荧光定量PCR检测结核杆菌PncA基因突变研究  

Detection of PncA Gene Mutation from Mycobacterium Tuberculos by Real-Time PCR

作　　者：杨小蓉[1] 陈少莲[1] 卢景辉[1] 丁彩屏[1] 

机构地区：[1]广东药学院附属第一医院检验科,广东广州510080

出　　处：《临床医学工程》2013年第12期1482-1485,共4页Clinical Medicine & Engineering

基　　金：广东省医学科研基金(2011321)

摘　　要：目的运用实时荧光定量PCR方法检测结核分支杆菌耐吡嗪酰胺(PZA)分离株PncA基因突变情况,从而探讨其水平与结核病的临床转归的相关性。方法用荧光定量PCR Taqman探针技术,以PncA基因片段设计引物,以突变发生率最高的47位(Thr→Ala)(PncA139)和85位(Leu→Pro)(PncA254)设计探针,10倍系列稀释含有目的基因的质粒,进行实时荧光定量PCR反应,建立标准曲线。并用于检测临床105份标本(TB-DNA>103)pncA基因突变表达量。结果①63例PncA139点突变表达量>102,占60%;②31例PncA254点突变表达量>102,占30%;③PncA139位突变表达量>103的患者同时也出现PncA254位突变表现,且TB-DNA表达量均>106;④PncA139位突变的高拷贝组与低拷贝组突变率比较χ2为50.44,85位点χ2为20.97,P值均<0.005,比较差异亦有明显统计意义。结论 TB-DNA高拷贝且PncA基因突变是临床结核病患者病情反复,以致迁延不愈的原因之一。Objective To explore the relation between PncA gene mutation resistant to pyrazinamide （PZA） to treat mycobacterium tuberculosis and the clinical outcome, and detect its expression level by real-time PCR. Methods According to mutation sequences of PncA, this standard curve was established by real-time PCR Taqman probe technology, then 105 cases were detected （TB-DNA 〉103）. Results 0 The gene expression levels were detected in 63 cases of 47 amino acid （PncA139） point mutation （60%）; （2）The gene expression levels were detected in 31 cases of 85 amino acid （PncA254） point mutation （30%）; （3）Patients with PncA gene mutation （47） level （〉103） also showed the mutation （85）, and TB-DNA 〉106; （4） PncA139 and PncA254 mutation of PncA between high copy group and low copy group was statistically different （x2 = 50.44 and 20.97, P 〈0.005）. Conclusions PncA gene mutation is one of the reasons for the difficulty of treating mycobactrium tuberculos.

关 键 词：实时荧光定量PCR 结核分支杆菌 吡嗪酰胺 PncA基因突变 

分 类 号：R378.911[医药卫生—病原生物学]