title: [Comparison of real-time PCR-optimized oligonucleotide probe method and Sanger sequencing for detection of KRAS mutations in colorectal and lung carcinomas].
abstract: 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%.
  
  
  CONCLUSIONS
  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.
authors.affiliation: Department of Pathology, Chinese Academy of Medical Sciences, Beijing, China.
authors.forename: Tian
authors.initials: T
authors.lastname: Qiu
authors.forename: Yun
authors.initials: Y
authors.lastname: Ling
authors.forename: Zhao
authors.initials: Z
authors.lastname: Chen
authors.forename: Ling
authors.initials: L
authors.lastname: Shan
authors.forename: Lei
authors.initials: L
authors.lastname: Guo
authors.forename: Ning
authors.initials: N
authors.lastname: Lü
authors.forename: Jian-ming
authors.initials: JM
authors.lastname: Ying
mesh_terms: D003062:Codon; D015179:Colorectal Neoplasms / Q000235:genetics*; D004252:DNA Mutational Analysis / Q000379:methods*; D005260:Female; D006801:Humans; D008175:Lung Neoplasms / Q000235:genetics*; D008297:Male; D008875:Middle Aged; D009154:Mutation; D015345:Oligonucleotide Probes; D011518:Proto-Oncogene Proteins / Q000235:genetics*; D016283:Proto-Oncogene Proteins p21(ras); D060888:Real-Time Polymerase Chain Reaction / Q000379:methods; D012680:Sensitivity and Specificity; D018631:ras Proteins / Q000235:genetics*
chemical_list: D003062:Codon; C117307:KRAS protein, human; D015345:Oligonucleotide Probes; D011518:Proto-Oncogene Proteins; D016283:Proto-Oncogene Proteins p21(ras); D018631:ras Proteins