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浙江大学学报（农业与生命科学版）
浙江大学学报(农业与生命科学版)  2011, Vol. 37 Issue (6): 603-609    DOI: 10.3785/j.issn.1008-9209.2011.06.003
生物科学与技术     
鮸鱼全长均一化cDNA 文库的构建及EST 序列信息学分析
徐田军, 孙悦娜, 石戈, 王日昕
浙江海洋学院海洋科学学院海洋生物资源与分子工程实验室,浙江舟山３１６０００
Establishment of a normalized full‐length cDNA library and bioinformatic analysis of ESTs of Miichthys miiuy
XU Tian‐jun ,SUN Yue‐na ,SHI Ge ,WANG Ri‐xin
Laboratory for Marine Living Resources and Molecular Engineering , College of Marine Science , Zhejiang Ocean University , Zhoushan , Zhejiang 316000 , China
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摘要： 采用双链特异性核酸酶(DSN)均一化技术与SMART 技术相结合,以经过鳗弧菌感染后获得的鮸鱼脾脏材料构建均一化全长cDNA 文库.该原始文库约含有７.５ × １０６个重组子,插入片段在１ ～ ３kb 之间,文库滴度为１.６ × １０６ CFU.mL － １ .从文库中随机挑选５９５２个克隆进行测序,共获得５０５３条表达序列标签( EST) ,其中高质量序列４６０９条.对序列进行组装后,发现非冗余基因序列３２２１条,重叠群６５６个,单拷贝序列２５６５条,冗余率为３０.１１％.对全部序列分析后发现,其平均读长为５５０bp ,大部分序列长度在５００ ～ ５９９ bp 之间,GC 含量大部分分布于４０％ ～ ６０％之间.１３９７个基因能够进行GO(geneontology) 功能注释,并初步筛选到大量与免疫功能相关的蛋白家族及功能域的基因序列,为进一步鮸鱼分子育种奠定了基础.
Abstract: To investigate molecular‐assisted breeding in M iichthys miiuy , a normalized full‐length cDNA library was established using the duplex‐specific nuclease (DSN) normalization method combined with switching mechanism at ５ end of RNA transcript (SMART) technique . The titer of un‐amplified cDNA library was about １.６ × １０６CFU.mL － １ ; the cDNA library contained ７.５ × １０６ independent clones with DNA inserts of １‐３ kb . A total of ５９５２clones were randomly selected to be sequenced and ５０５３expressed sequence tags ( EST) were obtained , and ４６０９ high quality EST sequences were generated .After processing , a total of ３２２１ unigenes comprising ６５６ contigs and ２５６５ singlets were obtained , and the redundancy ratio was about ３０.１１％. The average reading length was ５５０bp , and reading length of most sequences was ranged from ５００to ５９９bp and the GC contents of most sequences was ranged from４０to ６０percentages . A total of１３９７ functional known genes were categorized by gene ontology . Many putative immune genes were identified that encoded major histocompatibility complex , cytokines and cytokine receptors , adhesive proteins , stress proteins , transcription factors for immune response ,immunoglobulin and coagulation factors . The results above provide both a detailed annotation of immune genes and a collection of novel transcripts for this important species .
出版日期: 2011-11-25
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引用本文:

徐田军, 孙悦娜, 石戈, 王日昕. 鮸鱼全长均一化cDNA 文库的构建及EST 序列信息学分析[J]. 浙江大学学报(农业与生命科学版), 2011, 37(6): 603-609.

XU Tian‐jun,SUN Yue‐na,SHI Ge,WANG Ri‐xin. Establishment of a normalized full‐length cDNA library and bioinformatic analysis of ESTs of Miichthys miiuy. , 2011, 37(6): 603-609.

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http://www.zjujournals.com/agr/CN/10.3785/j.issn.1008-9209.2011.06.003    或    http://www.zjujournals.com/agr/CN/Y2011/V37/I6/603

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