Mutation detection by denaturing gradient gel electrophoresis (DGGE)

Fodde, Riccardo; Losekoot, Monique

doi:10.1002/humu.1380030202

Cited by 183 publications

(86 citation statements)

References 80 publications

(67 reference statements)

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“…The low rate of mutations detected in our cohort cannot be explained by relative low sensitivity of the method because the DGGE technique is a highly sensitive method and can detect > 90% of small base sequence alterations, such as splice site, missense, non-sense and frameshift mutations (Fodde and Losekoot, 1994;Myers et al, 1987). Nevertheless, large deletions and mutations in the region that the primers hybridize may not be detected by this technique.…”

Section: Discussionmentioning

confidence: 74%

Microsatellite instability and mismatch repair gene inactivation in sporadic pancreatic and colon tumours

et al. 1999

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Summary Genomic instability has been proposed as a new mechanism of carcinogenesis involved in hereditary non-polyposis colorectal cancer (HNPCC) and in a large number of sporadic cancers like pancreatic and colon tumours. Mutations in human mismatch repair genes have been found in HNPCC patients, but their involvement in sporadic cancer has not been clarified yet. In this study we screened 21 pancreatic and 23 colorectal sporadic cancers for microsatellite instability by ten and six different microsatellite markers respectively. Microsatellite alterations were observed at one or more loci in 66.6% (14/21) of pancreatic cancers and in 26% (6/23) colon tumours, but all the pancreatic and half of the colon samples showed a low rate of microsatellite instability. All the unstable samples were further analysed for mutations in the hMLH1 and hMSH2 genes and for hypermethylation of the hMLH1 promoter region. Alterations in the hMLH1 gene were found only in colorectal tumours with a large presence of microsatellite instability. None of the pancreatic tumours showed any alteration in the two genes analysed. Our results demonstrate that microsatellite instability is unlikely to play a role in the tumorigenesis of sporadic pancreatic cancers and confirm the presence of mismatch repair gene alterations only in sporadic colon tumours with a highly unstable phenotype.

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Section: Discussionmentioning

confidence: 74%

Microsatellite instability and mismatch repair gene inactivation in sporadic pancreatic and colon tumours

et al. 1999

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“…To analyse and characterize such extremely variable populations, there are a number of techniques that allow the estimation of genetic variability, such as heteroduplex mobility assays (Woodward et al, 1994), single-strand conformation polymorphism assays (Spinardi et al, 1991), multiple-site-specific tracking assays (Resch et al, 2001), mutant analysis by PCR and restriction enzyme cleavage (Chumakov et al, 1991) or denaturing gradient gel electrophoresis (Fodde & Losekoot, 1994;Woodward et al, 1994). However, to analyse many of the properties of such populations, it is necessary to know the nucleotide sequence of the constituting genomes.…”

Section: Introductionmentioning

confidence: 99%

Sampling and repeatability in the evaluation of hepatitis C virus genetic variability

Torres-Puente

Bracho

Jiménez

et al. 2003

Journal of General Virology

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Among the experimental techniques available to study the genetic variability of RNA virus populations, the most informative involve reverse transcription (RT), amplification, cloning and sequencing. The effects of several aspects of these techniques on the estimation of genetic variability in a virus population were analysed. Hepatitis C virus populations from four patients were examined. For each patient, ten series of data derived from independent PCR amplifications of a single RT reaction were obtained. The sample size of each data set was 10 sequences (in nine series) and 100 sequences (in one series). An additional data set derived from an independent RT reaction (about 10 sequences) performed on RNA extracted from the same serum sample was also analysed. The availability of data sets of different sample sizes allowed the effect of sample size on the amount and nature of the genetic variability recovered to be examined. The repeatability of the data obtained in different amplification experiments as well as from different RT reactions was also determined, together with the best strategy to obtain a given number of sequences by comparing the set of 100 sequences obtained from a single amplification with those obtained by pooling the nine sets of 10 sequences. In all cases, these results confirm the high repeatability of the conclusions and parameters derived from the sets of 10 sequences. These results validate the use of relatively small sample sets for the evaluation of genetic variability and for the estimation of phylogenetic relationships of RNA viruses in population and epidemiological studies.

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“…These include denaturing HPLC [21], mass spectrometry [22][23][24], DNA microarrays [25], chemical or enzymatic cleavage of mismatched duplexes [26,27], denaturing gel electrophoresis [28], sequence-specific conformation polymorphism [29], and techniques based on bacterial mismatch repair systems [30].…”

Section: Re-sequencingmentioning

confidence: 99%

Genome Projects and the Functional-Genomic Era

Sauer

Konthur

Lehrach

2005

CCHTS

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The problems we face today in public health as a result of the -fortunately -increasing age of people and the requirements of developing countries create an urgent need for new and innovative approaches in medicine and in agronomics. Genomic and functional genomic approaches have a great potential to at least partially solve these problems in the future. Important progress has been made by procedures to decode genomic information of humans, but also of other key organisms. The basic comprehension of genomic information (and its transfer) should now give us the possibility to pursue the next important step in life science eventually leading to a basic understanding of biological information flow; the elucidation of the function of all genes and correlative products encoded in the genome, as well as the discovery of their interactions in a molecular context and the response to environmental factors. As a result of the sequencing projects, we are now able to ask important questions about sequence variation and can start to comprehensively study the function of expressed genes on different levels such as RNA, protein or the cell in a systematic context including underlying networks. In this article we review and comment on current trends in large-scale systematic biological research. A particular emphasis is put on technology developments that can provide means to accomplish the tasks of future lines of functional genomics.

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Mutation detection by denaturing gradient gel electrophoresis (DGGE)

Cited by 183 publications

References 80 publications

Microsatellite instability and mismatch repair gene inactivation in sporadic pancreatic and colon tumours

Microsatellite instability and mismatch repair gene inactivation in sporadic pancreatic and colon tumours

Sampling and repeatability in the evaluation of hepatitis C virus genetic variability

Genome Projects and the Functional-Genomic Era

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