中华预防医学杂志
中華預防醫學雜誌
중화예방의학잡지
CHINESE JOURNAL OF
2012年
5期
465-467
,共3页
卢昕%王淑京%刘莎%阚飙%逄波
盧昕%王淑京%劉莎%闞飆%逄波
로흔%왕숙경%류사%감표%방파
弧菌%副溶血性%核酸扩增技术%环介导等温扩增%toxR基因
弧菌%副溶血性%覈痠擴增技術%環介導等溫擴增%toxR基因
호균%부용혈성%핵산확증기술%배개도등온확증%toxR기인
Vibrio parahaemolyticus%Nucleic acid amplification techniques%Loop-mediated isothermal amplification%toxR gene
目的 建立基于环介导等温扩增(LAMP)技术的副溶血弧菌快速检测方法.方法 针对副溶血弧菌toxR基因序列设计一套LAMP引物,应用LAMP技术对33株副溶血弧菌、22株其他种属细菌及含不同副溶血弧菌参考菌株( ATCC 17802)基因组拷贝数(5×100 ～5×105拷贝/μl)的样品进行检测,对平行样品分别应用普通PCR法和TaqMan探针实时PCR法进行检测,对3种检测方法的特异度、灵敏度及检测下限及反应时间进行比较.结果 LAMP技术、普通PCR法、TaqMan探针实时PCR法检测副溶血弧菌的特异度、灵敏度均为100％(分别为22/22,33/33),检测下限分别为5×101、5×103、5×102拷贝/μl,反应所需时长分别为22 main、3h、50 min.结论 与普通PCR、TaqMan 探针实时PCR检测相比,LAMP技术检测下限低,检测时长短,适于对副溶血弧菌进行快速检测.
目的 建立基于環介導等溫擴增(LAMP)技術的副溶血弧菌快速檢測方法.方法 針對副溶血弧菌toxR基因序列設計一套LAMP引物,應用LAMP技術對33株副溶血弧菌、22株其他種屬細菌及含不同副溶血弧菌參攷菌株( ATCC 17802)基因組拷貝數(5×100 ～5×105拷貝/μl)的樣品進行檢測,對平行樣品分彆應用普通PCR法和TaqMan探針實時PCR法進行檢測,對3種檢測方法的特異度、靈敏度及檢測下限及反應時間進行比較.結果 LAMP技術、普通PCR法、TaqMan探針實時PCR法檢測副溶血弧菌的特異度、靈敏度均為100％(分彆為22/22,33/33),檢測下限分彆為5×101、5×103、5×102拷貝/μl,反應所需時長分彆為22 main、3h、50 min.結論 與普通PCR、TaqMan 探針實時PCR檢測相比,LAMP技術檢測下限低,檢測時長短,適于對副溶血弧菌進行快速檢測.
목적 건립기우배개도등온확증(LAMP)기술적부용혈호균쾌속검측방법.방법 침대부용혈호균toxR기인서렬설계일투LAMP인물,응용LAMP기술대33주부용혈호균、22주기타충속세균급함불동부용혈호균삼고균주( ATCC 17802)기인조고패수(5×100 ～5×105고패/μl)적양품진행검측,대평행양품분별응용보통PCR법화TaqMan탐침실시PCR법진행검측,대3충검측방법적특이도、령민도급검측하한급반응시간진행비교.결과 LAMP기술、보통PCR법、TaqMan탐침실시PCR법검측부용혈호균적특이도、령민도균위100％(분별위22/22,33/33),검측하한분별위5×101、5×103、5×102고패/μl,반응소수시장분별위22 main、3h、50 min.결론 여보통PCR、TaqMan 탐침실시PCR검측상비,LAMP기술검측하한저,검측시장단,괄우대부용혈호균진행쾌속검측.
Objective This study aimed to establish a loop-mediated isothermal amplification (LAMP) assay for detection of Vibrio parahaenolyticus( V.parahaemolyticus ).Methods The specificity of this assay was evaluated by using a panel of 33 strains of V.parahaemolyticcus and 22 strains of other species bacteria.The sensitivity was determined by using serial dilutions of V.parahaemolyticas ( ATCC 17802 ) chromosomal DNA (5 × 100 -5 × 105 copies/μl).The samples were also tested by using qualification PCR assay and Taqman real-time PCR assay in parallel for comparison with LAMP.Results Both sensitivity and specificity of LAMP assay,PCR assay and Taqman real-time PCR assay were 100％ (22/22,33/33,respectively).The detection limits of above three methods assay were 5 × 101 copies/μl,5 × 103 copies/μl and 5 × 102 copies/μl,respectively.The reaction period of time needed of the above three assays was 22 min,3 h,50 main,respectively.Conclusion Compared to qualification PCR assay and Taqman real-time PCR assay,the established LAMP assay was better in low detection limit and less reaction time,which made it an ideal method for quick detection of V.parahaemolyticus.
