A modified PCR-based method for rapid non-radioactive detection of clinically important pathogens

被引：3

作者：

Gorelov, VN ^{[1
]}

Dumon, K ^{[1
]}

Barteneva, NS ^{[1
]}

Roher, HD ^{[1
]}

Goretzki, PE ^{[1
]}

机构：

[1] UNIV DUSSELDORF,DEPT SURG A,DUSSELDORF,GERMANY

来源：

MICROBIOLOGY AND IMMUNOLOGY | 1996年 / 40卷 / 09期

关键词：

polymerase chain reaction; direct nonradioactive labeling; bacterial pathogens;

D O I：

10.1111/j.1348-0421.1996.tb01117.x

中图分类号：

R392 [医学免疫学]; Q939.91 [免疫学];

学科分类号：

100102 ;

摘要：

We have devised a sensitive and rapid method for the detection of several bacterial pathogens in clinical specimens using PCR, This method has been named Direct Labeling and Detection Procedure (DLDP) and is based on the direct incorporation of a nonradioactive digoxigenin label (DIG-11-dUTP) into a microbial species-specific gene fragment during amplification, Following amplification, the resulting PCR products are cleansed of nonincorporated DIG-11-dUTP, spotted onto a nylon membrane, fixed by UV-crosslinking and the labeled DNA is visualized by digoxigenin detection reagents, Using cultivated reference bacteria (Staphylococcus aureus, Streptococcus pneumoniae, Pseudomonas aeruginosa) we were able to demonstrate a rapid and sensitive detection of <20 CFU of bacteria in human secretions (sputum, urine, mucous), The present study suggests that DLDP can be used as a reliable method for indication of bacteria in clinical or environmental specimens with the proviso that the selected corresponding oligonucleotide primers provide amplification of strong species-specific genes.

引用

页码：611 / 616

页数：6

共 19 条

[1]

BRAKSTAD OG, 1992, J CLIN MICROBIOL, V30, P1650

[2] DETECTION OF CHLAMYDIA-TRACHOMATIS IN CLINICAL SPECIMENS BY THE POLYMERASE CHAIN-REACTION [J].