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Volume 23 |
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Published by : PROFESSIONAL MEDICAL PUBLICATIONS |
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Volume 23 |
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Number 2 |
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Application of restriction enzyme analysis technique based on 65kda heat shock protein gene for fingerprinting and differentiation of mycobacterium tuberculosis clinical strains isolated from tuberculosis patients in Ahwaz, Iran
Azar D. Khosravi1, Abdulrazagh Hashemi2ABSTRACT
Objective: Application of identification methodology of restriction enzyme analysis (REA) for fingerprinting of the expanded population of Mycobacterium tuberculosis (MTB) isolates.
Methodology: A total of 150 clinical isolates from patients referred to TB reference laboratory, Public Health Centre, Ahwaz, Iran, were identified as MTB by using conventional culture and biochemical tests from January to December 2004. The PCR-REA method uses a PCR step based on amplification of a 439 bp fragment of hsp65 gene involving genus specific primers and restriction enzyme analysis by digestion of products with HaeIII & BstEII enzymes were employed.
Results: The identical restriction patterns similar to MTB reference strains equal to 160 / 145 / 72bp fragments for Hae III and 250 /120/82bp fragments for Bst EII digests were seen in 145 isolates (96.6%). The diverse patterns were observed for five isolates in Hae III digest as 180 / 100 / 80 bp, 194/ 72 bp and 160/ 145 bp representing the possible intra-species variation within studied MTB strains , while their Bst EII digestion patterns showed no variation.
Conclusions: The PCR-REA technique revealed three different new patterns for Hae III digest. However to verify that they are indeed MTB isolates, a sequence-based analysis of the exceptional isolates should be performed.
KEY WORDS: M. tuberculosis, Restriction enzyme, 65 heat shock protein gene, PCR-REA.
Pak J Med Sci April 2007 Vol. 23 No. 2 216-219
1. Dr. Azar Khosravi
2. Abdulrazagh Hashemi
1-2: Dept. of Microbiology, School of Medicine &
Infectious and Tropical Diseases Research Center,
Ahwaz Jondi Shapour University of Medical Sciences,
Ahwaz, Iran.
Correspondences:
Dr. Azar Khosravi
E-Mail: khosraviaz@yahoo.com
* Received for Publication: May 23, 2006
* Revision Received: August 16, 2006
* Revision Accepted: August 31, 2006
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