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.DOCVIP

东北马鹿鹿茸软骨组织cDNA文库构建及IGF2基因克隆×106 pfu·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1.4×1010 pfu·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。 关键词：马鹿；鹿茸软骨组织；SMART cDNA文库；胰岛素样生长因子-2 cDNA Library Construction from Antler Cartilage of the Red Deer (Cervus elaphus) and Cloning Insulin-like Growth Factor II Gene LIU Xue-dong, ZHANG Zi-dong, ZHENG Dong, MA Jian-zhang (Animal Genetics and Molecular Biology Laboratory, College of Wildlife Resources, Northeast Forestry University, Harbin 150040) Abstract: 【】【ethod】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. 【】 The primary cDNA library contained 1.8×106 independent clones, 89.8% of which were recombinant; titer of the amplified library was 1.4×10 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. 【】 Treliability 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. Key words: Red deer; Antler cartilage; SAMRT cDNA library; Insulin-like growth factorⅡ 0 引言 【研究意义】全世界共有鹿类47种，中国有16种，是鹿类资源最丰富的国家之一[1]。中国养鹿并利用鹿产品已有上千年的历史。东北马鹿Cervus elaphus）作为马鹿的一个亚种，因其鹿茸的品质优良和高产而蜚声国内外。再生指生物体以现存的细胞和组织重新生成或修复失去的组织或器官的过程[2]，鹿茸角是目前所知的惟一具有表形