罗氏沼虾Toll样受体基因cDNA片段的克隆及序列分析

Cloning and sequence analysis of cDNA encoding Toll-like receptor in Macrobrachium rosenbergii

提取罗氏沼虾(Macrobrachium rosenbergii)上海养殖群体活体鳃、肌肉、肝胰腺、血液和雄性腺5种组织的总RNA;应用RT-PCR技术,从鳃组织中克隆到Toll样受体(TLR)基因部分cDNA序列,并进行生物信息学分析。基因序列分析表明:所得罗氏沼虾TLR基因cDNA序列长1 875 bp,包含1 728 bp的开放阅读框,可编码575个氨基酸残基;该部分TLR是一个跨膜蛋白,存在胞外区LRR、LRR-CT、LRR-NT结构域及关键的胞内区TIR结构域,具备Toll样受体家族的典型结构特征;TIR结构域与凡纳滨对虾(Litopenaeus vannamei)和日本囊对虾(Marsupenaeus japonicus)、中国明对虾(Fenneropenaeus chinensis)、斑节对虾(Penaeus monodon)、中华绒螯蟹(Eriocheir sinensis)、拟穴青蟹(Scylla paramamosian)Toll样受体基因TIR区域的氨基酸序列具有很高的相似性。

In this study,we cloned a novel partial cDNA of toll-like receptor（ TLR） gene from total RNA of gill of living giant freshwater prawn（ Macrobrachium rosenbergii） of Shanghai population by RT-PCR by using primers designed from the partial sequence from transcriptone sequencing result,then it was sequenced and bioinformatically analyzed. Sequence analysis showed that the part of TLR cDNA was 1 875 bp in length and the open reading frame encoded a deduced protein with 575 amino acids residues. The part of TLR is a transmembrane protein that contains several domains like LRR,LRR-CT,LRR-NT motif in the extracellular domain and a TIR domain in the intracellular region,the common structural features of TLRs. The results of the amino acids sequence of TIR were compared with other species,indicating a highly conserved similarity among Litopenaeus vannamei,Marsupenaeus japonicus,Fenneropenaeus chinensis,Penaeus monodon,Eriocheir sinensi and Scylla paramamosian. Besides,similar RF PCR products of TLE gene were also amplified from total RNA of other different tissues including haemocytes,hepatopancreas,muscle and androgenic gland.