仁用杏SRAP-PCR体系的正交设计优化.PDFVIP

30 3 Vol. 30, No . 3 2009 9 Journal of Hebei University of Science and T echnology Se t. 2009 : 1008-1542( 2009) 03-0248- 05 SRAP-PCR 艾鹏飞, 甄志军, 方闪闪, 靳占忠 ( 河北科技大学生物科学与工程学院, 河北石家庄050018) 4 : 改良CTAB 法提取仁用杏基因组DNA, 用于建立SRA P- PCR 体系 用L16 (4 ) 正交设 计试验, 对影响仁用杏SRAP-PCR 反应体系的4 因素( dNT Ps 浓度Mg2+ 浓度引物浓度Taq 聚 合酶量) 4 水平进行筛选, PCR 结果经统计软件SPSS V 13. 0 分析, 并对退火温度进行了摸索, 确立 了仁用杏SRAP-PCR 的优化体系: 总体积为20 L, dNTPs 浓度为0. 2 mmol/ L, Mg2+ 浓度为2. 5 mmol/ L, 引物浓度为0. 9 mol/ L, T aq 聚合酶量为1. 5 U , 退火温度为52 本优化体系为仁用 杏资源遗传分析奠定了技术基础 : SRA P-PCR; 正交设计; 仁用杏 : S565. 5 : A O timization of SRAP-PCR system for kernelled a ricot by orthogonal design AI Peng-fei, ZHEN Zh-i jun, FANG Shan-shan, JIN Zhan-zhong ( College of Bioscience and Engineering, H ebei Universit y of Science and Technology, Shijiazhuang Hebei 050018, China) Abstract:The t o -qualit y genomic DNA extracted from leaves of kernelled a ricot with an im roved method of CT AB w as a - lied to o timize the SRAP-PCR system. The four levels of four factors ( dNTPs, Mg2+ , rimers, T aq DNA olymerase) in 4 SRAP-PCR system w ere studied by L ( 4 ) orthogonal design. T he electro horesis rofiles of SRAP-PCR w ere analyzed by 16 software SPSS V 13.0. T he annealing tem erature of PCR reaction was also o timized. The o timum SRAP-PCR system w as show n as follows: 0. 2 mmol/ L dNT Ps, 2. 5 mmol/ L Mg2+ , 0. 9 mol/ L rimers, 1. 5 U T aq DNA olymerase in reaction solu- t ion of 20L and annealing t em erat ure at 52 . T he o timum SRAP-PCR syst em