关键词：
羽衣甘蓝(Brassica oleracea var.acephala);SSR;正交设计;体系优化
摘 要：
为优化羽衣甘蓝(Brassica oleracea var.acephala)SSR反应体系,并利用优化的体系进行引物的筛选。采用单因素完全随机试验筛选各反应因素的最佳水平,并采用L16(45)正交设计进行试验优化,建立了羽衣甘蓝SSR-PCR最佳反应体系。结果表明,各因素不同水平浓度对扩增结果均有一定影响,羽衣甘蓝基因组DNA的最佳SSR反应体系为DNA模板量(50 ng/μL)1.0μL,10×PCR Buffer 1.0μL,引物量(2.0μmol/L)上下游各3.0μL,dNTPs(2.5 mmol/L)0.8μL,Taq聚合酶(5 U/μL)0.2μL,ddH2O 1.0μL,总体积10μL。检测了优化体系的稳定性,利用优化的反应体系对芸薹属植物20对SSR引物进行扩增并用聚丙烯酰胺凝胶电泳检测,在所选用的20对引物中,18对扩增出了清晰的条带,11对具有特异扩增条带,多态性引物比率为61%,验证了该体系可用于羽衣甘蓝SSR-PCR扩增和SSR引物的筛选。
译 名：
Optimizing SSR System of Brassica oleracea var. acephala
作 者：
LIU Chang;ZHU Peng-fang;FANG Xia;KANG Yao-hai;ZHAO Ying;College of Forestry, Shenyang Agricultural University;Liaoning Urban Construction Technical College;
关键词：
Brassica oleracea var.acephala;;SSR;;orthogonal design;;system optimization
摘 要：
The SSR system of Brassica oleracea var. acephala was optimized by screening primers. The single factor with random design was used to optimize the SSR-PCR amplification system and the L16(45) orthogonal design was used. Results showed that different levels of each factor had different effects on PCR. The optimal system was DNA template(50 ng / μL)1.0 μL, 10×PCR Buffer1.0 μL, 2.0 μmol / L upstream primer and downstream 3.0 μL each, dNTPs(2.5 mmol / L)0.8 μL, Taq DNA polymerase(5U / μL) 0.2 μL and ddH2O 1.0 μL. Twenty pairs of primers were used to test the optimized system by polyacrylamide gel electrophoresis. Eighteen pairs of primers had clear bands and eleven pairs of primers were with polymorphic, with the percentage of polymorphism of 61%. The optimized system could be used for SSR-PCR.
