Volume 14 Supplement 3

Abstracts from the 2nd International Science Symposium on HIV and Infectious Diseases (HIV SCIENCE 2014)

Open Access

Generation of differentially regulated genes in Mycobacterium tuberculosis isolates from cerebrospinal fluid and respiratory secretions using suppression subtractive hybridization

BMC Infectious Diseases201414(Suppl 3):O16

https://doi.org/10.1186/1471-2334-14-S3-O16

Published: 27 May 2014

Background

Suppression Subtractive Hybridization (SSH) is an established method for separating DNA molecules that distinguish two closely related DNA samples, which has become a powerful method for generating subtracted cDNA libraries. In this study, we postulated that SSH could be used to identify significant genetic differences between Mycobacterium tuberculosis causing meningitis and M. tuberculosis causing pulmonary tuberculosis.

Hypothesis

The unique DNA patterns present only in cerebrospinal fluid (CSF) TB isolates might be markers for neurotropism and neurovirulence in some strains of M. tuberculosis.

Methods

Bacterial DNAs were isolated and SSH was performed using PCR-Select Bacterial Genome Subtraction Kit (Clontech). Twenty subtracted DNAs were generated from 3 CSF isolates and 3 respiratory isolates. Molecular cloning was done on the subtracted DNAs and the positive isolates were extracted, purified and sequenced for analysis.

Results

So far, 9 of the subtracted DNAs have been successfully cloned into pGEM-T vector, resulting in 21 positive clones. Two preliminary BLASTP results revealed several mycobacterial PE/PPE proteins which play a role in the evasion of host immune responses, possibly via antigenic variation, together with conserved hypothetical proteins with unknown function.

Conclusion

Further studies are needed to determine the role of these subtracted DNAs.

Authors’ Affiliations

(1)
Department of Medical Microbiology, Faculty of Medicine, University of Malaya
(2)
Department of Biomedical Science, Faculty of Science, Universiti Tunku Abdul Rahman
(3)
School of Biosciences, Taylor’s University Lakeside Campus, Subang Jaya

Copyright

© Saw et al; licensee BioMed Central Ltd. 2014

This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.

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