红笛鲷LITAF基因的克隆与表达分析

通过同源克隆和RACE PCR技术从红笛鲷脾脏中鉴定得到了脂多糖诱导的肿瘤坏死因子(LITAF)基因,命名为Ls-LITAF。该基因c DNA全长815 bp,开放阅读框为447 bp,编码148个氨基酸,理论分子量为15.7 ku。序列分析显示Ls-LITAF蛋白具有保守的LITAF结构域,其中包含一个CXXC基序与一个(H)x Cxx C基序。Ls-LITAF蛋白与其他鱼类LITAF蛋白相似度较高,在系统进化树中也与其他鱼类该蛋白聚为一支。Ls-LITAF基因在健康鱼体多种组织均有表达,其中鳃、脾脏、皮肤与头肾的表达量较高;且哈氏弧菌刺激鱼体后,Ls-LITAF在脾脏和头肾中的表达量显著上调。以上研究结果为进一步了解Ls-LITAF在红笛鲷免疫反应中的作用提供了参考。

Molecular cloning and expression analysis of LITAF in Lutjanus sanguineus

The lipopolysaccharide-induced TNF factor (LITAF) gene was identified from humphead snapper (Lutjanus sanguineus) spleen using homologous cloning and RACE PCR, designated as Ls-LITAF. The full length cDNA of Ls-LITAF was 815 bp, contained an ORF of 447bp encoding a polypeptide of 148 amino acids with predicted molecular mass of 15.7kD. Sequence alignment showed that Ls-LITAF possessed a typical LITAF domain with a CXXC and (H)xCxxC motif, shared high similarities with other fish LITAF proteins. Phylogenetic analysis indicated that Ls-LITAF clustered closely with known fish LITAFs. Ls-LITAF distributed in all analyzed tissues with higher expressions in gill, spleen, skin and head kidney. After Vibrio harveyi infection, the expression levels of Ls-LITAF were up-regulated significantly in spleen and head kidney. These results provided a reference to further investigate the roles of Ls-LITAF in immune response of Lutjanus sanguineus.