CAJ | 학술논문

目的建立环介导等温扩增(LAMP)快速检测肺炎链球菌的方法.方法根据美国国家生物信息中心(GenBank)上提交的肺炎链球菌R6株的lytA基因序列(登录号AF2)08540)设计特异LAMP引物,以盐酸胍一苯甲醇法提取阳性血培养瓶中细菌DNA,然后以LAMP技术扩增lytA基因,反应条件为63℃45 min,采用肉眼观察浊度和琼脂糖电泳的方法来观察结果.结果在196份阳性血培养瓶中,采用LAMP法检测到肺炎链球菌16株,与传统方法比较,其检测肺炎链球菌的灵敏度和特异度均达到100%,且检测时间缩短为1 h左右.模拟标本的检测结果显示该方法的最低检测限为102CFU/ml.结论该方法灵敏度高,特异性强,操作方便快速,适合从临床标本中检测肺炎链球菌.
목적 건립배개도등온확증(LAMP)쾌속검측폐염련구균적방법.방법 근거미국국가생물신식중심(GenBank)상제교적폐염련구균R6주적lytA기인서렬(등록호AF2)08540)설계특이LAMP인물,이염산고일분갑순법제취양성혈배양병중세균DNA,연후이LAMP기술확증lytA기인,반응조건위63℃45 min,채용육안관찰탁도화경지당전영적방법래관찰결과.결과 재196빈양성혈배양병중,채용LAMP법검측도폐염련구균16주,여전통방법비교,기검측폐염련구균적령민도화특이도균체도100%,차검측시간축단위1 h좌우.모의표본적검측결과현시해방법적최저검측한위102CFU/ml.결론 해방법령민도고,특이성강,조작방편쾌속,괄합종림상표본중검측폐염련구균.
Objective To establish loop-mediated isothermal amplification(LAMP)method for detecting Streptococcus pneumoniae in clinical samples.Methods Four Streptococcus pneumoniae-specific LAMP primers were designed according to the published sequence of strain R6(GenBank accession number AE008540).Genomie DNA in positive blood cultures was extracted by nanidine hydrochloride and benzene-methanol.Then lytA was amplified by LAMP at 63℃ for 45 minutes.We observed the turbidity in thereaction tube.For further confirmation.The amplified products were also detected using electrophoresis in 2% agarose gels,followed by ethidium bromide staining.Resuits 16 strains of Streptococcus pneumoniae were detected in 196 positive blood cuhure bottles by LAMP.Compared with traditional method.It8 sensitivity and specificity were both 100%and the detection could be finished in an hour.The assay had a minimum detection limit of 102 CFU/m1.Conclusions This IJAMP-based assay is simple,rapid.Sensitive and specific.It can be used to detect trept OCOCCUS pneumoniae in clinical samples.