桃蚜基因组DNA提取及SSR反应体系优化和应用  

Extraction of genomic DNA from the peach-potato aphid Myzus persicae and optimization of SSR reaction system

作　　者：刘永刚[1] 漆永红[1] 赵丽娟[2] 吕和平[1] 

机构地区：[1]甘肃省农业科学院植物保护研究所,兰州730070 [2]甘肃省农业科学院生物技术研究所,兰州730070

出　　处：《植物保护》2009年第4期96-100,共5页Plant Protection

基　　金：农业部农业行业专项(200803024);甘肃省科技计划项目(0805XCNA071)

摘　　要：针对蚜虫体型微小，体表有外骨骼等特点对改进的醋酸钾（KAC）法作了优化，优化后的方法可在常温条件下更加简易和有效地提取高质量的单头蚜虫基因组DNA，结果表明，DNA样品的A260／280值普遍在1．6～1．9之间，电泳检测基因组DNA纯度和完整性较好。同时在20弘L的反应体系中将PCR的5个主要成分分别设定10个浓度梯度，研究了适合桃蚜SSR分子标记的优化体系，结果表明，最适宜的优化浓度分别为：1．5mmol／L Mg^2＋，0．25mmol／LdNTP，1．0UTaq酶，40ng／／μL模板DNA，25ng／μL引物。利用甘肃省酒泉地区马铃薯桃蚜来验证此优化体系，30％聚丙烯酰胺凝胶电泳检测结果显示，扩增产物大多在100-300bp，多态性高，且反应体系的稳定性和可重复性好。Genomic DNA of single aphids was simply and effectively extracted at room temperature with high quality. The results showed that the ratio of A260/280 of the extracted DNA ranged between 1.6-1.9. The extracted DNA was very pure and of integrity. Meanwhile, the reaction system of SSR was optimized. The reaction program of amplification was 1.5 mmol/L Mg^2＋ , 0. 25 mmol/L dNTP, 1.0U Taq, 40 ng/μL template DNA and 25 ng/μL random primer in 20 μL reaction system. To test the optimized SSR system, SSR fragments of 100--300 hp were detected in Myzus persicae from Jiuquan of Gansu Province by 30% polyacrylamide gel, indicating that the SSR reaction system had high polymorphism and was steady as well as reproducible.

关 键 词：桃蚜 基因组DNA 提取方法 SSR 体系优化 

分 类 号：Q523[生物学—生物化学] S436.621.21[农业科学—农业昆虫与害虫防治]