关键词: α甘露糖苷酶, 基因克隆, 荧光定量PCR, 生物信息学, 同源建模

Abstract: Goat lysosomal αAMA gene was amplified using RTPCR and the tissue specific expression profile, bioinformatics characteristic of αAMA were studied. Primers were designed based on the sequence of bovine lysosomal αAMA gene and were used in amplifying the goat αAMA, the tissue specific expression profile was analyzed by qRTPCR, and the bioinformatics analysis of αAMA was conducted. The results showed that CDS sequence of goat αAMA was 3 000 bp, encoding a deduced protein containing 999 amino acid residues in which the first 50 residues were signal peptide, and this nucleotide sequence of CDS and the deduced amino acids shared 95.93% and 94.79% homology with the αAMA mRNA of cattle. The qRTPCR revealed that the αAMA gene was expressed in various tissues at different levels. The expression level of this gene was higher in the lung, liver, and cerebellum. It was predicted that αAMA was belonged to Glycosyl hydrolases family 38, and composed of two conserved domains. There were thirtyfive phosphorylation sites, one phosphorylation site of specific protein kinase and nine Nglycosylation sites in αAMA protein. By SWISSMODEL homologous modeling, the predicted structure of goat was similar to the template and predictive. These results provide the theoretic basis for the mechanism research of αAMA and development of toxin antidote toward locoweed.

Key words: αmannosidase, gene clone, qRTPCR, bioinformatics, homology modelingα