Cloning And Expression Analysis Of Estrogen Receptor Gene In Pomacea Canaliculata

The apple snail,Pomacea canaliculata is native to the Amazon River Basin of South America,it has recently been introduced to several Asian countr ies where it has unexpectedly developed into a pest of rice.The World Conservation Union regards it as one of malignant Alien Species.The snails are not only harmful to aquatic crops,but also can spread diseases,like Angiostrongyliasis cantonensis.For the first time,the full-length c DNA sequence of estrogen receptor of Pomcea canaliculata was cloned by RAC E technology.And research on the tissue distribution of ER,expression of ovarian development in different stages,the expression of the ER in the testis and in the ovary under the action of exogenous hormones.The main results were as follows:(1)Molecular characteristics of ERPrimers were designed according to the conserved region of the ER gene of the close relationship of species and by cloning,sequencing and splicing,obtained full-length sequence of c DNA,total 1636 bp,including 5 'UTR 128 bp,3' UTR 101 bp.A 1407 bp ORF encoding 468 amino acids,the content of GC in encoding region was 50.68% and the isoelectric point was 7.40,molecular amount of 51.1KD,hydrophilic amino acid content was 67.32%,no signal peptide sequence and transmembrane peptide segment were detected.(2)Tissue distribution of ERTissue distribution of ER was performed by real-time fluorescence quantitative PCR assays,The results showed that ER were expressed in the snail testis,ovary,liver,head,foot,tentacles,stomach,gills,muscle and gland protein,and in these reproductive axis tissues such as the head,liver,testis,ovary and gland protein,the expression was relatively high.(3)Expression of E R in ovarian developmentER relative expression fluctuation during the ovary development was performed by real-time fluorescence quantitative PCR assays,the results showed that: the expression of ER tended to decline after rising initially,and reach the highest at maturing stage,conversely at recovering.The expression was up-regulated significantly at maturing stage with others and down-regulated significantly at proliferating stage,recovering stage with spawning stage.(4)Expression of ER under exogenous hormone treatmentThrough the study of real time fluoresce nce quantitative PCR in ER expression under the action of exogenous hormones,on the whole,the high concentration of estradiol can promote the expression of ER gene,and the expression level reached the highest on the third day;concentrations of tamoxife n can inhibit the expression of ER,and the inhibitory effect is more significant in male snails.The results of this study laid a foundation for further elucidate developmental regulation mechanism of apple snails and other gastropods reproduction,understanding exogenous steroid hormones influence sexual maturity in molluscs,at the same time also provides a theoretical basis for the regulation of reproduction of Pomacea canaliculata.