梅花鹿鹿茸软骨细胞的培养及X型胶原的克隆和序列分析

型胶原是肥大软骨细胞的标志物,据GenBank中收录的人、鼠、牛、猪型胶原基因的序列,进行同源性分析,在同源性高的相对保守区域设计了1对特异性引物。建立梅花鹿鹿茸软骨细胞的分离培养方法,培养鹿茸软骨细胞。提取细胞的总RNA,以总RNA为模板,用RT-PCR方法克隆了梅花鹿的型胶原基因序列为877的片段,此片段全部位于编码区,与已报道的牛的型胶原基因的序列比较,同源性达到96%。共有35个碱基发生变异。DNAStar软件分析表明,二者推导的氨基酸序列同源性为95.5%。

The Culture of Sika Deer Antler Cartilage Cells and Cloning of the collagen X from Antler Cartilage

The collagen X protein is the classical marker of hypertrophic chondrocytes. The homology was analyzed based on the reported collagen X sequence of human, mouse, cow and pig in Gen Bank. A pair of specific primers was designed in the conservative relatively sectors. The methods of isolation and culture sika deer antler chondrocytes were established. Sika deer antler chondrocytes were cultured, total RNA was extracted from the cultured cells. The sequence encoding deer antler collagen X was cloned from the cartilage cells of sika deer antler by means of the specific primers with the methods of RT-PCR. The cloned was 877 bp with an ORF. This sequence encoded 292 amino acids. The homology of the nucleotide sequence of sika deer collagen X with cow was 96%, and the homology of deduced amino acids between them was 95.5%.