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《Journal of Northwest A & F University(Natural Science Edition)》 2019-01
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Cloning of GeHSP70 gene of Gymnocypris eckloni and its response to Cu~(2+) stress

LI Ruihong;WEI Fulei;ZHANG Xia;WANG Yuqing;ZHU Shihai;WANG Guojie;WANG Zhenji;LI Changzhong;College of Eco-Environmental Engineering,Qinghai University;Qinghai Fisheries Environment Monitoring Station;  
【Objective】This study cloned Gymnocypris eckloni GeHSP70 gene and analyzed its bioinformatics and expression pattern under normal condition and Cu2+stress.【Method】The full-length cDNA of Gymnocypris eckloni GeHSP70 gene was cloned by molecular biology technology,and its bioinformatics characteristics were analyzed.The GeHSP70 mRNA expressions in liver,kidney,brain and gill under normal and 0.01 mg/L Cu2+stress(0,12,24 and 48h)were analyzed by Real-time PCR technology.TheELISA kit was used to determine the protein level of GeHSP70 in liver and gill.【Result】Gymnocypris eckloni GeHSP70 sequence is 1 387 bp in length with 123 bp 5′-UTR,592 bp 3′-UTR and 672 bp encoding region,encoding 223 amino acids.GeHSP70 is a hydrophobic protein with molecular weight of 11.15 ku and an isoelectric point of 5.01.It includes 2 HSP70 family feature sequences of IDLGTTYS(11-18 amino acid)and IFDLGGGTFDVSIL(199-212 amino acid).The nucleotide sequence of GeHSP70 gene has the highest homology with Carassius auratus(93%).Real-time PCR showed that the expression of Gymnocypris eckloni GeHSP70 gill tissue under normal conditions is the highest,followed by liver,brain and gill tissues.With the extension of 0.01mg/L Cu2+exposure,GeHSP70 mRNA expressions of Gymnocypris eckloni in liver,kidney and gill tissues firstly increased then decreased with the peak at 24 h,while the expression in brain tissue increased all the time.The protein levels in Gymnocypris eckloni liver and gill tissues first increased and then decreased,indicating that Cu2+induced the expression of GeHSP70.【Conclusion】This study successfully cloned the full-length Gymnocypris eckloni cDNA of GeHSP70,and GeHSP70 showed regular response at mRNA and protein levels to 0.01mg/L Cu2+exposure,indicating that GeHSP70 is a potential biomarker to reflect the quality of water environment.
【Fund】: 国家自然科学基金项目(31460700)
【CateGory Index】: X503.225;S917.4
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