| 黄鳝ISSR-PCR反应体系的建立及条件优化 |
| |
| 引用本文： | 杨浩,陈宏喜,陈欣,杨太有.黄鳝ISSR-PCR反应体系的建立及条件优化[J].生物技术通报,2009(7). |
| |
| 作者姓名： | 杨浩 陈宏喜 陈欣 杨太有 |
| |
| 作者单位： | 河南师范大学生命科学学院,新乡,453007 |
| |
| 基金项目： | 国家大学生创新性实验计划项目，河南省教育厅自然科学研究项目，河南省动物学重点学科资助;河南师范大学国家级生命科学实验教学示范中心资助 |
| |
| 摘 要： | 以黄鳝基因组DNA为模板,采用正交试验设计方法,对各反应因子、引物退火温度和循环参数进行优化.建立了黄鳝的最适ISSR-PCR反应体系,25 μl反应体系中含2.5 mmol/ L Mg2+,250 μmol/ L dNTPs,0.25 μmol/ L 引物,1.0 U Taq DNA聚合酶,30 ng DNA模板.最佳反应程序:94℃预变性5 min;94℃变性40 s,48～57℃复性40 s(随引物而确定),72℃延伸1.5 min,循环次数40;72℃延伸10 min.利用所建立的ISSR反应体系,获得了清晰、重复性好、多态性高的DNA谱带.
|
| 关 键 词： | 黄鳝 正交设计 反应体系 |
Optimization for ISSR Reaction System of Monopterus albus with Orthogonal Design |
| |
| Yang Hao,Chen Hongxi,Chen Xin,Yang Taiyou.Optimization for ISSR Reaction System of Monopterus albus with Orthogonal Design[J].Biotechnology Bulletin,2009(7). |
| |
| Authors: | Yang Hao Chen Hongxi Chen Xin Yang Taiyou |
| |
| Abstract: | Based on the genomic DNA of Monopterus albus, the factors influencing ISSR-PCR system were explored with the orthogonal design. The optimized ISSR-PCR system (25 μl) was determined as follows: 2.5 mmol / L Mg2+, 250 μmol / L dNTPs, 0.25 μmol / L primers, 1.0 U Taq DNA polymerase, 30 ng DNA template. The reaction program was set including initial denaturation for 5 minutes at 94℃, 40 cycles of denaturation for 40 seconds at 94℃,annealing for 40 seconds at 47 ～57℃, extension for 1.5 minutes at 72℃,with a final extension of 10 minutes at 72℃. The clear and reproducible amplification was obtained by the established ISSR reaction system. |
| |
| Keywords: | ISSR Monopterus albus ISSR Orthogonal design Reaction system |
| 本文献已被 万方数据 等数据库收录！ |
|
