籽用西瓜SSR体系的建立与优化

ESTABLISHMENT AND OPTIMIZATION OF SSR SYSTEM IN SEED-USED WATERMELON

为了建立籽用西瓜SSR-PCR反应的优化体系,以籽用西瓜幼苗叶片为实验材料,利用L16(45)正交设计实验对影响SSR-PCR反应体系的主要组分进行了优化探索。结果表明,籽用西瓜的SSR反应体系适宜体积为20μL,各组分含量分别为:Taq DNA聚合酶0.5 U、引物0.5μmol/L、Mg2+1.8 mmol/L、d NTPs 0.2mmol/L、DNA 80ng。并且优化引物最适退火温度为50.9℃。对20份籽用西瓜品种应用此反应体系进行SSR扩增并进行电泳检测,获得扩增谱带清晰且多态性较好,证明此体系稳定可靠。

To establish an appropriate SSR-PCR reaction system of seed-used watermelon. The experiment applied L16（ 4^5） orthogonal design method to study the mainly factors which affected the SSR-PCR reaction of seed-used watermelon. The results showed that 0.5 U Taq polymerase,0.5 μmol / L primer,1.8 mmol / L Mg^2＋,0.2mmol/L d NTPs,80 ng template DNA were the optimal conditions in the reaction of 20 μL volume. The optimal annealing temperature of primer SSR was 50.9℃. Under optimal conditions,SSR reactions were carried out with 20 cultivars of seed-used watermelon and tested with electrophoresis. The results revealed clear and high polymorphic bands which demonstrated that the reaction system established in this experiment was stable and reliable.