玉米弯孢霉叶斑病菌均一化全长cDNA文库构建与鉴定  

Construction and characterization of a normalized full-length cDNA library of Curvularia lunata

作　　者：刘铜[1,2] 刘力行[1] 刘志诚[1] 侯巨梅[2] 高士刚[1] 周斐红[1] 陈捷[1] 

机构地区：[1]上海交通大学农业与生物学院农业部都市农业(南方)重点开放实验室,上海200240 [2]黑龙江八一农垦大学农学院,大庆163319

出　　处：《植物病理学报》2010年第3期250-257,共8页Acta Phytopathologica Sinica

基　　金：国家重点基础研究发展计划"973"项目(2006CB101901);国家科技支撑课题(2006BAD08A06);国家自然科学基金资助项目(30871610)

摘　　要：利用DSN(duplex-specific nuclease)均一化与SMART(switching mechanism at5′end of RNA transcript)技术构建玉米弯孢霉叶斑病菌菌丝的均一化全长cDNA文库。经检测原始文库的滴度为1.4×106pfu/mL,重组率为96.9%,插入片段平均长度大于1.0 kb。从文库中随机挑取192个cDNA克隆测序,获得173个高质量的EST序列,其中有5个跨叠群(conting),168个独立克隆,全长cDNA克隆占57%。BLASTx分析表明,有18.5%EST与GenBank中无任何同源性,有81.5%EST与GenBank中报道的功能已知或未知蛋白具有相似性。A normalized full-length cDNA library was constructed with the mycelium of Curvularia lunata,a causal organism of maize leaf spot,by DSN duplex-specific nuclease normalization method combined with SMART（switching mechanism at 5′ end of RNA transcript） technique.This library was characterized by a plaque titer of 1.4×106 pfu/ mL and a 96.9% recombination ratio,of which the fragment length of inserted average cDNA sequences was above 1.0 kb.The 173 high quality expressed sequence tags（EST） including 5 coting and 168 singlet was obtained from 192 cDNA clones randomly picked.Based on bioinformatics analysis,the full-length cDNA accounted for 57% of total sequences.BLASTx analysis revealed that 81.5% of the EST displayed significant homology to known or unknown genes from GenBank database.

关 键 词：均一化 CDNA文库 弯孢霉 菌丝 

分 类 号：S435.131[农业科学—农业昆虫与害虫防治]