山羊MMP-9基因cDNA的克隆及生物信息学分析

Cloning and bioinformatics analysis of cDNA for the MMP-9 gene in dairy goat

为了克隆山羊基质金属蛋白酶-9(MMP-9)基因的cDNA片段,分析其基因序列,试验于无菌条件下采集泌乳期奶山羊乳腺组织并提取总RNA。根据已知其他物种的MMP-9基因保守性区域设计特异性引物,采用RT-PCR方法扩增MMP-9基因的CDS区,测序后对核苷酸和推导的氨基酸序列进行分析,并对8个物种进行聚类分析。结果表明:得到长度为2 245 bp的MMP-9基因cD-NA片段(GenBank登录号为JQ670877),其中包含2 130 bp的CDS全长;核酸序列分析结果显示,该基因编码709个氨基酸,与人、小鼠和牛的核苷酸序列进行比对,相似性分别为84%、80%、96%,氨基酸相似性分别为79%、73%、94%。

To clone the eDNA fragments of the matrix metalloproteinase - 9 （ MMP - 9） gene of dairy goat and to analyze its gene sequences, the mammary gland tissues of dairy goat during lactation were collected to extract the total RNA under sterile conditions. A pair of specific primers were designed based on the conserved regions of MMP -9 gene of other known species, and the RT - PCR was performed to amplify the complete CDS sequences of MMP -9 from dairy goat. The analyses were performed including the nucleotide and deduced amino acid sequence analysis after sequencing, and the cluster analysis for the eight species. The results showed that a cDNA fragment of 2 245 bp for the MMP -9 gene was obtained, and its GenBank accession number was JQ670877, which contained a full -length CDS with a length of 2 130 bp. The nucleic acid sequence analysis revealed that the gene encoded for 709 amino acids. Compared with the nucleotide sequences of human, mouse and cattle, the similarities of the cDNA nucleotide sequences of goat MMP - 9 genes were determined as 84%, 80% and 96%, respectively, and the similarities of its amino acid sequence compared with that of human, mouse and cattle were determined as 79%, 73% and 94%, respectively.