苹果愈伤组织SSR-PCR反应体系优化  

Optimization of SSR-PCR Reaction System for Apple Callus

作　　者：高兵[1,2] 安文杰[1] 孙俊 GAO Bing;AN Wenjie;SUN Jun(Department of Horticulture, Shanxi Forestry Vocational and Technical College, Taiyuan, Shanxi 030009;College of Horticulture, Nanjing Agricultural University, Nanjing, Jiangsu 210095;College of Horticulture, Anhui Agricultural University, Hefei, Anhui 230036)

机构地区：[1]山西林业职业技术学院园艺系,山西太原030009 [2]南京农业大学园艺学院,江苏南京210095 [3]安徽农业大学园艺学院,安徽合肥230036

出　　处：《北方园艺》2018年第10期53-56,共4页Northern Horticulture

基　　金：国家自然科学基金资助项目(30200190);港澳台科技合作专项资助项目(2015DFT30090)

摘　　要：以"嘎啦"苹果组培苗叶片诱导的愈伤组织作为DNA模板提取材料,采用正交设计,摸索了SSR反应体系中Mg^(2+)、Taq DNA聚合酶、引物、模板DNA、dNTP的最适浓度,可为苹果愈伤组织的遗传变异研究奠定基础。结果表明:各因素不同水平变化对反应体系的影响为Taq DNA聚合酶>dNTP用量>Mg^(2+)用量>引物用量=模板DNA浓度。确立了苹果愈伤组织SSR-PCR最佳反应体系总体积25μL,其中4μL 25mmol·L^(-1) Mg^(2+),0.25μL 5U·μL^(-1) Taq DNA聚合酶,3μL 0.01mmol·L^(-1) SSR引物,1μL 30ng·μL^(-1)模板DNA,4μL 2.5mmol·L^(-1)dNTP,2.5μL 10×PCR Buffer,10.25μL超纯水。In order to establish the basis of genetic variation research of apple callus,the ‘Gala＇induction of callus was DNA template extraction materials.the Mg^2＋,Taq DNA polymerase,primer dosage,template DNA concentration and dNTP which were suitable for concentration or dosage were explored.The results showed that the effect of different levels on reaction system was Taq DNA polymerased〉NTP dosage〉Mg^2＋dosage〉primer dosage=template DNA concentration.The optimal reaction system of apple callus was established,the optimal reaction system of apple callus was that25μL for total volume,which were 4μL 25 mmol·L^-1 Mg-（2＋）,0.25μL 5 U ·μL^-1 Taq DNA polymerase,3μL 0.01 mmol·L^-1 SSR,1μL 30 ng·μL^-1 template DNA,4μL 2.5 mmol·L^-1 dNTP,2.5μL 10×PCR Buffer,10.25μL ultra-pure water.

关 键 词：苹果 愈伤组织 SSR-PCR 体系优化 

分 类 号：S661.103.6[农业科学—果树学]