云南栘[木衣]ISSR-PCR反应体系的建立及引物筛选  

Establishment of ISSR-PCR Reaction System of Docynia delavayi(Franch.)Schneid and Primers Screening

作　　者：刘宇 朱泽莉 彭劲谕 王大玮 段安安 Liu Yu;Zhu Zeli;Peng Jingyu;Wang Dawei(Duan An'an College of Forestry,Southwest Forestry University,Kunming,650224)

机构地区：[1]西南林业大学林学院,昆明650224

出　　处：《分子植物育种》2023年第21期7097-7104,共8页Molecular Plant Breeding

基　　金：国家自然科学基金项目(32060350)资助。

摘　　要：为建立适合云南栘[木衣]的ISSR-PCR反应体系,本研究通过L_(16)(4~5)正交设计对影响PCR扩增反应的5个因素(模板DNA,引物,Mg~(2+),d NTPs,Taq DNA聚合酶)进行优化;在建立反应体系的基础上,筛选出多态性高的引物并确定其退火温度。结果表明,在25μL反应体系中,最佳反应体系用量为模板DNA 20 ng、引物0.3μmol/L、Mg~(2+)1.5 mmol/L、d NTPs 0.15 mmol/L、Taq DNA聚合酶1.5 U。建立的反应体系扩增稳定,筛选出的11条引物多态性较好、条带清晰。本研究反应体系的建立可为云南栘[木衣]遗传多样性研究提供技术支持。In order to establish ISSR-PCR reaction system suitable for Docynia delavayi(Franch.)Schneid,the five factors(template DNA,primers,Mg2+,dNTPs,Taq DNA polymerase)that affect the PCR amplification reaction were optimized by L16(45)orthogonal design;Based on the system,screen out primers with high polymorphism and determine their annealing temperature.The results showed that in the 25μL reaction system,the optimal reaction system dosage is 20 ng template DNA,0.3μmol/L primer,Mg2+1.5 mmol/L,dNTPs 0.15 mmol/L,Taq DNA polymerase 1.5 U.The established reaction system was stable in amplification,and 11 primers with clear bands and good polymorphism were screened out.The establishment of this reaction system can provide technical support for genetic diversity analysis of Docynia delavayi(Franch.)Schneid.

关 键 词：云南栘[木衣] ISSR-PCR 正交设计 引物筛选 

分 类 号：S792.99[农业科学—林木遗传育种]