论文部分内容阅读
为了得到适合研究瓜蒌农家品种的ISSR-PCR反应条件,本研究采用单因素实验和正交实验,对瓜蒌ISSR反应体系进行了优化,确立了适合瓜蒌的ISSR反应体系:20μL反应体系,含0.5μmol/L引物、2×Master Mix(Taq DNA Polymerase)10μL、35 ng模板DNA。通过性状差异较大样品的单因素试验,确定了能够适合尽量多样品的ISSR最佳反应程序为:94℃预变性5 min;94℃变性30 s,退火1 min,72℃延伸45 s,35个循环;72℃延伸7min,4℃保存。不同的引物所需要的退火温度不尽一致。
In order to obtain ISSR-PCR reaction conditions suitable for cultivating Agkistrodon fruits, single-factor experiments and orthogonal experiments were used to optimize the ISSR reaction system in Agkistrodon melon. The ISSR reaction system suitable for Agkistrodon Mucostris was established: 20μL reaction system, Including 0.5μmol / L primer, 2 × Master Mix (Taq DNA Polymerase) 10μL, 35 ng template DNA. The optimal reaction procedure of ISSR was as follows: pre-denaturation at 94 ℃ for 5 min, denaturation at 94 ℃ for 30 s, annealing at 1 min, extension at 72 ℃ for 45 s, 35 A cycle; 72 ℃ extended 7min, 4 ℃ preservation. Different primers require different annealing temperatures.