Antimicrobial peptides (AMPs), as key immune effectors, play important roles in the innate immune system of invertebrates. Different types of AMPs, including Penaeidin, Crustin, ALF (antilipopolysaccharide factor) have been identified in different penaeid shrimp; however, systematic analyses on the function of different AMPs in shrimp responsive to different types of bacteria are very limited. In this study, we analyzed the expression profiles of AMPs in the Chinese shrimps, Fenneropenaeus chinensis, simultaneously by real-time RT-PCR (reverse transcription-polymerase chain reaction) when shrimp were challenged with Micrococcus lysodeikticus (Gram-positive, G + ) or Vibrio anguillarium (Gram-negative, G+). Different AMPs showed different expression profiles when shrimp were injected with one type of bacterium, and one AMP also showed different expression profiles when shrimp were challenged with different bacteria. Furthermore, the expression of these AMPs showed temporal expression profiles, suggesting that different AMPs function coordinately in bacteria-infected shrimp. An RNA interference approach was used to study the function of the Relish transcription factor in regulating the transcription of different AMPs. The current study showed that Relish could regulate the transcription of different AMPs in shrimp. Differential expression profiles of AMPs in shrimp injected with different types of bacteria indicated that a complicated antimicrobial response network existed in shrimp. These data contribute to our understanding of immunity in shrimp and may provide a strategy for the control of disease in shrimp.
以中国对虾血液、眼柄、卵巢、雌虾头胸部、雄虾头胸部和三倍体对虾头胸部组织为材料，采用异硫氰酸胍一酸酚法提取总RNA；用磁珠法纯化mRNA；用UniZAPcDNA合成试剂盒合成双链cDNA并进行修饰，将cDNA定向连接在UniZAP载体上；经Gigapack GoldPackage包装试剂盒包装成为噬菌体颗粒，转染宿主XLl Blue MRF’菌株细胞，形成初级文库．初级文库经转染进一步扩增，形成稳定的cDNA文库．6个文库的库容在0．2×10^6～1．3×10^6之间，重组率都超过90％．从各文库随机取出6～10个清晰的噬菌斑进行PCR扩增，用琼脂糖凝胶电泳检测，其插入片段长度为500～2500bp．由初步功能基因克隆获得阳性结果．多项指标表明，所构建的对虾cDNA文库质量较高，为进一步筛选目的基因、EST测序和制作基因芯片提供了有效的工具．
