苹果SSR反应体系的建立

为摸索适宜苹果的SSR反应体系,以金冠苹果为试验材料,研究了苹果SSR技术中PCR反应体系的主要成分对SSR扩增结果的影响。对SSR反应体系中的Mg2+浓度、引物浓度、dNTP浓度、TaqDNA聚合酶浓度、模板浓度以及退火温度进行了探索,确立了适合苹果的SSR反应体系为:在20μL反应体系中,Mg2+、引物和dNTP的最适浓度分别为2.5mmol/L、0.8μmol/L、0.10mmol/L;反应体系中TaqDNA聚合酶宜加入1U,模板DNA应加入15-30ng;引物的最佳退火温度为48.5-52.0℃。并利用该反应体系对10个苹果代表品种进行SSR反应,用8%的非变性聚丙烯酰胺凝胶电泳检测,不同品种间DNA谱带多态性丰富,证实该体系稳定可靠。

Establishment of SSR analysis system in apple

In this study, SSR conditions on apple were optimizatied . The factors, which affected the SSR results of Golden Delicious apple cultivar were studied. The results showed that 2.5 mmol/L Mg2+, 0.8 μmol/L primer, 0.10 mmol/L dNTP, 1 U Taq polymerase, 15~30 ng template DNA and the optimal annealing temperature of primers of 48.5~52.0℃ in 20 μL SSR reaction system were the best. Trough above PCR system, SSR fragments of 10 apple cultivars were obtained and detected. Polymorphism between different cultivars was abundantly detected by 8% polyacrylamide gel, which showed this system was suitable and stable.