温度、盐度变化及鳗弧菌刺激对菲律宾蛤仔Ⅰ型溶菌酶基因表达的影响  

Expression analysis of two Ⅰ-type lysozyme genes from Ruditapes philippinarum under temperature and salinity stress and Vibrio anguillarum challenge

作　　者：祝艳霞[1,2] 母昌考[1,2] 王春琳[1,2] 

机构地区：[1]宁波大学海洋学院,浙江宁波315211 [2]浙江海洋高效健康养殖协同创新中心,浙江宁波315211

出　　处：《水产学报》2016年第12期1813-1822,共10页Journal of Fisheries of China

基　　金：国家科技支撑计划(2011BAD13B0903)~~

摘　　要：为研究在环境因素变化后,菲律宾蛤仔I型溶菌酶参与免疫应答反应的变化模式,实验采用c DNA末端快速扩增技术,从菲律宾蛤仔体内克隆获得了一种I型溶菌酶基因的全长c DNA序列(命名为Rpi LYZ-2),该序列开放阅读框(ORF)为471 bp,编码156个氨基酸。Rpi LYZ-2基因在所检测组织中均有表达,其中在外套膜中表达量最高,在肌肉组织中表达量最低。采用荧光定量PCR法,研究了不同温度[(29±1)、(21±1)和(13±1)°C]、盐度(32、22和12)及鳗弧菌刺激对菲律宾蛤仔I型溶菌酶基因表达的影响。结果显示,在温度21°C和盐度22胁迫处理后,Rpi LYZ-1基因的表达量呈现先上升后下降的趋势,而Rpi LYZ-2基因的表达量则呈现先下降后上升的趋势。在高温29°C和低盐12胁迫条件下,Rpi LYZ-1基因表达量总体呈下降趋势,而Rpi LYZ-2基因表达量总体呈上升趋势;经鳗弧菌刺激后,Rpi LYZ-1基因的表达量得到显著诱导,而Rpi LYZ-2基因表达量则表现为先降低后上升的趋势。研究表明,环境因子变化和病原菌刺激能够显著影响菲律宾蛤仔两种I型溶菌酶基因的表达量,且Rpi LYZ-1和Rpi LYZ-2可能存在着功能分化。I-type lysozymes have been found to play significant roles in the innate immune responses of mollusk.In the present study,the full-length c DNA of an I-type lysozyme（designated Rpi LYZ-2） was cloned from Ruditapes philippinarum by RACE approaches. The open reading frame of Rpi LYZ-2 was 471 bp and encoded a polypeptide of 156 amino acids. The Rpi LYZ-2 transcript was expressed in all detected tissues with the highest expression level in mantle and the lowest expression level in muscle. After being exposed to temperature[（29±1）,（21±1）,（13±1）°C],salinity（32,22 and 12） stress and Vibrio anguillarum challenge,the expression of Rpi LYZ-1and Rpi LYZ-2 displayed distinct pattern in the detected tissues. After being stimulated by elevated temperature（21°C） or salinity 22 stress,the expression pattern of Rpi LYZ-1 transcript was decreased after the initial increase,while the expression pattern of Rpi LYZ-2 transcript displayed an opposite trend. After being stimulated by elevated temperature 29 °C or salinity 12 stress,the expression pattern of Rpi LYZ-1 transcript was down-regulated as time progressed on,while the expression of Rpi LYZ-2 m RNA showed an opposite trend. After being challenged with V.anguillarum,a significant up-regulation of Rpi LYZ-1 was observed except 12 h,while the expression of Rpi LYZ-2m RNA was significantly inhibited at the early stage of bacterial challenge and recovered at 72 h. The above results suggested that environmental factors and pathogenic stimulation affected the clam＇s innate immune responses and the expression of Rpi LYZ-1 and Rpi LYZ-2,and functional division perhaps existed between Rpi LYZ-1 and Rpi LYZ-2.

关 键 词：菲律宾蛤仔 Ⅰ型溶菌酶 基因克隆 温度 盐度 鳗弧菌 

分 类 号：Q785[生物学—分子生物学] S968.3[农业科学—水产养殖]