小尾寒羊溶菌酶基因的克隆与差异表达分析

摘要

The male Small Tail Han sheep were inoculated with Brucella suis S2 vaccine strain, the suppression subtractive hybridization (SSH) library was constructed from buffy coat of sheep. After the recombinant plasmids were sequenced, the cDNA sequence of the conserved domain of lysozyme (LYZ) was recognized based on BLAST in GenBank, with the size of 770 bp, and continuously encoding 148 amino acid residuals, the encoded protein was a natural anti-infective substances. The sequence had been submitted to GenBank(JX263305). Quantitative Real-time PCR result showed that the LYZ gene was slightly up-regulated in buffy coat at 14 and 30 days post-challenge (dpc) without statistical significance (P>0. 05), and then recovered to the normal level at 40 dpc. The lysozyme was further proved as an essential natural immune factor which laid the foundation for the effective prevention and control of brucellosis.%本试验从猪种布鲁氏菌S2疫苗株免疫雄性小尾寒羊构建的白细胞层SSH cDNA文库中筛选1段cDNA序列,经测序和BLAST同源性搜索比对发现其为溶菌酶重组片段,该片段大小为770 bp,编码148个氨基酸残基,编码的蛋白质是-种天然抗感染物质,已提交GenBank,登录号为JX263305.经荧光定量PCR分析,与正常组羊相比,在S2免疫14和30 d的羊白细胞层中该溶菌酶基因表达轻微上调,但差异不显著(P＞0.05)；在免疫40 d时恢复到正常水平.这也进一步说明溶菌酶是一种存在机体内必不可少的天然免疫因子,为布鲁氏菌病有效防控方面的研究奠定了基础.