Molecular characterization and developmental expression of Ras-related nuclear protein in the oriental river prawn Macrobrachium nipponense and the effects of RNA interference on ovarian maturation

Abstract

PDF HTML阅读 XML下载 导出引用 引用提醒 青虾Ran基因的克隆、表达及其在卵巢发育中的功能 DOI: 作者: 作者单位: 1. 南京农业大学 无锡渔业学院, 江苏 无锡 214081;2. 中国水产科学研究院 淡水渔业研究中心, 农业部淡水渔业和种质资源利用重点实验室, 江苏 无锡 214081 作者简介: 卜宗元(1991-),男,硕士研究生,研究方向为水产动物遗传育种.E-mail:375334258@qq.com 通讯作者: 中图分类号: S917 基金项目: 江苏省重点研发计划（现代农业）重点项目（BE2016308）；中国水产科学研究院基本科研业务费专项项目（2016HY-ZD0402）；国家自然科学基金面上项目（31572617）；江苏省农业科技自主创新基金项目[CX（15）10124]；江苏省水产三新工程（D2015-16）；无锡科学科技发展基金项目（CLE02N1514） Molecular characterization and developmental expression of Ras-related nuclear protein in the oriental river prawn Macrobrachium nipponense and the effects of RNA interference on ovarian maturation Author: Affiliation: 1. Wuxi Fisheries College, Nanjing Agricultural University, Wuxi 214081, China;2. Key Laboratory of Freshwater Fisheries and Germplasm Resources Utilization, Ministry of Agriculture; Freshwater Fisheries Research Center, Chinese Academy of Fishery Sciences, Wuxi 214081, China Fund Project: 摘要 | 图/表 | 访问统计 | 参考文献 | 相似文献 | 引证文献 | 资源附件 | 文章评论 摘要:本研究应用RACE技术克隆了青虾（（Ras related nuclear protein，Ras 相关核蛋白）基因全长cDNA序列，该基因cDNA全长1191 bp，包括218 bp的5'UTR，648 bp的开放阅读框（ORF），405 bp的3'UTR，编码215个氨基酸。青虾基因属于P-loop-NTPase超级家族，拥有PTZ00132跨结构域，多肽分子量约为24.57 kDa，理论等电点7.13。系统进化树分析表明，在动物界进化中非常保守的青虾Ran多肽与罗氏沼虾（基因在成体青虾不同组织和卵巢不同发育期的表达差异，结果显示，基因的表达水平呈现上升趋势，在卵巢消退期又恢复到较低水平。RNA干扰后，实验组（vitellogenin）在卵巢中的表达量也显著低于对照组（基因的表达起到调控作用。 Abstract:Ras-related nuclear protein (Ran) is a small GTPase with many functions, such as hydrolysis of GTP, control of cell development, replication of DNA, and RNA transcription. In this study, the cDNA-encoding ) was cloned using expressed sequence tag (EST) analysis and a rapid amplification of cDNA ends (RACE) approach. The full-length cDNA of was 1191 bp, comprising a 5' untranslated region of 218 bp, a 3' untranslated region of 405 bp, and an open reading frame of 648 bp. The deduced protein had 215 amino acid residues with a molecular mass of 24.6 kD, and 7.13 point of theoretical isoelectric. Ran belongs to the P-loop NTPase super family. Ran has a PTZ00132 model that crosses multiple domains. The members of P-loop NTPase super family have extremely conservative nucleotide sequences. Phylogenetic analyses indicate evolution of Ran proteins within the animal kingdom is very conservative, with that of . Quantitative real-time RT-PCR analysis revealed the gene was expressed in testis, ovary, brain, muscle, eyestalk, abdominal nerve, heart and gill tissues. The ovary has the highest level of expression and the eyestalk has the lowest level of expression ( gene expression of ovary is seven-eight times higher than that of testis, and the expression level of gene increased with the development of ovary. After ovulation in ovarian regression period, the expression level of gene was at a low level. After RNA interference (RNAi), expression of gene in an experimental group of adult females was significantly lower than in the control group (<0.05). After RNA interference (RNAi) in the mature female prawns, the expression of gene in experimental group (injected dsRNA solution into the shrimp's pericardial cavity) was significantly lower than in the control group (injected equal amount of DEPC water into the pranw's pericardial cavity) ( gene changes with the development of ovary. The expression of gene increased from the early stage of ovarian development to the mature stage and decreased rapidly after ovulation in the ovary. Expression of in ovarian tissues in the experimental group was also significantly lower than that of control group (<0.05), indicating RNA interference was effective. Expression of vitellogenin was significantly affected by RNA interference, with expression in the experimental group significantly lower than in the control group ( gene plays a regulatory role in expression of the gene is involved in female ovary development. 参考文献 相似文献 引证文献