[Comparison of real-time PCR-optimized oligonucleotide probe method and Sanger sequencing for detection of KRAS mutations in colorectal and lung carcinomas].

OBJECTIVE: To investigate the feasibility of real-time PCR-optimized oligonucleotide probe method for detection of KRAS mutations in lung and colorectal carcinomas, as compared with Sanger sequencing method.

METHODS: Genomic DNA was extracted from formalin fixed paraffin embedded samples of 221 lung carcinomas and 131 colorectal carcinomas after tumor cell content assessment and macrodissection. Real-time PCR-optimized oligonucleotide probe method and Sanger sequencing were performed to detect KRAS gene mutations. The frequency of KRAS mutation, mutation types, and their concordance were analyzed.

RESULTS: KRAS mutation was detected in 6.3% (14/221) and 4.5% (10/221) of 221 lung cancer samples by using real-time PCR-optimized oligonucleotide probe method and Sanger sequencing, respectively, while in 41.2% (54/131) and 40.5% (53/131) of 131 colorectal cancer samples, respectively. There was no significant correlation between KRAS gene mutations and patients' gender and age (P > 0.05). The positive rate of KRAS codon 12 mutation was significantly higher than that of KRAS codon 13 mutation (P < 0.05). The overall concordance between real-time PCR-optimized oligonucleotide probe method and Sanger sequencing for KRAS mutation detection was 97.4%.

CONCLUSION: Real-time PCR-optimized oligonucleotide probe method provides an alternative method with high consistency and sensitivity as compared to Sanger sequencing in gene mutation detection.