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用改良的SDS法从云南箭竹硅胶干燥的叶片中提取基因组DNA进行RAPD扩增。通过正交法对RAPD反应条件优化,6个试验因子的最适条件为:模板DNA1.5ng/μL,随机引物0.8～1.0μmol/L,dNTPs浓度0.1～0.15mmol/L,Mg2+浓度2.0～2.5mmol/L,Taq酶用量0.5～1.0U。最佳热循环参数为:94℃预变性2min,之后进行40次循环(94℃变性30s,36℃退火60s,72℃延伸90s),最后72℃总延伸7min后在4℃终止反应。
Genomic DNA was extracted from the dried leaves of D. yunnanensis by a modified SDS method for RAPD amplification. The optimum conditions of RAPD reaction were optimized by orthogonal test. The optimal conditions for the six experimental factors were: template DNA 1.5 ng / μL, random primer 0.8-1.0 μmol / L, dNTPs 0.1-0.15 mmol / L, Mg 2+ concentration 2.0-2.5 mmol / L, Taq enzyme dosage 0.5 ~ 1.0U. The optimal thermal cycling parameters were as follows: pre-denaturation at 94 ° C for 2 min followed by 40 cycles of denaturation at 94 ° C for 30 s, annealing at 36 ° C for 60 s, extension at 72 ° C for 90 s, and final extension at 72 ° C for 7 min followed by termination at 4 ° C.