Toll样受体（TLR）基因家族在免疫系统中发挥着至关重要的作用。然而，鱼类多数TLR识别配体谱系尚不清楚，接头分子的数量和对应关系尚不明确。本研究以病毒性疾病损失严重且有基因组数据库的草鱼为例，全面筛查识别病毒相关分子模式（双链RNA）的TLR及其接头分子。我们前期研究发现草鱼有21个TLR成员，大多数对病毒感染表达量有变化。本研究首先重组表达LRR结构域，利用表面等离子共振等技术逐一筛查与poly(I:C)（双链RNA）的直接互作；接着通过过表达TLR、poly(I:C)刺激、转录组测序、敲降表达等对经典通路和新发现重要分子的调控来确认互作；然后免疫共沉淀结合定量蛋白组学及草鱼胞内TIR数据库筛选接头分子；最后反向免疫共沉淀结合激光扫描共聚焦显微镜共定位等技术鉴定接头分子。本研究将系统夯实鱼类识别双链RNA的TLR成员及其接头分子，为进一步阐释鱼类抗病毒免疫调控网络奠定坚实的基础。

Toll-like receptor (TLR) gene family play a crucial role in the immune system. However, the ligands remain largely unclear, and the quantities and corresponding relations of their adaptors also keep indistinct for most teleost TLR members. In the present study, a case of grass carp with severe viral disease loss and genomic database is taken as an example. We focus on the comprehensive screening of TLRs recognizing viral associated molecular pattern (double-stranded RNA) and investigation of their adaptors. Our previous studies found that there are 21 TLR members in grass carp, and mRNA expressions of most TLRs are induced post viral infection. In this study, we firstly reconstruct and express LRR (leucine-rich repeat) domains of TLRs, surface plasmon resonance (SPR) and other techniques are employed to screen the TLRs of direct interaction with poly(I:C) (double-stranded RNA). Then, TLRs overexpression, poly(I:C) stimulation, transcriptome sequencing, knockdown expression, and other techniques are used to confirm the interactions through triggering the classical pathways or the newly identified important molecules. After that, the adaptors of TLRs are investigated by Co-immunoprecipitation (Co-IP), quantitative proteome (iTRAQ) and the cytoplasmic TIR database of grass carp. In the end, the reverse Co-immunoprecipitation, laser scanning confocal microscopy and RNAi techniques are performed to identify the adaptors. The present study will systematically clarify the TLRs recognizing dsRNA and their adaptors in teleost, and lay a solid foundation for further elucidating the antiviral immune regulatory network in fish.