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Cloning and Sequencing of Tuberculosis Genes Rv0577 and Rv3846 for DNA Vaccine

Mussarat Qasim1, Asifa Hameed2*, Mirza Imran Shehzad1

Tuberculosis (TB) caused by Mycobacterium tuberculosis (M. tb) is the major health problem worldwide. It is a contagious, commonly prevalent disease in developing countries including Pakistan. Poverty in developing countries makes this disease an acute health problem. Almost one third of the world’s population is affected by TB. Pakistan ranks 5th among top Tuberculosis infected countries. Among 100,000 individuals about 280 people are infected with this disease in Pakistan. Vaccination is the way to prevent disease occurrence. Bacillus Calmette-Guerin (BCG) is the only licensed vaccine against TB throughout the world. However, protective immunity developed through this vaccine varies in different parts of the world. Hence, there is need to develop new DNA based anti TB vaccines based on information obtained from Mycobacterium tuberculosis whole genome sequencing, as DNA vaccines are capable to elicit both humoral and cell-mediated immune responses. In the present studies, we selected two Mycobacterium tuberculosis genes Rv0577 and Rv3846 for developing DNA vaccine. These genes were amplified through PCR, confirmed through restriction digestion, ligated in pTZ57R/T vector, transformed in DH5α cells. The clones were confirmed through restriction digestion and sequence analysis. In future, these clones can be used in sub-cloning into mammalian expression vectors and tested as DNA vaccines in mice model.