title: Detection of rare point mutation via allele-specific amplification in emulsion PCR.
abstract: It is essential to analyze rare mutations in many fields of biomedical research. However, the detection of rare mutations is usually failed due to the interference of predominant wild-type DNA surrounded. Herein we describe a sensitive and facile method of detecting rare point mutation on the basis of allele-specific amplification in emulsion PCR. The identification and selective amplification of rare mutation are accomplished in one-pot reaction. The allele-specific primers coupled on magnetic beads allow the exclusive amplification and enrichment of the mutant amplicons. The productive beads bearing mutant amplicons are subsequently stained with the fluorescent dyes. Thus, the rare point mutations with a percentage as low as 0.1%, can be detected by fluorescent analysis. The relative percentages of mutation among different samples can be roughly accessed by counting the fraction of fluorescent positive beads through flow cytometry.
authors.affiliation: National Engineering Center for Biochip at Shanghai, Shanghai 201203, China. cmcheng2005@gmail.com
authors.forename: Changming
authors.initials: C
authors.lastname: Cheng
authors.forename: Yin
authors.initials: Y
authors.lastname: Zhou
authors.forename: Chao
authors.initials: C
authors.lastname: Yang
authors.forename: Juan
authors.initials: J
authors.lastname: Chen
authors.forename: Jie
authors.initials: J
authors.lastname: Wang
authors.forename: Jie
authors.initials: J
authors.lastname: Zhang
authors.forename: Guoping
authors.initials: G
authors.lastname: Zhao
mesh_terms: D000483:Alleles; D045744:Cell Line, Tumor; D005434:Flow Cytometry; D005453:Fluorescence; D005784:Gene Amplification; D056945:Hep G2 Cells; D006801:Humans; D057230:Limit of Detection; D017354:Point Mutation / Q000235:genetics*; D016133:Polymerase Chain Reaction; D020641:Polymorphism, Single Nucleotide; D035583:Rare Diseases / Q000175:diagnosis / Q000235:genetics