Cloning And Expression Analysis Of Ecdysteroid Receptor(EcR) In Portunus Trituberculatus

To study the regulatory role of ecdysteroid receptor (EcR) in molting and ovarian development ofcrustaceans, the full-length EcR cDNA of Portunus trituberculatus(PtEcR) was cloned by using reversetranscript PCR (RT-PCR) and rapid amplification of cDNA ends (RACE). The full-length of PtEcR(GenBank accession number: KC354381) was2231bp, included a1269bp ORF which encoded422aminoacid residues. The alignment of amino acid sequence of PtEcR and that of other crustaceans showed that theiridentities were67%to97%. BLAST analysis revealed that the deduced amino acid sequence of PtEcRshared67%identity with Litopenaeus vannamei EcR,95%identity with Scylla paramamosain EcRa,95%identity with S. paramamosain EcRb and97%identity with S. paramamosain EcRc. Phylogenetic analysisof EcR showed PtEcR was clustered in crustaceans EcRs and separated from insect EcRs. Quantitative real-time PCR(qRT-PCR) was used to quantify the relative expression level of PtEcR in different tissues, moltingprocess and the second ovarian development in P. trituberculatus. PtEcR was expressed in various tissues(perider; gonad; hepatopancreas; eyestalk ganglia; mandibular organ; Y-organ; heart; muscle; thoracicganglion)and highest in the Y-organ(YO). During the molting process, the expression levels of PtEcR in YOremained low from postmolt period to substage of D2premolt period, then significantly increased at substageD3and D4.The changes in the expression levels of PtEcR was consistent with that in the levels ofhemolymphatic20-hydroxyecdysone, which demonstrated that PtEcR played an important role in moltingregulation of P. trituberculatus. During the first ovarian development, the expression levels of PtEcR in YO,hepatopancreas and ovary gradually increased to the maximum from earlier stage to the fast developingstages (I to IV), then gradually decreased to the minimum at mature stage (V); During the second ovariandevelopment, the expression levels of PtEcR in YO and hepatopancreas gradually increased to the maximumfrom stage II to IV; The expression levels of PtEcR in ovary at stage II and IV was significantly higher thanstage I and III. The results indicated that PtEcR may play an important role during ovarian development in P.Trituberculatus.