A novel RFLP-ARMS TaqMan PCR-based method for detecting the BRAF V600E mutation in melanoma

作者：Zhang, Yunqing; Qu, Shoufang; Zhao, Jinyin; Yu, Ting; Guo, Liping; Yin, Songchao; Hu, Xiaoxu; Chen, Weijun; Lai, Wei^*; Huang, Jie^*

来源：Oncology Letters, 2018, 16(2): 1615-1621.

DOI：10.3892/ol.2018.8844

• B-Raf proto-oncogene
• serine
• threonine kinase
• V600E
• restriction fragment length polymorphism-amplification refractory mutation system TaqMan polymerase chain reaction
• melanoma

To enable the rapid and sensitive screening of the BRAF V600E mutation in clinical samples, a novel method combining restriction fragment length polymorphism (RFLP) analysis with the popular amplification refractory mutation system (ARMS) TaqMan quantitative (qPCR) genotyping method in a single reaction tube was developed. A total of 2 primer pairs were designed to enrich for and genotype the BRAF mutational hotspot (RFLP primers and ARMS primers) and a restriction enzyme was used to remove the wild-type alleles. The analysis revealed that this method detected mutant alleles in mixed samples containing >0.1% mutant sequences. In a survey of 53 melanoma samples, this method detected 21 mutation-positive samples. This novel RFLP-ARMS TaqMan qPCR protocol may prove useful for detecting mutations in clinical samples containing only a small proportion of mutant alleles.

• 出版日期2018-8
• 单位中国科学院北京基因组研究所; 中山大学