To confirm the presence of inserts cells were lysed by three freeze–thawing
cycles, followed by 12 min sonication to get better results. Briefly, for PCR screening the vector specific primers SP6 (50-ATT TAG GTG ACA CTA TAG AAT AC-30) and T7 (50-TAA TAC GAC TCA CTA TAG GG-30) were used in a volume of 25 ll using
2.5 ll DNA extract as a template.