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段亚飞, 刘萍, 李吉涛, 李健, 高保全, 陈萍. 2013: 脊尾白虾组织蛋白酶L基因的克隆及其表达分析. 动物学研究, 34(1): 39-46. DOI: 10.3724/SP.J.1141.2013.01039
引用本文: 段亚飞, 刘萍, 李吉涛, 李健, 高保全, 陈萍. 2013: 脊尾白虾组织蛋白酶L基因的克隆及其表达分析. 动物学研究, 34(1): 39-46. DOI: 10.3724/SP.J.1141.2013.01039
Ya-Fei DUAN, Ping LIU, Ji-Tao LI, Jian LI, Bao-Quan GAO, Ping CHEN. 2013. Cloning and expression analysis of Cathepsin L cDNA of Exopalaemon carinicauda. Zoological Research, 34(1): 39-46. DOI: 10.3724/SP.J.1141.2013.01039
Citation: Ya-Fei DUAN, Ping LIU, Ji-Tao LI, Jian LI, Bao-Quan GAO, Ping CHEN. 2013. Cloning and expression analysis of Cathepsin L cDNA of Exopalaemon carinicauda. Zoological Research, 34(1): 39-46. DOI: 10.3724/SP.J.1141.2013.01039

脊尾白虾组织蛋白酶L基因的克隆及其表达分析

Cloning and expression analysis of Cathepsin L cDNA of Exopalaemon carinicauda

  • 摘要: 根据本实验室构建的脊尾白虾(Exopalaemon carinicauda)血细胞全长cDNA文库获得的EST序列,利用RACE技术克隆获得脊尾白虾组织蛋白酶L基因的cDNA全长,命名为EcCatL基因。该序列全长1 136 bp,包括5'非编码区24 bp,开放阅读框960 bp和3'非编码区152 bp,开放阅读框共编码319个氨基酸,预测相对分子量为35.30×103,理论等电点为5.27。同源性分析表明,脊尾白虾组织蛋白酶L EcCatL氨基酸序列与其它甲壳动物高度保守,与变色小长臂虾(Palaemonetes varians)及北极甜虾(Pandalus borealis) CatL的同源性分别为92%和76%。系统进化分析表明,EcCatL基因氨基酸序列与变色小长臂虾的CatL聚为一支。荧光定量PCR分析结果表明,EcCatL基因在血细胞、鳃、肝胰腺、肌肉、卵巢、肠、胃及眼柄中均有表达,其中肝胰腺中的相对表达量最高。感染鳗弧菌及WSSV后6 h和12 h,脊尾白虾血细胞和肝胰腺中EcCatL的表达量较对照组均极显著增加(P<0.01),且具有明显的时间差异性,表明EcCatL基因在脊尾白虾免疫反应中具有重要作用。

     

    Abstract: Based on the EST sequence from a hemocyte cDNA library, the cathepsin L cDNA of Exopalaemon carinicauda (EcCatL) was cloned by rapid amplification of cDNA ends (RACE). The EcCatL cDNA was 1136 bp in length, which contains an open reading frame (ORF) of 960 bp, encoding a 319 amino-acid polypeptide. Homology analysis revealed that the amino acid sequence of EcCatL was highly conserved with its homologs in other crustaceans. The similarities of EcCatL with the CatL of Palaemonetes varians and Pandalus borealis were 92% and 76%, respectively. Phylogenetic analysis showed that EcCatL was in the same branch as that of Palaemonetes varians. The expression levels of EcCatL in different tissues were analyzed by quantitative real-time PCR. Expression of EcCatL was detected in all tested tissues of E. carinicauda, including hemocytes, gill, hepatopancreas, muscle, ovary, intestine, stomach and eyestalk, with the highest expression level in hepatopancreas. After challenged with Vibrio anguillarum or white spot syndrome virus, the expression of EcCatL were up-regulated in the hemocytes and hepatopancreas of E. carinicauda. Our results implied that EcCatL might play an important role in the prawn immune response.

     

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