金黄色葡萄球菌特异性PCR检测靶点的自动化筛选  

Screening of specific target sequences for the PCR detection of Staphylococcus aureus by automatic genomic comparison

作　　者：范一灵[1,2] 朱东升[1,3] 胡瑜[1] 史贤明[1] 

机构地区：[1]上海交通大学农业与生物学院陆伯勋食品安全研究中心中美食品安全联合研究中心,上海200240 [2]上海市食品药品检验所,上海201203 [3]联合利华(上海)研发中心联合利华中国研究所,上海200335

出　　处：《生物工程学报》2011年第4期637-644,共8页Chinese Journal of Biotechnology

基　　金：国家科技支撑计划(No.2009BAK43B31);上海市科委项目(Nos.10142201300;08142200700;08391911000)资助~~

摘　　要：旨在挖掘用于鉴定金黄色葡萄球菌的高特异性靶点及其PCR检测引物。采用C++语言编程,以金黄色葡萄球菌Staphylococcus aureus MRSA 252基因组编码序列为对象,对2 656个可编码区进行分析,获得特异性靶点序列,并设计PCR扩增引物。对包括葡萄球菌属11个种及其他细菌属在内的共计137株细菌验证引物特异性,筛选获得9个DNA序列,并设计了4对引物。经验证2对引物的特异性较好,其中引物SA3的基因组DNA检测限为13.7 fg/μL,菌体检测限为9.25×102 CFU/mL。结果验证了特异性DNA靶点筛选平台的实用性,该方法突破了传统特异性靶点挖掘方法对检测对象的限制,适用性广,可移植性强。The aim of this study was to establish a fast and accurate method for developing specific DNA sequences and PCR primers for the detection of Staphylococcus aureus.An automatic C＋＋ program for genomic comparison was used to identify specific DNA sequences from the genome of S.aureus MRSA 252.Four primer pairs were obtained from 9 specific target sequences by comparison of 2656 coding sequences with our local genome database,and 2 pairs of primers were confirmed to be specific to S.aureus by PCR evaluation against 137 bacterial strains,including 11 species of Staphylococcus.Furthermore,the DNA detection sensitivity of primer SA3 was 13.7 fg/μL and the cell sensitivity for this primer was 9.25×102 CFU/mL.This method has overcome the limitations of specific target mining in conventional assays,and it could be easily and widely used for other foodborne pathogens.

关 键 词：金黄色葡萄球菌 基因组 生物信息比对分析 编码序列 C++编程 

分 类 号：Q93[生物学—微生物学]