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Efficient screening of p53 mutations by denaturing gradient gel electrophoresis in colorectal tumors. 1993

R Hamelin, and N Jego, and P Laurent-Puig, and M Vidaud, and G Thomas
Laboratoire de Génétique des Tumeurs, Institut Curie, Paris, France.

Alterations in exons 5-8 of the p53 gene have been found in the vast majority of human tumors. Although some specific codons have been revealed as preferential mutational sites in defined tumor types, mutations are generally so scattered within this region that various screening methods for their detection have been widely used. However, the capacity of these techniques to detect exhaustively all mutations has not been evaluated. In this report, conditions for analysing exons 5-8 of the p53 gene by denaturing gradient gel electrophoresis (DGGE) have been elaborated using the Melt87 and SQHTX computer programs. The procedure requires five different amplifications. The screening of 90 colorectal tumors revealed 56 amplification products demonstrating abnormal DGGE behavior. Direct sequencing of these amplification products after asymmetric polymerase chain reaction (PCR) showed, besides polymorphism, 49 somatically mutated DNA samples (54.4%) corresponding to 38 different p53 variants. Moreover, 25 additional p53 variants identified in other tumor types were also detectable with our conditions. Thus, altogether the procedure has been successfully used in the detection of 63 different p53 variants. The experimental differences in migration between the wild-type homoduplex DNA molecule and the heteroduplex(es) containing one mismatch were systematically compared with those calculated by the SQHTX program. The computer program was found to be reliably predictive. This DGGE procedure appears thus to be effective and reliable for detecting mutations in exons 5-8 of the p53 gene.

UI MeSH Term Description Entries
D008969 Molecular Sequence Data Descriptions of specific amino acid, carbohydrate, or nucleotide sequences which have appeared in the published literature and/or are deposited in and maintained by databanks such as GENBANK, European Molecular Biology Laboratory (EMBL), National Biomedical Research Foundation (NBRF), or other sequence repositories. Sequence Data, Molecular,Molecular Sequencing Data,Data, Molecular Sequence,Data, Molecular Sequencing,Sequencing Data, Molecular
D004591 Electrophoresis, Polyacrylamide Gel Electrophoresis in which a polyacrylamide gel is used as the diffusion medium. Polyacrylamide Gel Electrophoresis,SDS-PAGE,Sodium Dodecyl Sulfate-PAGE,Gel Electrophoresis, Polyacrylamide,SDS PAGE,Sodium Dodecyl Sulfate PAGE,Sodium Dodecyl Sulfate-PAGEs
D005091 Exons The parts of a transcript of a split GENE remaining after the INTRONS are removed. They are spliced together to become a MESSENGER RNA or other functional RNA. Mini-Exon,Exon,Mini Exon,Mini-Exons
D006801 Humans Members of the species Homo sapiens. Homo sapiens,Man (Taxonomy),Human,Man, Modern,Modern Man
D001483 Base Sequence The sequence of PURINES and PYRIMIDINES in nucleic acids and polynucleotides. It is also called nucleotide sequence. DNA Sequence,Nucleotide Sequence,RNA Sequence,DNA Sequences,Base Sequences,Nucleotide Sequences,RNA Sequences,Sequence, Base,Sequence, DNA,Sequence, Nucleotide,Sequence, RNA,Sequences, Base,Sequences, DNA,Sequences, Nucleotide,Sequences, RNA
D015179 Colorectal Neoplasms Tumors or cancer of the COLON or the RECTUM or both. Risk factors for colorectal cancer include chronic ULCERATIVE COLITIS; FAMILIAL POLYPOSIS COLI; exposure to ASBESTOS; and irradiation of the CERVIX UTERI. Colorectal Cancer,Colorectal Carcinoma,Colorectal Tumors,Neoplasms, Colorectal,Cancer, Colorectal,Cancers, Colorectal,Carcinoma, Colorectal,Carcinomas, Colorectal,Colorectal Cancers,Colorectal Carcinomas,Colorectal Neoplasm,Colorectal Tumor,Neoplasm, Colorectal,Tumor, Colorectal,Tumors, Colorectal
D016133 Polymerase Chain Reaction In vitro method for producing large amounts of specific DNA or RNA fragments of defined length and sequence from small amounts of short oligonucleotide flanking sequences (primers). The essential steps include thermal denaturation of the double-stranded target molecules, annealing of the primers to their complementary sequences, and extension of the annealed primers by enzymatic synthesis with DNA polymerase. The reaction is efficient, specific, and extremely sensitive. Uses for the reaction include disease diagnosis, detection of difficult-to-isolate pathogens, mutation analysis, genetic testing, DNA sequencing, and analyzing evolutionary relationships. Anchored PCR,Inverse PCR,Nested PCR,PCR,Anchored Polymerase Chain Reaction,Inverse Polymerase Chain Reaction,Nested Polymerase Chain Reaction,PCR, Anchored,PCR, Inverse,PCR, Nested,Polymerase Chain Reactions,Reaction, Polymerase Chain,Reactions, Polymerase Chain
D016158 Genes, p53 Tumor suppressor genes located on the short arm of human chromosome 17 and coding for the phosphoprotein p53. Genes, TP53,TP53 Genes,p53 Genes,Gene, TP53,Gene, p53,TP53 Gene,p53 Gene
D017354 Point Mutation A mutation caused by the substitution of one nucleotide for another. This results in the DNA molecule having a change in a single base pair. Mutation, Point,Mutations, Point,Point Mutations

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