Cloning of MyD88 partial cDNA sequence of Chinese soft shelled turtle (Trionyx sinensis) and its expression in different tissues

Myeloid differentiation factor 88(MyD88),which is considered as a pivot adaptor molecule for signal transduction of all toll-like receptors(TLRs) except TLR3,possesses important function.In this study,degenerate primer pairs were designed from the conserved regions of the Toll-like/IL-1 receptor(TIR) domain of myeloid differentiation factor 88(MyD88).A 351-bp length target sequence was amplified by PCR successfully from the spleen cDNA of Chinese soft-shelled turtle(CSST),and is found as the TIR domain of CSST MyD88 as shown by sequence alignment and domain search.Its identity is 72.9%-86%and 77.6%-83.6%respectively at the nucleotide and amino acid levels as compared with the MyD88 TIR domains of other vertebrates(e.g.yellow croca［Pseudosciaena crocea］,chicken).Real-time PCR revealed that mRNA expression of CCST MyD88 in the liver,spleen and kidney increased to varying extent within 48 hours after intramuscular stimulation of CSST by heat-inactivated Aermoans hydrophila.In particular, MyD88 expression in the spleen was 6.89 fold higher than that from unstimulated control. When the heart fibroblast cell line of CSST was stimulated by 20 ng LPS,as stimulator,MyD88 expression increased slightly from hour 1 to 8,but significantly at hour 24.This study has laid good foundation for further study on innate immunity of CTTS.