库尔勒香梨mRNA差异显示技术体系的建立  

Establishing the System of Korla Fragrant Pear mRNA Differential Display Technology

作　　者：张飞[1] 牛建新[1] 叶春秀[1] 刘娜[1] 高静涛[1] 朱天丁[1] 

机构地区：[1]石河子大学农学院园艺系,石河子832003

出　　处：《石河子大学学报（自然科学版）》2011年第2期173-178,共6页Journal of Shihezi University(Natural Science)

基　　金：国家科技攻关计划引导项目(2003BA546C);国际科技合作(2009YD32);新疆兵团"科技人员服务南疆专项"(2010GG59)

摘　　要：为了研究库尔勒香梨萼片脱落与宿存的本质,试验以库尔勒香梨盛花前期的同一时间同一棵树同一花序的第2位序花和第4序位花为试验材料,采用mRNA差异显示的方法进行研究。扩增体系为20μL:cDNA 2.0μL,10μmol/L锚定引物2.0μL,10μmol/L随机引物2.0μL,2.5 mmol/L dNTPs 2.0μL,5 U/μL Taq 0.25μL,10×PCR buffer 2.5μL,ddH2O 9.25μL。结果显示,扩增产物经6%的非变性聚丙烯酰胺凝胶电泳、银染,获得了比较清晰的差异条带,回收并克隆差异条带,分离得到一些差异表达的片段,优化了库尔勒香梨mRNA差异显示技术,为克隆差异表达基因的全长cDNA奠定了良好基础。In this study,DDRT-PCR was used to study the nature of calyx leaving and persistent from fruit of Korla fragrant pear.We chose the flowers of early full bloom,and total RNA was extracted from the second flowers and the fourth flowers of the same inflorescence of the same tree at the same time.PCR reactions（20 μL） were performed with 2.0 μL RT reaction first strand,10 μmol/L anchor primer 2.0 μL,10 μmol/L arbitrary primer 2.0 μL,2.5 mmol/L dNTPs 2.0 μL,5 U/μL Taq DNA polymerase 0.25 μL,10×PCR buffer 2.5 μL,ddH2O 9.25 μL.The PCR product was confirmed by 6% non-denaturing polyacrylamide gel electrophoresis followed by silver staining,access to a relatively clear-cut difference between bands,and then recovering differential bands and cloning,preliminary isolating and getting some of differential genes between the second flowers and the fourth flowers.Finally,the Korla fragrant pear mRNA differential display was established,so this research laid a foundation for cloning full length of relative cDNAs.

关 键 词：库尔勒香梨 萼片 体系 MRNA差异显示 

分 类 号：S661.23[农业科学—果树学] Q781[农业科学—园艺学]