Simultaneous Detection of Three Genotypes of Gene Methylene Tetrahydrofolate Reductase and Methionine Synthase Reductase Based on Multiplex Asymmetric Real-Time PCR-HRM Biosensing

作者全名："Yu, Wen; Yao, Juan; Zhang, Zhang"

作者地址："[Yu, Wen] Chongqing Univ Canc Hosp, Dept Clin Lab, Chongqing Key Lab Translat Res Canc Metastasis & I, Chongqing 400030, Peoples R China; [Yu, Wen] Chongqing Canc Inst, Chongqing 400030, Peoples R China; [Yao, Juan] Southwest Med Univ, Dept Lab Med, Affiliated Tradit Chinese Med Hosp, Luzhou 646000, Peoples R China; [Zhang, Zhang] Southwest Med Univ, Dept Lab Med, Affiliated Hosp, Luzhou 646000, Peoples R China; [Zhang, Zhang] Chongqing Med Univ, Key Lab Lab Med Diagnost Educ, Dept Lab Med, Chongqing, Peoples R China"

通信作者："Zhang, Z (通讯作者)，Southwest Med Univ, Dept Lab Med, Affiliated Hosp, Luzhou 646000, Peoples R China.; Zhang, Z (通讯作者)，Chongqing Med Univ, Key Lab Lab Med Diagnost Educ, Dept Lab Med, Chongqing, Peoples R China."

来源：ANALYTICAL CHEMISTRY

ESI学科分类：CHEMISTRY

WOS号：WOS:000858449000001

JCR分区：Q1

影响因子：6.7

年份：2023

卷号：　

期号：　

开始页：　

结束页：　

文献类型：Article; Early Access

关键词：　

摘要："Genotyping of folate metabolism genes is of great importance in disease diagnosis and prevention. However, most current detection methods used for folate metabolism gene genotyping are based on sequencing and chips, which suffer from a high cost and laborious and time-consuming procedures. Herein, we reported a multiplex asymmetric PCR-HRM strategy for identifying genotypes of folate metabolism genes in a single tube. The proposed multiplex PCR-HRM assay has been successfully applied to identify the genotypes of folate metabolism genes, methylene tetrahydrofolate reductase (C677T, A1298C) and methionine synthase reductase A66G, on 1 mu L of genomic DNA (gDNA) samples directly released from blood specimens, and the genotyping results were 100% consistent with the results of sequencing. The assay allows us to accurately detect the genotypes of gDNA with the detection limit down to 1 ng, which meets the clinical requirement. What is more, the capacity of resistance to aerosol pollution of the multiplex asymmetric PCR-HRM biosensing was first addressed and has been evaluated as it can withstand contamination of roughly 12.5-25% interfering nucleic acids. Because of the advantages of multiplex detection, high accuracy, and resistance to aerosol pollution and having no open tube procedure, this approach would pave the way for establishing a fast and cost-effective platform for folate metabolism gene genotyping and other SNP genotyping in clinical diagnostics."

基金机构：China Postdoctoral Science Foundation [2021TQ0395]; National Natural Science Foundation of China [81902169]

基金资助正文：This research was supported by the China Postdoctoral Science Foundation (2021TQ0395) and the National Natural Science Foundation of China (no 81902169) .