title: Efficient mutation detection in mismatch repair genes using a combination of single-strand conformational polymorphism and heteroduplex analysis at a controlled temperature.
abstract: Single-strand conformational polymorphism analysis (SSCP) and heteroduplex analysis (HD) were tested as methods for mutation screening with respect to experimental variation, sensitivity, and specificity. Thirty-nine fluorescently labeled PCR products covering the two mismatch repair genes, hMLH1 and hMSH2, were tested in 15 patients for pattern changes, using SSCP and HD at two temperatures, in a total of 2340 runs. SSCP was most efficient in detecting base changes, whereas HD was the method of choice when detecting deletions. SSCP and HD at 20 degrees C were most effective (sensitivity 97%, specificity 49%), and SSCP and HD at 10 degrees C gave no additional information, except in one case where an exon had two base changes. Several mutations only showed a small pattern change in one of the two strands, most explicit at 20 degrees C. No correlation between the type of base change and the size or direction of the pattern changes could be found.
authors.affiliation: University Hospital of Aarhus, Skejby Sygehus, Denmark. fpw@kba.sks.au.dk
authors.forename: F P
authors.initials: FP
authors.lastname: Wikman
authors.forename: N
authors.initials: N
authors.lastname: Katballe
authors.forename: M
authors.initials: M
authors.lastname: Christensen
authors.forename: S
authors.initials: S
authors.lastname: Laurberg
authors.forename: T F
authors.initials: TF
authors.lastname: Orntoft
mesh_terms: D048868:Adaptor Proteins, Signal Transducing; D020137:Base Pair Mismatch / Q000235:genetics*; D002352:Carrier Proteins; D003123:Colorectal Neoplasms, Hereditary Nonpolyposis / Q000175:diagnosis / Q000235:genetics*; D004252:DNA Mutational Analysis / Q000379:methods*; D004260:DNA Repair / Q000235:genetics*; D004268:DNA-Binding Proteins; D005069:Evaluation Studies as Topic; D006580:Genetic Carrier Screening; D005820:Genetic Testing / Q000379:methods; D020180:Heteroduplex Analysis / Q000191:economics / Q000379:methods*; D006801:Humans; D000070957:MutL Protein Homolog 1; D051718:MutS Homolog 2 Protein; D009363:Neoplasm Proteins / Q000235:genetics; D009687:Nuclear Proteins; D016133:Polymerase Chain Reaction; D011110:Polymorphism, Genetic / Q000235:genetics; D018807:Polymorphism, Single-Stranded Conformational; D011518:Proto-Oncogene Proteins / Q000235:genetics; D012680:Sensitivity and Specificity; D013696:Temperature
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; D004268:DNA-Binding Proteins; C501486:MLH1 protein, human; D009363:Neoplasm Proteins; D009687:Nuclear Proteins; D011518:Proto-Oncogene Proteins; C497172:MSH2 protein, human; D000070957:MutL Protein Homolog 1; D051718:MutS Homolog 2 Protein