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当前位置: 首页 > 文章 > 德保苏铁珊瑚根均一化全长cDNA文库的构建和EST分析 亚热带植物科学 2014,43 (2) 93-97
Position: Home > Articles > Construction of Normalized Full-length cDNA Library for the Coralloid Roots of Cycas debaoensis and Analysis of its Expressed Sequence Tags Subtropical Plant Science 2014,43 (2) 93-97

德保苏铁珊瑚根均一化全长cDNA文库的构建和EST分析

作  者：
王运华;陈庭;李楠
单  位：
深圳市中国科学院仙湖植物园深圳市南亚热带植物多样性重点实验室
关键词：
德保苏铁;珊瑚根;均一化;cDNA文库
摘  要：
以德保苏铁珊瑚根为材料,利用DSN(duplex-specific nuclease)均一化与SMART(switching mechanism at 5′end of RNA transcript)技术,构建均一化全长cDNA文库。经检测原始文库滴度为1.5×106 cfu·mL-1,文库重组率达97%,插入片段平均长度为1 kb。从文库中随机挑取192个cDNA克隆测序,获得175条高质量EST序列,装备成165条unique序列,其中包括7条contigs和158条singlets,EST冗余度为5.71%。将获得的174条干净的高质量EST序列与NCBI非冗余核酸库(NT)和非冗余蛋白库(NR)数据库进行Blast比对(分值≥200,一致性≥60),其中66.1%(115)的EST序列发现了与其同源的序列,而33.9%(59)的EST序列未发现与其显著同源的序列。构建德保苏铁珊瑚根均一化全长cDNA文库为功能性标记开发和功能基因研究提供了丰富的信息平台。
译  名：
Construction of Normalized Full-length cDNA Library for the Coralloid Roots of Cycas debaoensis and Analysis of its Expressed Sequence Tags
作  者：
WANG Yun-hua;CHEN Ting;LI Nan;Shenzhen Key Laboratory of Southern Subtropical Plant Diversity,Fairylake Botanical Garden,Shenzhen & Chinese Academy of Sciences;
关键词：
Cycas debaoensis;;coralloid root;;normalized;;cDNA library
摘  要：
A normalized full-length cDNA library was constructed with the coralloid roots of Cycas debaoensis by DSN(duplex-specific nuclease) normalization method combined with SMART(Switching Mechanism At 5′ end of the RNA Transcript) technique. The titer of original cDNA library was about 1.5×106 cfu·mL-1 and the average insertion size was about 1 kb with high recombination rate(97%). The 175 high quality expressed sequence tags(ESTs) were obtained from 192 randomly picked cDNA clones. Clusting and assembly of ESTs resulted in 165 unique sequences, consisting of 7 contigs and 158 singlets. Using a generic similarity threshold(score ≥ 200, identity ≥ 60), approximately 115(66.1%) of all 174 clean ESTs matched corresponding homologous sequences according to BLAST analysis of the nr-nt(non-redundant protein and non-redundant nucleotide sequence) database in GenBank. The 33.9%(59) ESTs have no significant match. These results showed preliminarily that we successfully constructed a normalized full-length cDNA library of coralloid roots in C. debaoensis, which could serve as an abundant information platform for functional marker development and functional gene study.

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