南方农业学报2013，Vol.44Issue(6)：898-902,5.DOI:10.3969/j:issn.2095-1191.2013.6.898
苦瓜抗白粉病SRAP标记筛选的PCR体系优化与验证
Optimizing and verification of PCR system for SRAP marker screening related to powdery disease resistance in bitter melon (Momordica charantia)
摘要
Abstract
[Objective]The present study was conducted to optimize the PCR system of SRAP molecular marker related to powdery disease resistance in bitter melon for providing reference on breeding new bitter melon cultivars resistant to powdery disease.[Method]On the basis of the known SRAP-PCR amplification procedure,the concentration combinations of DNA template,dNTPs,Mg2+,primer,Taq DNA polymerase were optimized and verified by orthogonal design L16 (45) with the genome DNA of powdery disease-resistant and-susceptible parents used as templates and four pair primers.[Result]The extrated bitter melon genomic DNA showed better purity and integrity,and had no pollution for protein and RNA,etc.The primer pair ME9/EM11 was used for PCR amplification and got obvious difference in 16 PCR systems of genomic DNA in parents MC18 and MC1-2.Among 16 PCR systems,the 3th system was the optimum system of SRAP-PCR for getting clear and more amplified bands,which contained DNA 5 ng/μL,dNTPs 0.05 mmol/L,Mg2+ 2.5 mmol/L,primer 0.6 μmol/L,Taq DNA polymerase 0.088 U/μL,in total 20 μL.Four primer pairs were used to verify the optimized SRAP-PCR system for different bitter melon materials,It showed good repetition and no polymorphism.[Conclusion]The optimized SRAP-PCR system was suitable for screening SRAP marker related to powdery disease resistance in bitter melon.关键词
苦瓜/SRAP-PCR/体系优化/筛选/验证Key words
bitter melon/SRAP-PCR/system optimization/orthogonal design/screening/verification分类
农业科技引用本文复制引用
米军红,尚小红,周生茂,车江旅,黄如葵,梁任繁,文俊丽,郭元元,梁和..苦瓜抗白粉病SRAP标记筛选的PCR体系优化与验证[J].南方农业学报,2013,44(6):898-902,5.基金项目
国家自然科学基金项目(31240060) （31240060）
广西科学研究与技术开发计划项目(桂科攻10100005-6) （桂科攻10100005-6）
广西自然科学基金项目(2012GXNSFAA053061) （2012GXNSFAA053061）
广西农业科学院基本科研业务专项项目(桂农科2012YT05) （桂农科2012YT05）
广西农业科学院公益性维持费项目(桂农科2012GW04) （桂农科2012GW04）
