The total volume of the nested PCR reactions was 50 ll, which contained 1 ll amplified DNA, 10 ll 1 mM deoxy-ribonucleotide triphosphates (dNTPs, MBI Fermentas), 0.5 ll of each primer, 5 ll 10X Taq buffer (MBI Fermentas), 2.5 ll 25 mM MgCl2, 0,2 ll Taq polymerase (2 U) (MBI Fermentas) and 28.5 ll water.