OPTIMIZATION OF THE PCR REACTION MIXTURE COMPONENT COMPOSITION FOR THE OBTAINING OF MYCOBACTERIAL GENES

Bovine tuberculosis is an infectious respiratory disease of cattle without specific clinical signs, caused by the bacterium Mycobacterium bovis (M. bovis), which can cross the species barrier, infect and cause disease in many other wild and domestic animals. Immunization of cattle by vaccination is a standard preventive measure in the fight against tuberculosis. The only currently available vaccine against human and bovine tuberculosis is live attenuated Bacillus Calmette-Guérin (BCG) and its modified variants. The need to develop a domestic vaccine that provides higher and more stable protection than the BCG vaccine is relevant. And the purpose of this work is to select the optimal component composition of the PCR reaction mixture for the production of protective proteins M. bovis Esat-6 and TB10.4. As a result of work to optimize the PCR setting for the obtaining mycobacterial genes of M. bovis, Taq DNA polymerase containing MgCl2 – 0.3 μl; 10x PCR buffer – 2.5 µl; specific primers (20 pM) – 1 µl each, dNTP mixture (10 mM)-1 µl; target DNA – 5 µl were used as part of the reaction mixture in a total volume of 25 µl with optimized mode: 94°C – 5 min, 94°C – 1 min, 55/62°C – 40 sec, 72°C – 1.30 min (35 cycles), and 72°C – 7 min, 4°C – cooling and storage indefinitely

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