盐度对天津厚蟹Na^(+)/K^(+)-ATPase、GDH酶活性及GDH基因表达的影响  

Effects of Salinity on Activities of Na^(+)/K^(+)-ATPase and Glutamate Dehydrogenase(GDH) and GDH Gene Epression in Mudflat Crab Helice tientsinensis

作　　者：左迪 马长安 吴东蕾 张云飞[3] ZUO Di;MA Chang'an;WU Donglei;ZHANG Yunfei(School of Health and Social Care,Shanghai Urban Construction Vocational College,Shanghai 201415,China;Institute of Agri-food Standards and Testing Technology,Shanghai Academy of Agricultural Sciences,Shanghai 201403,China;Natural History Research Center,Shanghai Natural History Museum,Shanghai Science&Technology Museum,Shanghai 200041,China)

机构地区：[1]上海城建职业学院健康与社会关怀学院,上海201415 [2]上海市农业科学院农产品质量标准与检测技术研究所,上海201403 [3]上海科技馆,上海自然博物馆自然史研究中心,上海200041

出　　处：《水产学杂志》2022年第5期34-42,共9页Chinese Journal of Fisheries

基　　金：国家自然科学基金(31572221);国家高等职业教育食品营养与检测专业教学资源库一级子项目《水产品检测技术》课程资源(A3-2018-046);上海城建职业学院人才引进与师资队伍建设项目(B101-0101-20-001).

摘　　要：将体质量(9.91±2.01)g的经暂养的天津厚蟹(Helice tientsinensis Rathbun)暴露于6个盐度梯度:低盐组(0、2、4)、高盐组(16、32)和对照组(8),每组设三个平行,分别于0 h、6 h、12 h、24 h、48 h、72 h和96 h时,冰浴麻醉30 min后取肌肉和鳃组织,测定鳃和肌肉组织中Na^(+)/K^(+)-ATPase和谷氨酸脱氢酶(GDH)活性的变化,并从肌肉中克隆获得了GDH基因全长(GenBank登录号:KJ649743),以研究湿地围垦导致的盐度变化对天津厚蟹渗透压相关酶及基因表达的影响。结果表明,低盐胁迫48 h内,天津厚蟹Na^(+)/K^(+)-ATPase和GDH活性随水体盐度的降低呈上升趋势,96 h后,0和2盐度组酶活性随盐度降低而显著下降(P<0.05);高盐胁迫组,24 h内天津厚蟹组织中两酶活性升高,随盐度胁迫时间延长,16盐度组天津厚蟹鳃和肌肉组织中Na^(+)/K^(+)-ATPase活性低于对照组,GDH活性高于对照组,差异显著(P<0.05);32盐度组Na^(+)/K^(+)-ATPase和GDH活性低于对照组,显著高于0、2盐度组(P<0.05)。这表明低盐胁迫对两酶活性的影响更强。RACE技术获得天津厚蟹GDH基因cDNA全长为2078 bp,包含编码546个氨基酸的1641 bp的开放阅读框(ORF),预测分子量为60.54 kDa;Realtime-PCR实验结果表明:GDH基因在肌肉组织中表达量最高,且盐度胁迫后表达量变化明显,其中低盐胁迫组GDH基因表达量极显著低于对照组,表明GDH基因可能参与天津厚蟹的渗透压调节。Mudflat crab Helice tientsinensis with body weight oof(9.91±2.01)g was exposed to 6 salinity gradients:low-salinity groups(0,2,4),high-salinity groups(16,32)and the control group(8)with triplication,and activities of Na^(+)/K^(+)-ATPase and glutamate dehydrogenase(GDH)were determined in the gills and muscles at 0 h,6 h,12 h,24 h,48 h,72 h and 96 h in the crab esposed to ice bath anesthesia for 30 minutes.The full-length GDH gene was cloned from the muscle tissue by molecular biological methods to probe into the effects of changes in salinity caused by wetland reclamation on the expression of osmotic pressure-related enzymes and genes of mudflat crab in Tianjin.It was found that there was increase in activities of Na^(+)/K^(+)-ATPase and GDH with the decrease in water salinity within 48 h in low-salinity groups.At 96 h,however,the enzyme activities was shown to be decreased significantly with the decrease in salinity in 0 and 2 salinity groups(P<0.05).In the high salinity groups,after 96 h,Na^(+)/K^(+)-ATPase activity was slightly lower in the 16 group than that in the control group(P<0.05).In the 32 salinity group,the activities of both enzymes were significantly higher than those in the 0 and 2 salinity groups.It was approved that enzyme activity was changed in the gills and muscles subjected to salinity stress,and low salt stress had greater influence on the osmotic status of the mudflat crab than high salt stress did.The GDH gene cDNA sequence was found to be an overall length of 2,078 bp,a 1,641 bp open reading frame encoding 546 amino acids,and a predicted molecular weight of 60.54 kDa.Quantitative reverse transcriptase polymerase chain reaction(qRT-PCR)assays revealed that there was the maximal GDH gene expression in the muscle tissue,with effect of salinity stress and GDH gene expression,indicating that the GDH gene may involve in osmotic pressure regulation of mudflat crab.

关 键 词：天津厚蟹 盐度 渗透压调节 Na^(+)/K^(+)-ATPase 谷氨酸脱氢酶(GDH)基因 

分 类 号：S917[农业科学—水产科学]