古樟ISSR扩增条件的优化  

Optimization of ISSR Amplification Conditions in Ancient Camphor-trees

作　　者：黄萌[1,2] 陈尚钘[1,2] 杨光耀[1,2] 王宗德[1,2] 季春峰[1,2] 

机构地区：[1]江西农业大学园林与艺术学院,江西南昌330045 [2]江西农业大学,江西省竹子种质与利用重点实验室,江西南昌330045

出　　处：《北方园艺》2013年第7期116-120,共5页Northern Horticulture

基　　金：国家林业公益性行业科研专项资助项目(201004073)

摘　　要：采用改良CTAB法提取古樟叶片基因组DNA。利用单因素试验设计,对反应体系中退火温度、模板DNA含量、Mg2+浓度、dNTP浓度、引物浓度、Taq DNA聚合酶用量、去离子甲酰胺等7种因素不同梯度对扩增结果的影响进行了比较分析。结果表明:古樟ISSR-PCR最佳反应体系为,在20μL PCR反应体积中,含50ng DNA模板,2.5mmol/L MgCl2,0.20mmol/LdNTPS,1UTaq DNA聚合酶,0.5μmol/L引物浓度,2%去离子甲酰胺。该反应体系的建立为今后利用ISSR分子技术对古樟的遗传多样性分析提供了技术支撑。Abstract. The genomic DNA of ancient camphor-trees was extracted from leaves by improved CTAB method. With the design of single factor experiment method, the influences of seven factors including Mg2＋ , dNTP,DNA content in the template, Taq DNA polymerase, amounts of primer,deionized formamide,and annealing temperature on PCR amplification were analyzed. The results showed that the best ISSR-PCR reaction system of ancient camphor-trees was as follows.. 50 ng DNA template, 2. 5 mmol/L MgCl2,0. 20 mmol/L dNTPS, 1.0 U Taq polymerase, 0.5 μmol/L primer and 2% formamide deionized in 20 /IL reaction volume. The results would lay a technology foundation for further study of the genetic diversity of ancient camphor-trees with ISSR molecular marking.

关 键 词：古樟 ISSR 反应条件 优化 

分 类 号：S792.23[农业科学—林木遗传育种]