Cloning and expression analysis of ecdysteroid receptor (EcR)inPortunus trituberculatus

Abstract

To study the regulatory role of ecdysteroid receptor( EcR) in molting and ovarian development of crustaceans,the full-length EcR cDNA of Portunus trituberculatus( PtEcR) w as cloned by using reverse transcript PCR( RT-PCR) and rapid amplification of cDNA ends( RACE). The full-length of PtEcR( GenBank accession number: KC354381) was 2 231 bp,included a 1 269 bp ORF which encoded 422 amino acid residues. The alignment of amino acid sequence of PtEcR and that of other crustaceans show ed that their identities w ere 67% to 97%. Phylogenetic analysis of EcR show ed PtEcR w as clustered in crustaceans EcRs and separated from insect EcRs. Quantitative real-time PCR( qRT-PCR) w as used to quantify the relative expression level of PtEcR in different tissues,molting process and the second ovarian development in P. trituberculatus. PtEcR w as expressed in various tissues and highest in the Y-organ( YO). During the molting process,the expression levels of PtEcR in YO remained low from postmolt period to substages of D2premolt period,then significantly increased at substages D3 and D4. The changes in the expression levels of PtEcR w ere consistent w ith those in the levels of hemolymphatic 20-hydroxyecdysone,w hich demonstrated that PtEcR played an important role in molting regulation of P. trituberculatus. During the second ovarian development,the expression levels of PtEcR in YO and hepatopancreas gradually increased to the maximum from stage Ⅱ to Ⅳ; The expression levels of PtEcR in ovary at stages Ⅱ and Ⅳ w ere significantly higher than those at stage Ⅰ and Ⅲ. The results indicated that PtEcR may play an important role during ovarian development in P. trituberculatus.