title: An effective method based on real time fluorescence quenching for single nucleotide polymorphism detection.
abstract: In the Human Genome Project, the most common type of these variations is single nucleotide polymorphisms (SNPs). A large number of different SNP typing technologies have been developed in recent years. Enhancement and innovation for genotyping technologies are currently in progress. We described a rapid and effective method based on real time fluorescence quenching for SNP detection. The new method, Quenching-PCR, offering a single base extension method fully integrated with PCR which used a probe with quencher to eliminate fluorophor of the terminal base according to dideoxy sequencing method. In this platform, dideoxy sequencing reaction and obtaining values of real-time fluorescence occur simultaneously. The assay was validated by 106 DNA templates comparing with Sanger's sequencing and TaqMan assay. Compared with the results of DNA sequencing, the results of Quenching-PCR showed a high concordance rate of 93.40%, while the results of TaqMan platform showed a concordance rate of 92.45%, indicating that Quenching PCR and TaqMan assay were similar in accuracy. Therefore, Quenching PCR will be easily applicable and greatly accelerate the role of SNP detection in physiological processes of human health.
authors.affiliation: State Key Laboratory of Bioreactor Engineering & Shanghai Key Laboratory of Chemical Biology, School of Pharmacy, East China University of Science and Technology, Shanghai 200237, China.
authors.forename: Yichun
authors.initials: Y
authors.lastname: Xu
authors.affiliation: Department of General Surgery, Zhujiang Hospital, Southern Medical University, Guangzhou, 510282, China.
authors.forename: Shuai
authors.initials: S
authors.lastname: Han
authors.affiliation: Shanghai Aoyin Biotechnology Research and Development Limited Corporation, Shanghai 201203, China.
authors.forename: Xinhua
authors.initials: X
authors.lastname: Huang
authors.affiliation: Department of Pathology, Central Hospital of Xuzhou, Xuzhou 221009, China.
authors.forename: Shichao
authors.initials: S
authors.lastname: Zhuo
authors.affiliation: Shanghai Aoyin Biotechnology Research and Development Limited Corporation, Shanghai 201203, China.
authors.forename: Huiqing
authors.initials: H
authors.lastname: Dai
authors.affiliation: Laboratory of Integrative Medicine Surgery, Shuguang Hospital affiliated to Shanghai University of Traditional Chinese Medicine, 201203 Shanghai, China. Electronic address: wangke8430@163.com.
authors.forename: Ke
authors.initials: K
authors.lastname: Wang
authors.affiliation: Department of General Surgery, Zhujiang Hospital, Southern Medical University, Guangzhou, 510282, China. Electronic address: Leezhou888@126.com.
authors.forename: Zhou
authors.initials: Z
authors.lastname: Li
authors.affiliation: State Key Laboratory of Bioreactor Engineering & Shanghai Key Laboratory of Chemical Biology, School of Pharmacy, East China University of Science and Technology, Shanghai 200237, China. Electronic address: liujian@ecust.edu.cn.
authors.forename: Jianwen
authors.initials: J
authors.lastname: Liu
mesh_terms: D004247:DNA / Q000032:analysis / Q000737:chemistry / Q000235:genetics; D004847:Epithelial Cells / Q000737:chemistry; D006801:Humans; D009061:Mouth Mucosa / Q000166:cytology; D016133:Polymerase Chain Reaction / Q000379:methods*; D020641:Polymorphism, Single Nucleotide / Q000235:genetics*; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA / Q000379:methods*; D013050:Spectrometry, Fluorescence / Q000379:methods*
chemical_list: D004247:DNA
keywords: Fluorescence quenching; PCR; Sensitivity; Single nucleotide polymorphisms (SNPs)