动物医学进展2023，Vol.44Issue(12)：29-35,7.
多房棘球蚴RNA结合蛋白EWS的原核表达及其细胞定位与功能的初步鉴定
Prokaryotic Expression of the RNA Binding Protein EWS in Echinococcus multilocularis and Preliminary Identification of Its Cellular Localization and Function
摘要
Abstract
Through prokaryotic expression,antibody preparation and tissue distribution of RNA binding protein EWS(EmEWS)gene of Echinococcus multilocularis(E.multilocularis),the gene characteristics and binding ability of EmEWS with RNA were preliminarily explored.A pair of primers were designed ac-cording to the sequence from WormBase.EmEWS gene was amplified by RT-PCR and a recombinant ex-pression plasmid was constructed.The recombinant EmEWS protein was induced expressed and purified.EmEWS polyclonal antibody was prepared and identified by Western blot,real-time fluorescence quantita-tive PCR,immunofluorescence assay and dual fluorescence assay.The results showed that the full length of EmEWS gene was 1 485 bp,encoding 495 amino acids.The EmEWS gene was highly expressed in the pri-mary cells of E.multilocularis and lowest in the protoscoleces.Bioinformatics analysis showed that EWS of Echinococcus granulosus was 98.3％similar to EmEWS,and the RBD domain of EmEWS was highly con-served in tapeworms.The size of recombinant EmEWS protein was about 54 ku,and the prepared poly-clonal antibody could specifically recognize the native EmEWS protein.Immunoflurescence showed that EmEWS mainly distributed on the cell membrane.Double fluorescence assay demonstrated that EmEWS could bind to UG-rich RNA sequences.Taken together,it is speculated that EmEWS may be involved in the formation of small RNA as a transcription regulator and contribute to nuclear protein transport.关键词
多房棘球蚴/RNA结合蛋白EWS基因/原核表达/多克隆抗体/免疫荧光Key words
Echinococcus multiloculais/RNA binding protein EmEWS gene/prokaryotic expression/poly-clonal antibody/immunoflurescence分类
农业科技引用本文复制引用
严玉婷,郭小腊,孙玥,何学东,丁军涛,郑亚东..多房棘球蚴RNA结合蛋白EWS的原核表达及其细胞定位与功能的初步鉴定[J].动物医学进展,2023,44(12):29-35,7.基金项目
国家自然科学基金项目(31702224) （31702224）
浙江农林大学科研发展人才基金项目(2021LFR038) （2021LFR038）
