甜瓜cDNA-AFLP分析体系的建立  

Establishiment of cDNA-AFLP Analysis System for Melon

作　　者：赵惠新[1] 唐亚萍[1] 祝长青[1] 覃建兵[1] 

机构地区：[1]新疆师范大学分子生物学与生物信息研究室,乌鲁木齐830053

出　　处：《种子》2009年第11期45-48,共4页Seed

基　　金：新疆维吾尔自治区重大科技专项科技支疆项目(20084010249);国家自然科学基金(30960044;30960092)

摘　　要：选用新疆甜瓜伽师和皇后为试材,研究了影响cDNA-AFLP反应体系的几个关键因素,建立了适合于甜瓜的cDNA-AFLP反应体系。结果表明:适用于甜瓜cDNA-AFLP分析的RNA可以用Trizol试剂提取的总RNA,双链cDNA合成应用置换法,双链酶切用VspⅠ、TaqⅠ和EcoRⅠ、MseⅠ,效果均能达到目标。相比较而言,VspⅠ、TaqⅠ酶切组合更为理想。酶切cDNA用量、连接产物、预扩增产物稀释倍数参照常规AFLP技术中的常用即可满足要求。检测采用8%的非变性聚丙烯酰胺凝胶电泳、银染显色,操作较文献介绍的变性胶方法简单、方便。实验得到了较为理想的甜瓜mRNA指纹图谱,建立的反应体系可以应用于甜瓜差异基因表达分析等方面的研究。The key factors affecting cDNA-AFLP analysis system of cucumis melon was studied, and an optimized system was established which showed a distinct electrophoretogram of cDNA-AFLP. The result indicated that the total RNA from melon using Trizol reagent was good for this study and replacement was sit for synthesizing dsCDNA. The two pairs restriction enzyme combinations were good for melon both although EcoR I and Mse I was better. The dosage of dscDNA was 300 ng, the pre-amplification procducts using template of selectamplification were diluted to 30 times and else were perfect for melon. Otherwise, 8 % no-denaturalization poly- propylene gel for electrophoresis and display with siller were simpler and more facility than denaturalization polypropylene gel that literatures shown.

关 键 词：甜瓜 cDNA-AFLP体系 影响因素 

分 类 号：S562[农业科学—作物学] Q503[生物学—生物化学]