论文部分内容阅读
采用均匀设计方法,对影响兴安落叶松ISSR-PCR体系的MgCl2、dNTPs、引物、TaqDNA聚合酶和模板5个组分的浓度进行了U16(45)和U12(35)两轮优化,建立了适合于兴安落叶松的ISSR-PCR反应体系。该优化的20μL反应体系包含MgCl23.0 mmol/L、dNTPs 0.15 mmol/L、引物0.5μmol/L、TaqDNA聚合酶1.0U、模板20ng/20μL。并进一步进行梯度退火试验,找到了最适的兴安落叶松ISSR退火温度为56.5℃。利用所确立的体系对部分兴安落叶松种质进行扩增的结果清晰可靠,多态性好。
The uniform design method was used to optimize the concentrations of MgCl2, dNTPs, primer, TaqDNA polymerase and template in the ISSR-PCR system of Larix gmelinii for U16 (45) and U12 (35) ISSR-PCR reaction system of larch in Xing’an. The optimized 20μL reaction system contained 23.0mmol / L MgCl 2, 0.15mmol / L dNTPs, 0.5μmol / L primer, 1.0U Taq DNA polymerase and 20ng / 20μL template. And further gradient annealing test, find the optimum ISSR Larix gmelinii annealing temperature of 56.5 ℃. The result of using the established system to amplify the germplasm of some Larix gmelinii was clear and reliable and the polymorphism was good.