GeXP系统在恶性疟耐青蒿素基因K13SNP检测中的应用  

A GeXP system combined with multiplex PCR technology for detection of Plasmodium falciparum

作　　者：石莹[1] 樊学军[1] 田绿波[1] 龙娟[2] SHI Ying;FAN Xue-jun;TIAN Lv-bo;LONG Juan(Chengdu Customs Sichuan International Trav-elHealthcare Center,Chengdu,China 610041;Department o fLaboratory Medicine,Sichuan Academy of Medical Sciences&Sichuan Provincial People's Hospital)

机构地区：[1]成都海关四川国际旅行卫生保健中心,四川成都610041 [2]四川省人民医院检验科

出　　处：《中国病原生物学杂志》2020年第3期249-252,258,共5页Journal of Pathogen Biology

基　　金：四川省科技项目(No.2017RZ0053,No.2018jy0339)。

摘　　要：目的探讨GeXP系统在恶性疟耐青蒿素基因K13SNP检测中的应用价值。方法根据4种SNP的序列特征制备标准品质粒。收集2016-2018年出入境人群中检出的28例临床标本,利用GenBank下载4种SNP的核苷酸序列,利用GeXPexpressProfiler设计特异性引物,优化条件采用GeXP系统结合多重PCR检测对应SNP及28份临床标本。结果优化的引物能特异性检测4种K13SNP位点,无交叉反应,灵敏度为100%,特异性为99.81%,检测效能为99.15%,检出限为5×103copies/μl,应用于临床标本检测,结果与测序法的一致性为99.32%。结论GeXP方法检测恶性疟耐青蒿素基因4种K13SNP位点快速、准确可用于输入性恶性疟的检测。Objective To use the GenomeLab Genetic Analysis System(GeXP)and multiplex-PCR to establish a high-throughput,rapid,and accurate method for detection of P.falciparum.Methods K13 is a common artemisinin resistance gene.Standard granules were prepared based on the sequence characteristics of four K13 SNPs.The sequences of these SNPs were obtained from the NCBI’s GenBank,and gene-specific primer pairs were designed using GeXP express Profiler.Optimized conditions allowed the GeXP system to detect the corresponding SNPs in a specific and rapid manner.Twenty-eight clinical specimens were collected at the International Travel Healthcare Center from 2016 to 2018 and tested with the GeXP system and multiplex PCR.The sensitivity and specificity of this assay were evaluated.Results The optimized primers specifically detected four K13 SNP loci without cross-reactivity.The assay had a sensitivity of 100%,a specificity of 99.81%,and an efficiency of 99.15%;its detection limit was 5×103 copies/μl.Its consistency with gene sequencing was 99.32%.Conclusion The combined GeXP and multiplex-PCR method quickly and accurately detected four SNP loci of K13,an artemisinin resistance gene,in P.falciparum.This facilitates the detection of imported P.falciparum.

关 键 词：恶性疟 耐青蒿素 K13 GeXP 多重PCR 

分 类 号：R382.31[医药卫生—医学寄生虫学]