梅花鹿Smad2与Smad4基因的克隆及其在鹿茸顶端组织的表达  

Cloning of Smad2 and Smad4 genes from sika deer and their expression in antler tissue

作　　者：刘洪运 韩香玉[1] 刘明晓[1] 胡薇[1] LIU Hongyun;HAN Xiangyu;LIU Mingxiao;HU Wei(College of Life Sciences,Jilin Agricultural University,Changchun 130118,China)

机构地区：[1]吉林农业大学生命科学学院,长春130118

出　　处：《黑龙江畜牧兽医》2019年第3期6-10,173,共6页Heilongjiang Animal Science And veterinary Medicine

基　　金：国家自然科学基金项目(31572372;30972083)

摘　　要：为了从梅花鹿鹿茸顶端组织中克隆Smad2与Smad4基因的编码区(CDS)序列,分析其分子特性及在鹿茸顶端不同组织中的表达情况,试验采用TRIzol法提取鹿茸顶端组织总RNA,以PCR方法获得Smad2与Smad4基因,并利用生物信息学软件对其进行生物信息学分析,免疫组化法检测Smad2与Smad4基因在鹿茸顶端不同组织中的表达水平。结果表明:Smad2基因完整的CDS序列长度为1 404 bp,编码467个氨基酸,与牛、人、马和猪的Smad2核苷酸序列同源性分别为98.29%、94.52%、95.30%和95.51%;Smad4基因完整的CDS序列长度为1 662 bp,编码553个氨基酸,与牛、人、马和猪的Smad4核苷酸序列同源性分别为98.26%、94.89%、95.85%和95.97%;Smad2与Smad4蛋白的分子质量分别为52.24 ku与60.50 ku,理论等电点分别为6.13与6.50,均具有较强的亲水性;梅花鹿Smad2与Smad4基因在茸皮层、间充质层和软骨层中均有表达,在软骨层中表达水平较高。说明梅花鹿Smad2与Smad4基因在鹿茸软骨层中表达水平较高,对鹿茸再生发育具有一定的调节作用。In this study,we extracted total RNA in antler tip by improved TRIzol method and got the Smad2 and Smad4 gene of sika deer antlers by PCR method.Bioinformatics software was used to analyze the sequence and structure of the Smad2 and Smad4 gene. The immunohistochemical method was used to determine the gene expression of Smad2 and Smad4 in different tissues of antler tissue.The results showed that the complete coding region of Smad2 gene ( CDS) sequence was 1 404 bp and encoding 467 amino acids, sequence homology of the nucleotide were 98. 29%、 94. 52%、95. 30% and 95. 51% comparing with cattle,human,horse and pig,respectively.The complete coding region of Smad4 gene ( CDS) sequence was 1 662 bp and encoding 553 amino acids,sequence homology of the nucleotide were 98. 26%、94. 89%、95. 85% and 95. 97% comparing with cattle,human,horse and pig,respectively. Physical and chemical properties analysis found that the molecular weight of Smad2 and Smad4 protein were 52. 24 ku and 60. 50 ku,the theoretical PI were 6. 13 and 6. 50,respectively,both have strong hydrophilicity.The Smad2 and Smad4 protein were expressed in the top of antler tissue,including skin layer mesenchymal layer and cartilage layer,and it was higher expressed in the cartilage layer.It indicates that the expression of Smad2 and Smad4 gene of cartilage layer in the top of antler are relatively high, and it plays an important role in antler growth process.

关 键 词：梅花鹿 SMAD蛋白 免疫组化 克隆 序列分析 

分 类 号：S825[农业科学—畜牧学] Q785[农业科学—畜牧兽医]