Abstract:

In this study, the genome DNA was extracted from Xuehua pear and was used for inter-simple sequence repeat(ISSR)analysis. By evaluating amplification concentrations and conditions,the optimum reaction system of ISSR-PCR for Pyrus L. were established in a total volume of 20μL, containing template DNA 40 ng, 1.0 U Taq-DNA polymerase, 0.2μmol?L-1ISSR primer, 200μmol?L-1, dNTP, Buffer (Tris-HCl (pH8.3) 10m mol?L-1、KCl 50 mmol?L-1、 MgCl21.5 mmol?L-1). Amplified procedure was programmed for 5 minutes at 94℃, at annealing temperature for 60s,extension 120s at 72℃; 39 cycles: 60 seconds at 94℃, at annealing temperature(52℃) for 60s, extension 120s at 72℃and then a final extension 72℃ for 10 min after the last cycle and then keep at 4℃.Amplified products were detected by electrophoresis in 1.4% agarose gel (containing 1μg?mL-1 EB).