东北马鹿鹿茸软骨组织cDNA文库构建及IGF2基因克隆与结构分析  

cDNA Library Construction from Antler Cartilage of the Red Deer (Cervus elaphus) and Cloning Insulin-like Growth Factor II Gene

作　　者：刘学东[1] 张子栋[1] 郑冬[1] 马建章[1] 

机构地区：[1]东北林业大学野生动物资源学院动物遗传学与分子生物学实验室,哈尔滨150040

出　　处：《中国农业科学》2008年第9期2806-2812,共7页Scientia Agricultura Sinica

基　　金：黑龙江省科技攻关项目资助(GC03B514);黑龙江省留学回国基金资助(LC06C36)

摘　　要：【目的】从功能基因组学角度深入研究鹿茸软骨组织生长发育机理,探讨胰岛素样生长因子-II(IGF2)在鹿茸软骨组织生长发育过程中的作用。【方法】利用SMART技术构建了东北马鹿鹿茸软骨组织全长cDNA文库,并设计IGF2基因保守引物对该文库进行筛选。【结果】构建的未扩增文库滴度为1.8×106pfu·ml-1,文库重组率大于89.8%;插入片段平均长度约为1.2 kb;扩增文库滴度为1.4×1010pfu·ml-1;克隆东北马鹿IGF2基因(GenBank登录号:EF177491)CDS区为540 bp,编码179 aa长度的多肽,与GenBank中检索得到的猪、马、牛、羊、人、小鼠和大鼠IGF2基因进行比对分析显示,马鹿IGF2多肽无色氨酸(Trp),组氨酸(His)与苏氨酸(Thr)含量在8个物种中最高;重建系统树结果显示,东北马鹿IGF2与牛和羊IGF2基因具有最高的相似性。【结论】成功构建东北马鹿鹿茸软骨组织cDNA文库并克隆到IGF2基因。[Objective] To investigate molecular mechanisms of ander cartilage growth and development at the functional genornic level and further clarify fimction of the Insufin-like growth factor Ⅱ （IGF2） gene. [Method] The SMART technique was used to construct a cDNA library from antler cartilage of the red deer （Cervus elaphus） and conserved primers were further designed to screen cDNA library and isolate IGF2 gene. [Result] The primary cDNA library contained 1.8X 106 independent clones, 89.8% of which were recombinant; titer of the amplified library was 1.4×10^l0 pfu·ml^-1 and average length of exogenous inserts was 1.2 kb. Bioinformatic analysis based on IGF2 genes of pig, horse, cattle, sheep, human, mouse and rat showed CDS region of the red deer IGF2 gene （GenBank accession No： EF177491） was 540 bp, which encoded a 179-amino-acids-long peptide. Compared with other seven species, IGF2 of the red deer had no tryptophan, and contained the most histidine and threonine. Moreover, phylogenetic tree reconstructed disclosed the highest similarities among IGF2 of deer, cattle and sheep. [Conclusion] The results demonstrated that the reliability and representativeness of the cDNA library were constructed, which would provide a further platform for studying regeneration mechanisms of antler cartilage of the red deer.

关 键 词：马鹿 鹿茸软骨组织 SMART CDNA文库 胰岛素样生长因子-2 

分 类 号：S865.4[农业科学—野生动物驯养]