Isolation of Genomic DNA of Dangshansu Pearand Optimization of Its RAPD Analysis Condition
The isolation effect of genomic DNA from mature leaf, tender leaf and bud of Dangshansu pear was studied respectively by SDS method. After comparing the quantity and quality of genomic DNA isolated from different materials,the highquality DNA was used as template to optimize the reaction condition of RAPD analysis system. The result showed that the genomic DNA isolated from tender leaf and bud could be used for RAPD analysis and that isolated from mature leaf couldn't be used directly. The optimum reaction condition of RAPD analysis system for Dangshansu pear was as follows: total reaction volume 50μl, including about 80 ng template DNA,5 μL 10×PCR Buffer purchased, 20 mmol/L MgCl2,120 μmol/L dNTPs,3 unite Taq polymerase, 05 μmol/L random primer; the program of heating:predenaturation 5 min at 94℃ to make template DNA denaturating sufficiently,then undergo a proceeding of reaction cycles as denaturation 30 s at 94℃,anneal 45 s at 36℃, polymerization 90 s at 72℃, 40 cycles, finally polymerization 7 min at 72℃。