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为了发掘罗氏沼虾(Macrobrachium rosenbergii)免疫相关基因,研究Rab蛋白(Ras-related proteins in brain)在罗氏沼虾免疫应答中发挥的作用,研究采用RACE-PCR技术克隆了罗氏沼虾Rab11基因全长c DNA序列,记为Mr Rab11。全长1381 bp,包括226 bp的5′UTR,511 bp的3′UTR和645 bp的开放阅读框,编码214个氨基酸,含有一个Rab结构域。氨基酸序列比对显示,罗氏沼虾与冈比亚按蚊(Anopheles gambiae)、蚤状溞(Daphnia pulex)、叶蝉(Homalodisca vitripennis)Rab11一致性分别为82%、83%和82%。软件预测,Mr Rab11编码的蛋白分子量约为23.75 k D,等电点约为5.34。实时荧光定量表达分析表明,Mr Rab11基因在罗氏沼虾各组织中都有表达,肝胰腺中的表达量最高,其次是肌肉和肠道。在阴沟肠杆菌(Enterobacter cloacae)感染12h后,罗氏沼虾肝胰腺中Mr Rab11的表达量上升,显著高于对照组(P<0.05),推测这是Mr Rab11对阴沟肠杆菌的应激表达,Mr Rab11在肝胰腺中参与了罗氏沼虾免疫应答过程。
In order to explore the immune related genes of Macrobrachium rosenbergii and investigate the role of Ras-related proteins in brain in the immune response of Macrobrachium rosenbergii, the full-length Rab11 gene was cloned by RACE-PCR c DNA sequence, denoted as Mr Rab11. It has a length of 1381 bp and consists of 226 bp 5’UTR, 511 bp 3’UTR and 645 bp open reading frame, which encodes 214 amino acids and contains a Rab domain. Amino acid sequence alignment showed that the consistency of Rab11 was 82%, 83% and 82% with Anopheles gambiae, Daphnia pulex and Homalodisca vitripennis respectively. The software predicts that Mr Rab11 encodes a protein with a molecular weight of about 23.75 kD and an isoelectric point of about 5.34. Real-time quantitative PCR analysis showed that the Mr Rab11 gene was expressed in all tissues of Macrobrachium rosenbergii, with the highest expression in hepatopancreas, followed by muscle and intestine. The expression of Mr Rab11 in hepatopancreas of M. rosenii increased significantly 12 h after Enterobacter cloacae infection, which was significantly higher than that of the control group (P <0.05). It was speculated that this was the stress expression of Mr Rab11 against Enterobacter cloacae, Mr Rab11 is involved in the immune response to M. rosenbergii in the hepatopancreas.