马铃薯种质遗传多样性分析的AFLP反应体系优化与引物筛选

Optimization of AFLP Reaction System and Primer Screening for Analysis of Genetic Diversity of Potato Germplasm

本试验主要对AFLP反应体系进行优化,并用该体系筛选适用于马铃薯种质资源遗传多样性分析的引物。酶切连接、预扩增和选择性扩增体系的优化结果表明,建立的马铃薯AFLP反应体系为:模板DNA160ng,37℃酶切连接12h;预扩增体系中dNTP浓度为0.2mmol/L,TaqDNA聚合酶用量为1U;选择性扩增体系中预扩增产物稀释20倍,引物终浓度为2.5ng/μL。利用优化后的AFLP反应体系,以5个马铃薯品种为试材筛选选择性引物,可以从81对引物组合中选出16对条带丰富且多态性较高的引物组合。本研究为种质资源鉴定、马铃薯遗传多样性以及马铃薯抗病性研究等奠定了良好的基础。

In this research, we have optimized AFLP reaction system, then we used the optimized system to screen applicable primers for studying genetic diversity of potato germplasm. After optimizing the system of digestion-ligation, pre-amplification and selective amplification, the established AFLP reaction system in patato were as follows： template DNA 160 ng, digestion-ligation 37℃ for 12 h; 0.2 mmol/L dNTP, 1 U Taq DNA polymerase in pre-amplification system; amplification products should be diluted 20 times, and the final concentration of primer was 2.5 ng/uL in selective pre-amplification system. We also employed 5 potato germplasm as the tested material to screeen selectivity primers by using the optimized AFLP reaction system, and we acquired 16 primers combination with rich bands and high polymorphism from tested 81 primers combination. This studies would be establish a good foundation for germplasm identification, genetic diversity of potato as well as the potato disease resistance research.