首页 >  广东海洋大学学报 >  凡纳滨对虾含酪氨酸酶结构域血蓝蛋白的免疫功能

2022, 42(5): 1-8. doi: 10.3969/j.issn.1673-9159.2022.05.001

凡纳滨对虾含酪氨酸酶结构域血蓝蛋白的免疫功能

1. 广东海洋大学食品科技学院/现代生化中心, 广东 湛江 524088;

2. 中山大学生命科学学院有害生物控制与资源利用国家重点实验室, 广东 广州 510275;

3. 南方海洋科学与工程广东省实验室(珠海), 广东 珠海 519000;

4. 广东海洋大学深圳研究院, 广东 深圳 518108

收稿日期:2022-06-08
修回日期:2022-06-08

基金项目:   广东省基础与应用基础研究基金项目(2021A1515110900);中山大学中央高校基本科研业务费专项资金(22qntd2615) 

关键词: 凡纳滨对虾 , 含酪氨酸酶结构域血蓝蛋白 , 免疫刺激 , 副溶血弧菌 , 免疫应答

Immune Function of Tyrosinase-Domain Containing Hemocyanin(TDCH)from Litopenaeus vannamei

1. College of Food Science and Technology/Modern Biochemistry Experimental Center, Guangdong Ocean University, Zhanjiang 524088, China;

2. State Key Laboratory of Biocontrol, School of Life Sciences, Sun Yat-Sen University, Guangzhou 510275, China;

3. Southern Marine Science and Engineering Guangdong Laboratory(Zhuhai), Zhuhai 519000, China;

4. Shenzhen Institute of Guangdong Ocean University, Shenzhen 518108, China

Received Date:2022-06-08
Accepted Date:2022-06-08

Keywords: Litopenaeus vannamei , tyrosinase-domain containing hemocyanin , immune stimulation , Vibrio parahaemolyticus , immune response

摘要

【目的】克隆凡纳滨对虾(Litopenaeus vannamei)一种新的含酪氨酸酶结构域血蓝蛋白(Tyrosinase-domain containing hemocyanin,TDCH),研究其免疫功能。【方法】根据NCBI上公布的tdch基因序列设计克隆引物,通过RT-PCR方法获得tdch的编码区;采用生物信息学方法分析tdch的结构特征;使用荧光定量PCR技术分析tdch在凡纳滨对虾12 个组织中的表达,以及在副溶血弧菌(Vibrio parahaemolyticus)、金黄色葡萄球菌(Staphhylococcu saureus)、黑曲霉菌(Aspergillus niger)、白斑综合征病毒、脂多糖及聚肌苷酸-聚胞苷酸[Poly(I:C)]刺激后在鳃和血细胞中的表达变化;利用RNAi 技术敲降tdch 在对虾中的表达,研究其在对虾抗副溶血弧菌免疫中的作用。【结果与结论】克隆获得tdch 的编码区全长,编码551 氨基酸的蛋白,理论分子质量为63.37 ku,等电点为7.74;tdch除含有血蓝蛋白结构域以外,还含有酪氨酸酶Tyrosinase 结构域;tdch 在对虾肌肉、肝胰腺、鳃、胃、表皮、触角、心脏、肠、幽门盲囊、眼柄和神经索中均有表达,眼柄中表达最高,在幽门盲囊中表达最低。Poly(I:C)刺激后,tdch在血细胞中的表达显著升高,在鳃组织中表达变化不明显;其余5 种免疫原刺激后,tdch 在鳃和血细胞中的表达均显著升高,沉默tdch 的表达可导致副溶血弧菌感染对虾存活率显著下降,表明tdch 参与了抗副溶血弧菌免疫应答。

【目的】克隆凡纳滨对虾(Litopenaeus vannamei)一种新的含酪氨酸酶结构域血蓝蛋白(Tyrosinase-domain containing hemocyanin,TDCH),研究其免疫功能。【方法】根据NCBI上公布的tdch基因序列设计克隆引物,通过RT-PCR方法获得tdch的编码区;采用生物信息学方法分析tdch的结构特征;使用荧光定量PCR技术分析tdch在凡纳滨对虾12 个组织中的表达,以及在副溶血弧菌(Vibrio parahaemolyticus)、金黄色葡萄球菌(Staphhylococcu saureus)、黑曲霉菌(Aspergillus niger)、白斑综合征病毒、脂多糖及聚肌苷酸-聚胞苷酸[Poly(I:C)]刺激后在鳃和血细胞中的表达变化;利用RNAi 技术敲降tdch 在对虾中的表达,研究其在对虾抗副溶血弧菌免疫中的作用。【结果与结论】克隆获得tdch 的编码区全长,编码551 氨基酸的蛋白,理论分子质量为63.37 ku,等电点为7.74;tdch除含有血蓝蛋白结构域以外,还含有酪氨酸酶Tyrosinase 结构域;tdch 在对虾肌肉、肝胰腺、鳃、胃、表皮、触角、心脏、肠、幽门盲囊、眼柄和神经索中均有表达,眼柄中表达最高,在幽门盲囊中表达最低。Poly(I:C)刺激后,tdch在血细胞中的表达显著升高,在鳃组织中表达变化不明显;其余5 种免疫原刺激后,tdch 在鳃和血细胞中的表达均显著升高,沉默tdch 的表达可导致副溶血弧菌感染对虾存活率显著下降,表明tdch 参与了抗副溶血弧菌免疫应答。

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凡纳滨对虾含酪氨酸酶结构域血蓝蛋白的免疫功能