仿刺参纤维胶凝蛋白基因的分子特征及表达分析

摘要

纤维胶凝蛋白是非特异性免疫系统中的重要分子.通过转录组测序及cDNA末端快速克隆技术得到一条仿刺参纤维胶凝蛋白基因的全长cDNA序列,并将其编码的蛋白命名为仿刺参纤维胶凝蛋白-1.获得的基因cDNA全长为1951 bp,其中5′-末端非翻译区为397 bp,3′-末端非翻译区为666 bp,开放阅读框为888 bp,编码295个氨基酸,N端16个氨基酸为信号肽,信号肽后面有两个G-X-Y重复序列,C端为纤维蛋白素原结构域.采用实时荧光定量PCR方法分析了仿刺参纤维胶凝蛋白-1基因在仿刺参幼参不同组织及细菌脂多糖刺激后的时序表达规律,结果显示仿刺参纤维胶凝蛋白-1基因在仿刺参的肠道、呼吸树、体腔细胞和体壁均有表达,且肠道的表达量最高;脂多糖刺激后,4种组织的仿刺参纤维胶凝蛋白-1基因表达量均有变化,且以肠道和体壁表达量的变化最为显著;此外,仿刺参纤维胶凝蛋白-1基因在仿刺参4种组织中的表达量变化具有不同的时序性,表明仿刺参纤维胶凝蛋白-1可能在仿刺参免疫应答中起重要作用.%We identified the cDNA sequence of ficolin gene from sea cucumber (Apostichopus japonicus) through RNA-seq sequencing and RACE (rapid amplification of cDNA ends) technology, and the protein known as AjFCN had length of 1951 bp including a 397 bp 5′ untranslated region (UTR), a 666 bp 3′-UTR and an ORF of 888 bp encoding 295 amino acids.The AjFCN protein contained a typical signal peptide, two G-X-Y repetitive sequences and a brinogen-like domain (FBG).The relative expression of AjFCN gene was detected by the qRT-PCR in the intestine, respiratory tree, coelomocyte and body wall, with the maximal expression level in the intestine.After lipopolysaccharide (LPS) challenge, the expressions in intestine and body wall tissues were changed most dramatically.Furthermore, the relative expression levels of AjFCN gene in the four tissues showed different time-sequence patterns.The findings indicated that the AjFCN played an important role in the immune response of sea cucumber.