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[目的]进行鲤鱼白细胞介素-1β(IL-1β)全长cDNA的克隆、鉴定及其差异表达分析。[方法]利用DD-RTPCR方法获得差异表达cDNA片段,对有丝分裂原刺激的鲤鱼外周血白细胞cDNA文库进行筛选,克隆了鲤鱼IL-1β的全长cDNA,并进行了序列分析和差异表达分析。[结果]获得的阳性克隆含有1个大小为831bp编码276个氨基酸的完整开放阅读框。聚类分析表明,鲤鱼IL-1β氨基酸序列与日本鲤鱼紧密聚为一支,氨基酸序列的同源性达95%,之后聚类顺序依次为鲫鱼、斑马鱼、猪、牛、马、人和小鼠。差异表达分析表明,经有丝分裂原刺激后前期(2h)白细胞中IL-1β的表达量显著增大,但随着时间推移(12,24h)并非一直较同期大,表达量总体趋势成峰形。[结论]为进一步研究IL-1β在体内的表达方式、功能特点和调控机理以及在炎症反应、应急反应和免疫应答的作用机制奠定了基础。
[Objective] The research aimed to clone, identify and differentiate the full-length cDNA of IL-1β in common carp. [Method] The differentially expressed cDNA fragments were obtained by DD-RTPCR method. The mitochondria-stimulated carp peripheral blood leucocyte cDNA library was screened. The full-length cDNA of IL-1β in common carp was cloned and sequenced and differentially expressed. [Result] The positive clone obtained contained a complete open reading frame with a size of 831bp encoding 276 amino acids. The results of cluster analysis showed that the amino acid sequence of IL-1β was closely related to Japanese carp, and the homology of the amino acid sequence was 95%. The order of clustering was carp, zebra fish, pig, cow, horse, human and small mouse. The differential expression analysis showed that the expression of IL-1β in leukocytes stimulated by mitogen was significantly increased, but it was not always higher than that in the same period (12,24h). The overall trend of expression was peaked. [Conclusion] This study lays the foundation for further study on the expression, function and regulation of IL-1β in vivo and the mechanism of action in response to inflammation, emergency and immune response.