梅花鹿VEGF基因的克隆及其在鹿茸顶端组织的表达

Cloning of Vascular Endothelial Cell Growth Factor Gene and Its Expression in Antler Tissue

为了得到VEGF成熟肽基因,利用Trizol试剂法提取鹿茸顶端组织总RNA,反转录形成c DNA。根据Gen Bank已发表的相关基因序列设计1对特异性引物,并克隆VEGF成熟肽基因,再利用免疫组化法检测VEGF基因在鹿茸顶端不同组织的表达水平。结果表明:成功获得梅花鹿VEGF成熟肽基因,该基因长495 bp,共编码164个氨基酸;经Blast同源性分析,结果显示与牛、羊、猪的核苷酸序列的同源性分别为98.79%、97.98%、96.36%;经DNAMAN软件比对,其氨基酸序列的同源性分别为98.78%、97.56%、96.95%。免疫组化法结果显示,VEGF在茸皮层、间充质层和软骨层中均有表达,其中在软骨层中表达水平较高。

To obtain vascular endothelial cell growth factor（ VEGF） mature peptide gene,total RNA was isolated from growing antler tip tissue. A pair of specific primers was designed according to homologous gene sequence of VEGF among other species. c DNA sequence of VEGF gene was cloned by RT-PCR technology. Enzyme immunohistochemistry was used to detect expression level of VEGF in antler tip. Results showed that VEGF mature peptide gene was successfully obtained. It had 495 base pairs in length,encoding a predicted protein of 164 amino acids. Bioinformatic analysis based on VEGF genes of cattle,sheep and pig showed that homologies of VEGF were 98. 79%,97. 98%and 96. 36%,and homologies of protein were 98. 78%,97. 56% and 96. 95%,respectively. Results of enzyme immunohistochemistry showed that VEGF was expressed in cortex,mesenchyme and cartilage layers,and its expression level in cartilage layer was higher.