通用引物PCR检测临床常见致病菌的实验研究  

Universal Primer PCR for Detection of Clinic Bacteria

作　　者：王建春[1] 刘静[1] 黄文红 张秀岭 叶林柏[1] 梅国华 

机构地区：[1]武汉大学生命科学学院,武汉430072 [2]武汉钢铁公司职工医院,武汉430080

出　　处：《微生物学通报》2005年第2期73-77,共5页Microbiology China

摘　　要：通用引物可一次性扩增18种临床常见致病菌和耐药菌株的DNA,扩增片段长度在220bp左右,18种特异性探针分别与18种标准菌株的PCR扩增产物杂交结果显示探针都具有高度特异性;5种37例经法国梅里埃API细菌鉴定系统确定的临床分离菌株进行杂交鉴定,鉴定结果与分离株一致,表明设计的探针具有高度特异性及准确性。80例临床标本分别用法国梅里埃API细菌鉴定系统及PCR杂交法进行检测,阳性率分别为(52.5%)和(67.5%),表明PCR结合寡核苷酸杂交法比传统的生物学培养法更为灵敏,值得推广。Universal primes and oligonucleotide probe with genus specific were designed. PCR targeted conserved sequence of bacterial 16S rDNA gene fragment, combined with dot-blot hybridization was used. .18 clinical bacteria and drug-resistant bacterial strains were amplified by PCR, and about 220 bp fragment were produced in the same condition. The probe only hybridized with corresponding PCR products. Results showed that every probe was highly specific. Compare to the bacterial strains of 37 clinical samples of 5 genuses. Those were authenticated by identified system of Biomeyieux API bacteria in France with those by this method. The result was the same Results showed that those probes were highly specific and highly accurate.80 clinical samples bested by identified system of Biomeyieux API bacteria in France The positive vote was 52.5% but bested by PCR and dot-blot by hybridization technique, the positive rate was 67.5%. The result showed that the samples' positive rate tested by this method was highly than traditional bioetional biolog culture technique. The sensitivity of bacterial samples was 18CFU/mL.This method is highly specific, sensitive, rapid, simple as well as economical and reduces the chances of cross contamination.

关 键 词：通用引物 PCR 寡核苷酸探针 致病菌 

分 类 号：R446.5[医药卫生—诊断学]