Optimization of ISSR-PCR reaction system and its verification in common bean

• Published in: Molecular plant breeding Vol. 11; no. 4
• Main Authors: Yang Jing, Jilin Agricultural University, Changchun (China), Zhang Guangchen, Jilin Agricultural University, Changchun (China)
• Format: Journal Article
• Language: Chinese
• Published: 01.08.2013
• Subjects: ADN; DNA; http://www.fao.org/aos/agrovoc#c_2347; http://www.fao.org/aos/agrovoc#c_34079; http://www.fao.org/aos/agrovoc#c_5993; PCR; PLANTE; PLANTS
• ISSN: 1672-416X

用试剂盒法提取菜豆（Phaseolus vulgarisL.）嫩叶的基因组DNA，并以此为模板对影响ISSR-PCR反应体系的Taq酶量、模板浓度、dNTP浓度、Mg2＋浓度、引物浓度5个因素进行了优化。结果表明总体积为25μL的菜豆ISSR-PCR反应最优体系为：45ng模板DNA，1UTaq酶，2mmol/LMg2＋，0.6μmol/L引物和0.1mmol/LdNTP，在此基础上对循环次数进行优化，最终确定最佳循环次数为45次。利用该体系，选取引物823对8份材料进行扩增，验证了该体系的稳定性。本研究旨在建立菜豆ISSR反应的最优体系，为今后利用该分子标记技术进行有关菜豆种质及遗传方面的研究奠定技术基础。 The genomic DNA of common bean was extracted from tender leaves by DNA secure plant kit method, and optimized ISSR reaction condition with it as template. We established optimum ISSR-PCR system for common bean using ISSR-PCR amplification with five factors (Taq DNA polymerase, DNA template, dNTP, Mg2＋ concentr