Mining of EST-SSR in sweetpotato and its optimization of PCR system
In this study,20 EST-SSR(expressed sequence tag-simple sequence repeat)primer pairs were designed based on sweetpotato cDNA sequences in public database.The genomic DNAs were isolated from two sweetpotato cultivars,one(named asXu781)of them is highly resistant to Ditylenchus destructor,and another(Xu 18)is highly susceptible to Ditylenchus destructor.The designed primers were screened against two DNAs.The PCR reaction components and reaction conditions were optimized for DNA amplification.An optimized PCR system was established as following：20μL reaction solution contained PCR 10× buffer 2.0μl,25 mmol/I MgCl2 1.5μl,10 mmol/I dNTP 0.2μl,7.5μmol/I SSR primers 0.5μl,150ng DNA template,5U/μl Taq polymerase 0.2μl,and ddH2O up to terminal volume.The PCR condition was initially run 5 min at 95℃,followed by 35 cycles of 30s at 95℃,60s at 59℃ and 60s at 72℃,and then extended 10 min at 72℃.The polyacrylamide gel electrophoresis and silver staining were employed for detecting and scoring the banding patterns.Specific bands amplified by 10 EST-SSR primers were presented in resistant cultivar,but absent in susceptible one using optimized PCR system which would be a potential tool to identify cultivars resistance to Ditylenchus Destructor in sweetpotato.
Sweetpotato EST-SSR PCR system Optimization Primer pair
Lin Jiang Lili Xu Yu Wang
School of Life Science,Anhui University,Anhui Key Laboratory of Eco-engineering and Bio-technique,Hefei 230039,China
国际会议
3rd China-Japan-Korea Workshop on Sweetpotato(第三届中日韩甘薯学术研讨会)
北京
英文
372-378
2008-10-12（万方平台首次上网日期，不代表论文的发表时间）