摘要
本研究以中国古老月季新鲜幼叶为试材,进行RAPD分子标记反应扩增体系的优化。对RAPD-PCR反应结果影响较大的五个因素(模板DNA的浓度,引物的浓度, Mg Cl2的浓度, dNTP的浓度, Taq DNA聚合酶的浓度)进行了单因素多水平设计。结果表明:20μL总反应体积中含模板DNA (10 ng/μL) 1.0μL、dNTP(2.5 mmol/L) 1.6μL、引物(2.0μmol/L) 1.5μL、Mg Cl2(25 mmol/L) 1.6μL、Taq DNA聚合酶(5 U/μL) 0.3μL、10×Buffer用量2.0μL,其余用去离子水补充。通过优化后的RAPD-PCR反应体系可获得清晰、稳定、条带多的电泳图,表明以上反应条件适合中国古老月季RAPD分子标记反应的体系。该体系的建立有利于中国古老月季的亲缘关系和遗传多样性分析。
In this research, we studied the optimization of RAPD molecular marker reaction amplification system based on the fresh young leaves of Chinese old garden roses. We carried out single factor and multi-level design on five factors(concentration of template DNA, concentration of primers, concentration of Mg Cl2, concentration of dNTP, and concentration of Taq DNA polymerase) that had great influence on RAPD-PCR reaction results. The results showed that the 20 μL overall reaction volume contained template(10 ng/μL) 1.0 μL, primers(2.0 μmol/L)1.5 μL, Mg Cl2(25 mmol/L) 1.6 μL, dNTP(2.5 mmol/L) 1.6 μL, Taq DNA polymerase(5 U/μL) 0.3 μL, 10×Buffer dosage of 2.0 μL, and the rest was supplemented with deionized water. The optimized RAPD-PCR reaction system could obtain a clear, stable, and multi strip electrophoretic diagram, indicating that the above reaction conditions were suitable for the RAPD molecular marker reaction system of Chinese old garden roses. The establishment of this system was conducive to the analysis of genetic relationship and genetic diversity of Chinese old garden roses.
作者
李玉洁
赵成日
Li Yujie;Zhao Chengri(Agricultural College of Yanbian University,Yanji,133002)
出处
《分子植物育种》
CAS
CSCD
北大核心
2018年第24期8073-8079,共7页
Molecular Plant Breeding
基金
国家自然科学基金项目(31660319)资助
关键词
中国古老月季
反应体系
RAPD
优化
Chinese old garden rose
Reaction system
RAPD
Optimization
