transcript is a consequence of exon 2 skipping.</span></p></div>","PeriodicalId":100939,"journal":{"name":"Mutation Research/Mutation Research Genomics","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"1997-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1016/S1383-5726(97)00010-1","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"20293266","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}

{"title":"Apolipoprotein E R112; R251G: a carboxy-terminal variant found in patients with hyperlipidemia and coronary heart disease","authors":"Amrit K Kang ,&nbsp;David J.A Jenkins ,&nbsp;Thomas M.S Wolever ,&nbsp;Murray W Huff ,&nbsp;Graham F Maguire ,&nbsp;Philip W Connelly ,&nbsp;Robert A Hegele","doi":"10.1016/S1383-5726(97)00009-5","DOIUrl":"10.1016/S1383-5726(97)00009-5","url":null,"abstract":"<div><p><span>A 49 year-old hypercholesterolemic male with marked electrocardiographic ST segment depression<span> on exercise testing was found to have an apo E E3/3 phenotype by isoelectric focusing, but an </span></span><em>APOE</em> E4/3 genotype using <em>Hha</em><span>I restriction isotyping. DNA sequence analysis<span> of the proband's </span></span><em>APOE</em><span> gene found a G→C point mutation at codon 251. This predicted a change in the amino acid encoded by codon 251, from arginine to glycine. The mutation occurred on an allele that encoded arginine at position 112 and this variant was named </span><em>APOE</em> R112; R251G. The R251G change altered a recognition site for the endonuclease <em>Stu</em>I and was the basis for a restriction isotyping method to rapidly screen for this mutation. In relatives of the proband, <em>APOE</em><span><span> R112; R251G was consistently found in subjects with both hyperlipidemia and </span>atherosclerosis<span><span>. Apo E R112; R251G-containing very low density lipoproteins bound normally to macrophages in vitro. However, the proband had an abnormal post-prandial </span>lipoprotein response to a dietary fat challenge. The association of </span></span><em>APOE</em> R112; R251G with abnormal phenotypes suggests that the amino acid change in the carboxy-terminal, perhaps in combination with the common amino acid polymorphism at codon 112, has a functional impact upon lipoprotein metabolism in members of this family.</p></div>","PeriodicalId":100939,"journal":{"name":"Mutation Research/Mutation Research Genomics","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"1997-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1016/S1383-5726(97)00009-5","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"20291425","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}

{"title":"Detection of a point mutation (A to G) in exon 5 of the murine Mgf gene defines a novel allele at the Steel locus with a weak phenotype1","authors":"Jochen Graw,&nbsp;Angelika Neuhäuser-Klaus,&nbsp;Walter Pretsch","doi":"10.1016/S1383-5726(97)00005-8","DOIUrl":"https://doi.org/10.1016/S1383-5726(97)00005-8","url":null,"abstract":"<div><p><span>A new mutation at the locus encoding the mast cell growth factor (Mgf) is described and designated as </span><em>Mgf^Sl-3Neu</em><span>. Homozygous mutants have a light grey fur, sometimes with white patches. Homozygotes are fertile, but with reduced litter size, when mated inter se. Analysis of haematological parameters indicated no difference between mutant and wild-type mice. Sequence analysis of the cDNA obtained from the brain of homozygous mutants revealed an A→G exchange at position 400 leading to a predicted amino acid exchange from Asn→Leu at position 122. As a consequence of the predicted amino acid exchange an extension of the α-helical context and a decreased hydropathicity of the region at positions 101–125 can be deduced. This single amino acid exchange is outside of the known important domains of MGF and explains the weak phenotype of </span><em>Mgf^Sl-3Neu</em>.</p></div>","PeriodicalId":100939,"journal":{"name":"Mutation Research/Mutation Research Genomics","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"1997-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1016/S1383-5726(97)00005-8","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"72111849","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}

{"title":"Identification and characterization of two polymorphic Ya5 Alu repeats","authors":"Santosh S Arcot ,&nbsp;Margaret M DeAngelis ,&nbsp;Stephen T Sherry ,&nbsp;Aaron W Adamson ,&nbsp;Jane E Lamerdin ,&nbsp;Prescott L Deininger ,&nbsp;Anthony V Carrano ,&nbsp;Mark A Batzer","doi":"10.1016/S1383-5726(97)00002-2","DOIUrl":"10.1016/S1383-5726(97)00002-2","url":null,"abstract":"<div><p><span>Two new polymorphic Alu elements (HS2.25 and HS4.14) belonging to the young (Ya5/8) subfamily of human-specific Alu repeats have been identified. </span>DNA sequence analysis<span> of both Alu repeats revealed that each Alu repeat had a long 3′-oligo-dA-rich tail (41 and 52 nucleotides in length) and a low level of random mutations. HS2.25 and HS4.14 were flanked by short precise direct repeats of 8 and 14 nucleotides in length, respectively. HS2.25 was located on human chromosome 13<span>, and HS4.14 on chromosome 1. Both Alu elements were absent from the orthologous positions within the genomes of non-human primates, and were highly polymorphic in a survey of twelve geographically diverse human groups.</span></span></p></div>","PeriodicalId":100939,"journal":{"name":"Mutation Research/Mutation Research Genomics","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"1997-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1016/S1383-5726(97)00002-2","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"20291420","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}