CAJ | 학술논문

为了建立柿属植物目标区域扩增多态性（ TRAP）标记技术体系，对影响TRAP-PCR的Mg2＋、dNTPs、Taq DNA聚合酶、固定引物和随机引物浓度比5个因素进行了优化。确定优化的反应体系为：模板DNA 40 ng，buffer 1×，Mg2＋1．4 mmol/L，dNTPs 0．2 mmol/L，固定引物和随机引物浓度比为11∶1，Taq 酶0．5 U，反应总体积为15μL。利用柿属植物EST数据库信息设计锚定引物10条，与11条随机引物组合共110对引物。利用这些引物对柿属植物6个基因型进行TRAP-PCR扩增，其中84个引物组合能扩增出清晰条带，占引物总量的76．36％；36个引物组合表现出良好的多态性扩增，并在20份柿属植物中进行了引物验证。
위료건립시속식물목표구역확증다태성（ TRAP）표기기술체계，대영향TRAP-PCR적Mg2＋、dNTPs、Taq DNA취합매、고정인물화수궤인물농도비5개인소진행료우화。학정우화적반응체계위：모판DNA 40 ng，buffer 1×，Mg2＋1．4 mmol/L，dNTPs 0．2 mmol/L，고정인물화수궤인물농도비위11∶1，Taq 매0．5 U，반응총체적위15μL。이용시속식물EST수거고신식설계묘정인물10조，여11조수궤인물조합공110대인물。이용저사인물대시속식물6개기인형진행TRAP-PCR확증，기중84개인물조합능확증출청석조대，점인물총량적76．36％；36개인물조합표현출량호적다태성확증，병재20빈시속식물중진행료인물험증。
The concentrations of Mg 2+, dNTPs, Taq DNA polymerase and primers which affected TRAP ( target region ampli-fied polymorphism)-PCR reaction were optimized for the establishment of TRAP molecular marker system in Diospyros spp.The optimum reaction system was acquired as follows:template DNA 40 ng, buffer 1 ×, Mg2+1.4 mmol/L, dNTPs 0.2 mmol/L, fixed primer-arbitrary primer concentration ratio 11∶1, Taq DNA polymerase 0.5 U, and total volume of reaction system 15μL.Eighty-four TRAP primer combinations which showed robust PCR amplification in 6 persimmon genotypes were screened out from 110 primer combinations (10 fixed primers times 11 arbitrary primers).Thirty-six polymorphic primer combinations were verified in 20 persimmon genotypes .