For molecular study small pieces (50 mg) of suitable samples
from different tissues described above were removed and stored
at 70 C. DNA was extracted from the samples using the DNP™
Kit (Cinna-Gene) as per manufacturer’s instructions. Then, following
extraction the DNA was stored at 70 C prior to analysis by
PCR. Primer pair Np21/Np6 was used for amplification of the Nc-
5 gene (Kang et al., 2009). The expected size of the amplicon with
this primer pair is 328 bp. Each 25 ll PCR reaction contained 2.5 ll
PCR buffer (10), 3 mM MgCl2, 200 lM of dNTP, 400 nM of each
primer and 4 ll of DNA. Thermal cycling was done as follows:
94 C for 4 min (as initial denaturing), 25–30 cycles of 94 C for