淋病奈瑟菌隐蔽性质粒的克隆及PCR的建立与应用  

CLONING OF THE CRYPTIC PLASMID FROM NEISSERIA GONORRHOEAE AND THE CLINICAL APPLICATION WITH PCR

作　　者：叶芳耘[1] 蔡龙荣[1] 施少林 何建文[1] 辛宏[1] 于洋[1] 晏碧君 

机构地区：[1]上海长海医院实验诊断科

出　　处：《临床检验杂志》1995年第2期71-73,共3页Chinese Journal of Clinical Laboratory Science

摘　　要：用分子生物学方法自淋病奈瑟菌标准分离株中克隆了４．２ｋｂ的隐蔽性质粒，并根据该质粒ＤＮＡ序列，设计引物，建立淋病奈瑟菌ＰＣＲ检测方法。该方法标本处理简单、反应程序优化，可在２．５小时内完成一次检测。对２１株淋病奈瑟菌标本进行检测，结果全为阳性；而对２２株其它菌株检测均为阴性，证实了该法的特异性。与涂片、培养法比较，ＰＣＲ阳性率最高。结合临床检测了７０６例性病患者，总阳性率为２８．６％（２０２／７０６）。实验表明该ＰＣＲ方法简便快速、敏感特异，对淋病的流行病学调查和早期防治有重要意义。所克隆的含有４．２ｋｂ隐蔽性质粒ＤＮＡ还可用作制备探针和ＰＣＲ阳性对照。The 4.2 kb cryptic plasmid was isolated from the standard reference strain of N.gonorrhoeae and purified by CsCl-ultracentrifugation.The 4.2 kb plasmid DNA was cloned into pUC 18 DNA Hind Ⅲ-site and then transformed into the E.coli JM103.A specific fragment of the cryptic plasmid was amplified by polymerase chain reaction to detect gonococcus.The cloned cryptic plasmid DNA from N.gonorrhoeae could be used for DNA probe as well the PCR positive controls.21 isolates of N.gonorrhoeae were positive by PCR, while the other 22 isolates of non-gonococcus were negative,It demonstrated that the PCR assay has a exquisite specificity.The samples collected from 69 high-risk patients were detected by three methods,microscope smear. cultured and PCR.The positve rate in PCR was 60.8%,much higher than that of microscope smear(26.08%) and cultured(18.84%).We examined 706 clinical specimens by PCR.the positive rate was 28.6%(202/706).Results show that the PCR is a simple,rapid,sensitive and specific method and is suited for routine assay.

关 键 词：淋病奈瑟菌 质粒 克隆 聚合酶链反应 

分 类 号：R378.16[医药卫生—病原生物学]