Simultaneous Genotyping of Multiple Somatic Mutations by Using a Clamping PNA and PNA Detection Probes

• Published in: Chembiochem : a European journal of chemical biology Vol. 16; no. 2; pp. 209 - 213
• Main Authors: Kim, Yong-Tae, Kim, Jin Woo, Kim, Sung Kee, Joe, Goon Ho, Hong, In Seok
• Format: Journal Article
• Language: English
• Published: Weinheim WILEY-VCH Verlag 19.01.2015; WILEY‐VCH Verlag; Wiley Subscription Services, Inc
• Subjects: Alanine - chemistry; cancer; Cell Line; Dimerization; gamma-PNA; Genes, ras - genetics; Genotyping Techniques - methods; HeLa Cells; Humans; Lysine - chemistry; Molecular Probes; multiple genotyping; Mutation; peptide nucleic acid; Peptide Nucleic Acids - genetics; Polymerase Chain Reaction - methods; quantitative PCR; somatic mutations; Temperature; somatic mutations; cancer; gamma-PNA; multiple genotyping; quantitative PCR; peptide nucleic acid
• ISSN: 1439-4227; 1439-7633
• DOI: 10.1002/cbic.201402640

It has been very difficult to detect and quantify multiple somatic mutations simultaneously in single‐tube qPCR. Here, a novel method for simultaneous detection of multiple mutations and a melting curve analysis was developed by using clamping PNA and detection PNA probes. Each PNA probe was designed to have a specific melting temperature by the introduction of γ‐PNA monomer. This technique was successfully applied to the detection of six genotypes in two different mutations of K‐RAS at the same time. Such simultaneous analysis of an amplified curve and a melting curve in qPCR can be widely used for the early diagnosis of cancer and determining the prognosis of drug treatments. A novel method of simultaneous detection of multiple mutations and a melting curve analysis was developed using clamping PNA/detection PNA probes. Each PNA probe was designed to have a specific melting temperature by the introduction of γ‐PNA monomers. This technique was successfully applied to the detection of six genotypes in two different mutations of K‐RAS at the same time.