Amplification was done using a total of 50 µL reaction mixture containing 1 µg DNA template, 1× PCR Buffer with MgCl2, 50 µM of dNTP mix, 0.4 pmole of each forward and reverse primer, 2.5 U Taq polymerase, and 38.5 µL of autoclaved ddH2o to top up to the final reaction volume.