安徽农业科学2009，Vol.35Issue(19)：8876-8878,8880,4.
火棘基因组DNA的提取和RAPD稳定的反应体系的建立
Research on the Genomic DNA Extraction and Establishment of Reproducible RAPD Reaction System for Pyracantha fortuneana
摘要
Abstract
[Objective] The reference for the research on the diversity and resource of Pyracantha fortuneana and the identification of the relation of its varieties were provided through the study on the optimal method of DNA extraction and stable system of RAPD reaction.[Method] The genomic DNA of Pyracantha fortuneana was successfully extracted with the improved CTAB method and the optimal condition in the system was selected with the experimental design of 4 factors(Taq enzyme, MgcL2, dNTPs and primer) at 4 levels.[Results] The electrophoresis result indicated that the quality and concentration of extracted DNA were very high(500 ng/μl).After that, the reproducible RAPD reaction system was set up with extracted DNA as template.RAPDs were performed in volumes of 25 μl, containing 25 ng of DNA, 1× reaction buffer(TaKaRa Dalian), 2.3 mmol/L MgCl2(TaKaRa Dalian), 1.0 μmol/L primer(SBS Beijing), 0.15 mmol/L mixture of dNTPs(TaKaRa Dalian) and 2.0 unit Taq DNA Polymerase(promega Shanghai).The procedure of RAPD was thermal cycler (HYBAID) was programmed for 1 cycle at 94 ℃ for 4 minutes, followed by 40 cycles at 94 ℃ for 40 seconds, at 36.8 ℃ for 50 seconds and at 72 ℃ for 70 seconds, and finally by 1 cycle at 72℃ for 4 minutes.[Conclusion] The genomic DNA of Pyracantha fortuneana was extracted with the improved CTAB method and the reaction system was established for the stable and reliable the procedure of RAPD.关键词
火棘/RAPD/反应体系Key words
Pyracantha fortuneana/RAPD/Reaction system分类
农业科技引用本文复制引用
韩颖..火棘基因组DNA的提取和RAPD稳定的反应体系的建立[J].安徽农业科学,2009,35(19):8876-8878,8880,4.基金项目
西南科技大学人才引进基金项目(zk043081) . （zk043081）
