Abstract:
In order to establish a suitable ISSR-PCR reaction system for Zizania latifolia, the wetlands plant Zizania latifolia genomic DNA was optimized with choosing the primer UBC818. We used the orthogonal design to establish an ISSR-PCR reaction system and amplification program. The results indicated that 25 μL reaction system containing 1×PCR buffer, 250 μmol/L dNTP, 2.0 mmol/L MgCl₂, 0.5 μmol/L primer, 1 U Taq DNA polymerase and 100 ng template DNA was suitable to ISSR-PCR amplifying. Amplification program as followed: pre-denaturation at 94℃ for 5 min, denaturrtion at 94℃ for 1 min, annealing at 56℃ for 45 s, extending at 72℃ for 90 s, after 35 cycles, then extending again at 72℃ for 10 min and keeping at 4℃.