Optimization of restriction fragment length polymorphism and single strand conformation polymorphism analysis for the detection of point mutations of ras oncogenes in paraffin-embedded tissues.
69 samples of formalin-fixed and paraffin-embedded tumor tissues (colon carcinomas n = 15, colon adenomas n = 15, transitional cell carcinomas of the urinary bladder n = 17, transitional cell papillomas of the urinary bladder n = 22) were investigated for point mutations of ras oncogenes using polymerase chain reaction (PCR) and nested-PCR followed by restriction fragment length polymorphism (RFLP) and single strand conformation polymorphism (SSCP), respectively. Results were proved by sequence analysis. Nested-PCR combined with SSCP analysis revealed five mutations of the Ki-ras oncogenes in five colon tumors. RFLP analysis revealed one point mutation in one sample of invasive transitional cell carcinomas. Using optimized PCR protocols, the methods described are applicable to paraffin- embedded tissues and offer a tool for mutation analysis in retrospective and prospective studies.
Weise
B
B
Department of Pathology, University of Göttingen, Germany.
Ghebremedhin
E
E
Zschunke
F
F
Radzun
H J
HJ
D001483:Base Sequence; D003110:Colonic Neoplasms / Q000235:genetics / Q000473:pathology; D011905:Genes, ras; D006801:Humans; D008969:Molecular Sequence Data; D016612:Paraffin Embedding; D017354:Point Mutation; D016133:Polymerase Chain Reaction; D012150:Polymorphism, Restriction Fragment Length; D018807:Polymorphism, Single-Stranded Conformational; D017422:Sequence Analysis, DNA; D001749:Urinary Bladder Neoplasms / Q000235:genetics / Q000473:pathology