摘要
本研究旨在扩增出条带清晰、非特异性条带少的SSR/InDel标记产物。选择来自6个国家的12份不同基因型甜菜品种,利用SSR及InDel两种标记方法,Touch-down PCR、梯度PCR及常规固定退火温度PCR三种反应程序扩增产物。通过比较扩增产物的多态性、稳定性、清晰度选择适合两种分子标记方法的PCR反应程序。结果表明:Touch-down PCR法可以通过一次反应即顺利扩增出带型清晰,无无效带,扩增效果稳定的样本产物。梯度PCR法扩增结果不稳定,有非特异性条带出现并且弥散现象严重。固定退火温度PCR扩增结果因引物不同而差异较大,同时伴有弥散现象。Touch-down PCR扩增产物特异性,稳定性均高于梯度PCR及固定退火温度PCR,且较梯度PCR法省略了摸索最适退火温度的过程。因此,推荐使用Touch-down PCR程序进行甜菜SSR与InDel分子标记法研究。
To amplify the SSR/InDel labeled products with clear and less nonspecific bands, twelve differentgenotype beet cultivars from six countries were selected. The products were amplified using SSR markers,InDel markers and three PCR procedures, including touch-down PCR, gradient PCR, and conventional fixedannealing temperature PCR. By comparing the polymorphism, stability, and clarity of the amplificationproduct, a PCR reaction program suitable for both molecular markers was selected. The results show that thetouch-down PCR method can successfully amplify the sample products with clear bands and stableamplification effect, without invalid product through a single reaction. Gradient PCR amplification results areunstable, there are non-specific bands and diffuse phenomenon is serious. The fixed annealing temperaturePCR has different amplification results due to different primers, and accompanied by diffusion phenomenon.The specificity and stability of touch-down PCR amplification products are higher than that of gradient PCRand fixed annealing temperature PCR, meanwhile the process of groping the optimal annealing temperature isomitted compared with gradient PCR method. Therefore, touch-down PCR procedure is recommended forsugar beet SSR and InDel molecular markers.
作者
刘乃新
刘蕊
吴玉梅
吴则东
Liu Naixin;Liu Rui;Wu Yumei;Wu Zedong(Academy of Crop Science,Heilongjiang University,Harbin 150080;Faculty of Life Science and Technology,Harbin Normal University,Harbin 150025)
出处
《中国农学通报》
2018年第30期76-80,共5页
Chinese Agricultural Science Bulletin
基金
国家现代农业产业技术体系(糖料)建设专项资金资助(CARS-17-0503)
农业行业标准<甜菜等级规格糖料甜菜>(181721301092371116)
农业行业标准制定和修订<甜菜种子贮藏技术规程>(181821301092371022)
农业农村部种子管理局项目<甜菜登记品种分子数据库建设与完善>(111821301092361010)
