兴安落叶松SSR反应体系的建立与优化  

Optimization of SSR-PCR Reaction System for Larix gmelinii

作　　者：冷海楠[1] 张玉[1] 徐明仪 王丽媛[1] 付晓玲[1] 崔福星[1] 杨立宾[1] 朱道光[1] LENG Hainan(Institute of Natural Resources and Ecology,HAS.National and Provincial Joint Engineering Laboratory of Wetlands and Ecological Conservation,Heilongjiang Harbin 150040)

机构地区：[1]黑龙江省科学院自然与生态研究所湿地与生态保育国家地方联合工程实验室

出　　处：《林业科技》2020年第1期12-14,共3页Forestry Science & Technology

基　　金：黑龙江省科学院青年创新基金面上项目(CXMS2018ZR03);国家自然科学基金青年基金项目(31700424)

摘　　要：采用L16(4^5)正交试验设计,对影响兴安落叶松SSR-PCR反应体系的主要因素(模板DNA、d NTPs、Mg^2+、Taq聚合酶浓度及引物浓度)进行了试验分析,其结果表明:兴安落叶松SSR-PCR体积为20μL的最佳反应体系是模板DNA用量50 ng、正反向引物浓度均为5μmol·L^-1、d NTPs浓度为2.5 mmol·L^-1、Mg^2+的浓度为2.5 mmol·L^-1、Taq聚合酶用量为1.0 U、最佳退火温度为60℃。试验所建立的反应体系可进一步用于兴安落叶松SSR遗传图谱构建、多样性分析、良种选育等方面的研究中。The important basis of the researches on SSR marker assisting selective breeding of Larix gmelinii is to establish an SSR-PCR reaction system and get some polymorphism primers. In order to lay a foundation for the researches on applying SSR marker technology to establishment of genetic diversity and molecular breeding, some main impact factors in the Larix gmelinii SSR-PCR reaction system were analyzed by using combination method of L1 6(4^5) orthogonal test and single factor tests, including DNA templates,primer, dNTPs, Mg^2+, Taq polymerase, and annealing temperature. The results showed that the optimized PCR reaction mixture was 20 μL total volume including 50 ng template DNA, 5 μmol/L forward and reverse primers, 2.5 mmol/L dNTPs, 2.5 mmol/L Mg^2+ and 1.0 U Taq DNA polymerase, and the optimum annealing temperature was 60℃. This result indicated that the optimized SSR-PCR reaction system could provide a vital theoretic and technical support for genetic maps establishment and molecular marker assisted breeding in Larix gmelinii.

关 键 词：兴安落叶松 SSR-PCR 正交优化试验 

分 类 号：S791.222[农业科学—林木遗传育种]