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白背飞虱Sogatella furcifera(Horváth)是水稻重要害虫之一。研究采用L16(45)正交设计优化试验,构建了白背飞虱RAPD最优反应体系,并对引物退火温度和反应循环次数进行了优化。结果表明:白背飞虱RAPDPCR最优反应体系为2μL 10×PCR Buffer(10 mmol/L Tris-HCl;50 mmol/L KCl),2.4 mmol/L Mg2+,0.15 mmol/L dNTPs,0.7μmol/L引物,0.8U Taq DNA Polymerase和30 ng模板DNA,ddH2O补充至20μL。本研究采用的10条引物的最佳退火温度在35-39℃间,以引物相应最优退火温度和40次PCR反应循环可获得较好的白背飞虱RAPD-PCR结果。
White-backed planthopper Sogatella furcifera (Horváth) is one of the major pests in rice. In order to optimize the RAPD reaction system, the L16 (45) orthogonal design optimization experiment was carried out. The annealing temperature and the number of reaction cycles were optimized. The results showed that the optimal reaction system of RAPDPCR was 2 μL 10 × PCR Buffer (10 mmol / L Tris-HCl, 50 mmol / L KCl), 2.4 mmol / L Mg2 +, 0.15 mmol / L dNTPs, 0.7 μmol / L Primer, 0.8U Taq DNA Polymerase and 30 ng template DNA, ddH2O supplemented to 20 μL. The best annealing temperature of the 10 primers used in this study was between 35-39 ℃. The RAPD-PCR results of the whitebacked planthopper, Siphonaptera lugens, were obtained by using the corresponding optimal annealing temperature and 40 PCR cycles.