PCR restriction fragment length polymorphism analysis (PRA)-algorithm targeting 644 bp Heat Shock Protein 65 (hsp65) gene for differentiation of Mycobacterium spp.

被引:36
作者
Kim, H
Kim, SH
Shim, TS
Kim, MN
Bai, GH
Park, YG
Lee, SH
Cha, CY
Kook, YH
Kim, BJ
机构
[1] Seoul Natl Univ, Coll Med, Dept Microbiol, Seoul 110799, South Korea
[2] Seoul Natl Univ, Coll Med, Liver Res Inst, Seoul 110799, South Korea
[3] Univ Ulsan, Coll Med, Dept Internal Med, Div Pulm & Crit Care Med,Asan Med Ctr, Seoul 138600, South Korea
[4] Univ Ulsan, Coll Med, Dept Lab Med, Asan Med Ctr, Seoul 138600, South Korea
[5] Korean Natl Tuberculosis Assoc, Korean Inst Tuberculosis, Seoul 137140, South Korea
[6] Konkuk Univ, Coll Med, Dept Microbiol, Chungju 380230, South Korea
基金
新加坡国家研究基金会;
关键词
heat shock protein gene (hsp65); Mycobacterium; identification; PRA;
D O I
10.1016/j.mimet.2005.02.010
中图分类号
Q5 [生物化学];
学科分类号
071010 ; 081704 ;
摘要
A method based on PCR-restiiction fragment length polymorphism analysis (PRA) using a novel region of the hsp65 gene was developed for the rapid and exact identification of mycobacteria to the species level. A 644 bp region of hsp65 in 62 mycobacteria reference strains, and 4 related bacterial strains were amplified, and the amplified DNAs were subsequently digested with restriction enzymes, namely, AvaII, HphI, and HpaII. Most of the mycobacteria species were easily differentiated at the species level by the developed method. In particular, the method enabled the separation of M avium, M intracellulare and M tuberculosis to the species level by Avall digestion alone. An algorithm was constructed based on the results and a blind test was successfully performed on 251 clinical isolates, which had been characterized by conventional biochemical testing. Our results suggest that this novel PRA offers a simple, rapid, and accurate method for the identification of mycobacteria culture isolates at the species level. (c) 2005 Elsevier B.V. All rights reserved.
引用
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页码:199 / 209
页数:11
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