三角帆蚌LRR基因的分子特征及表达分析

Molecular Characteristicsand Expression Analysis of Leucine-Rich Repeat (LRR) Gene from Hyriopsis cumingii

近年来珠蚌养殖业一直饱受病害困扰,现有资料表明富亮氨酸重复序列（leucine-rich repeat,LRR）蛋白在无脊椎动物的蛋白互作、识别过程和免疫反应中都起到至关重要的作用,本实验通过RACE法克隆并表达了三角帆蚌LRR基因,全长为3 492 bp,其中开放阅读框2 142 bp,编码713个氨基酸,其中5个氨基酸组成信号肽,而其余688个氨基酸则形成成熟肽,氨基酸序列经相关软件预测存在跨膜结构。运用荧光定量PCR技术分析LRR在正常组织中及受到嗜水气单胞菌侵染后相关组织表达量的变化,分析发现,在非胁迫状态下,LRR基因在斧足中高表达,在其他组织中几乎不表达。嗜水气单胞菌刺激侵染后可以看到斧足和肝胰腺中LRR基因表达显著下调,而其他组织表达量在不同时间点均出现表达峰值,显示能被显著诱导,说明LRR基因表达可能与抵挡病原体入侵有关。

Infecti ous diseases have seriously inhibited the pearl oyster breeding in recent years. Available data showed that leucine-rich repeat（LRR） proteins play a crucial role in protein-protein interactions, recognition processes and immune reactions of invertebrates. In the present study, we cloned and characterized a LRR cDNA from the Hyriopsis cumingii by using the RACE strategy. Hyriopsis Cumingii LRR gene＇s full length was 3 492 bp,the open reading frame of which was 2 142 bp, encoding 713 amino acids, including the period of the 25 amino acid signal peptide and 688 amino acid mature peptide. Amino acid sequence analysis showed that the sequence contained a transmembrane structure. We used fluorescent quantitative PCR to examine the LRR expression changes in normal tissues and related tissues infected by Aeromonas hydrophila. The results showed that, under non-stress state, LRR gene highly expressed in foot, barely in other tissues. After been infected by Aeromonas hydrophila, it could be observed that the expression of LRR gene was significantly down-regulated in foot and hepatopancreas, while in other tissues the expression of LRR reached its peaked at different time points, indicating it could be significantly induced. It suggested the expression of LRR gene might be associated with the organism immunity against pathogens.