Construction of Suppression Subtractive Hybridization Library for Testis of Male Tilapia Under the Stress of Methomyl
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摘要
[Objectives] This study was conducted to construct forward and reserve libraries of suppression subtractive hybridization(SSH) in the testis of male tilapia under the stress of methomyl by using SSH technology. [Methods] Using male tilapia as the test animal, the forward and reserve libraries of SSH in the testis of tilapia under the stress of methomyl were constructed by using the SSH technology. [Results] 45 expressed sequence tags(ESTs) were obtained, and 25 expressed sequence tags were successfully noted, including 13 forward libraries and 12 reverse libraries. The genes with confirmed functions were classified into five types. Genes related to catalytic activity and cell characteristics were up-regulated, while genes related to structural molecule's activity and biological process were down-regulated. The expression amount of integrin β1 was up-regulated, while serine/threonine protein kinase pim-3, Ca~(2+)-ATPase, Na~+-K~+-ATPase and ribosomal protein L22 were down-regulated. [Conclusions] The research results could lay a foundation for revealing the molecular mechanism of methomyl's reproductive toxicity to tilapia.
[Objectives] This study was conducted to construct forward and reserve libraries of suppression subtractive hybridization(SSH) in the testis of male tilapia under the stress of methomyl by using SSH technology. [Methods] Using male tilapia as the test animal, the forward and reserve libraries of SSH in the testis of tilapia under the stress of methomyl were constructed by using the SSH technology. [Results] 45 expressed sequence tags(ESTs) were obtained, and 25 expressed sequence tags were successfully noted, including 13 forward libraries and 12 reverse libraries. The genes with confirmed functions were classified into five types. Genes related to catalytic activity and cell characteristics were up-regulated, while genes related to structural molecule's activity and biological process were down-regulated. The expression amount of integrin β1 was up-regulated, while serine/threonine protein kinase pim-3, Ca~(2+)-ATPase, Na~+-K~+-ATPase and ribosomal protein L22 were down-regulated. [Conclusions] The research results could lay a foundation for revealing the molecular mechanism of methomyl's reproductive toxicity to tilapia.
[Objectives] This study was conducted to construct forward and reserve libraries of suppression subtractive hybridization(SSH) in the testis of male tilapia under the stress of methomyl by using SSH technology. [Methods] Using male tilapia as the test animal, the forward and reserve libraries of SSH in the testis of tilapia under the stress of methomyl were constructed by using the SSH technology. [Results] 45 expressed sequence tags(ESTs) were obtained, and 25 expressed sequence tags were successfully noted, including 13 forward libraries and 12 reverse libraries. The genes with confirmed functions were classified into five types. Genes related to catalytic activity and cell characteristics were up-regulated, while genes related to structural molecule's activity and biological process were down-regulated. The expression amount of integrin β1 was up-regulated, while serine/threonine protein kinase pim-3, Ca~(2+)-ATPase, Na~+-K~+-ATPase and ribosomal protein L22 were down-regulated. [Conclusions] The research results could lay a foundation for revealing the molecular mechanism of methomyl's reproductive toxicity to tilapia.
引文
[1]GUO XB.Environmental health[M].Beijing:Peking University Medical Press,2006.(in Chinese)
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[5]MENG SL,QIU LP,HU GD,et al.Responses and recovery pattern of sex steroid hormones in testis of Nile tilapia(Oreochromis niloticus)exposed to sublethal concentration of methomyl[J].Ecotoxicology,2016,25(10):1805-1811.
[6]MENG SL,QIU LP,HU GD,et al.Effect of methomyl on sex steroid hormone and vitellogenin levels in serum of male tilapia(Oreochromis niloticus)and recovery pattern[J].Environmental toxicology,2017,32(7):1869-1877.
[7]DIATCHENKO L,LAU YFC,CAMPBELL AP,et al.Suppression subtractive hybridization:A method for generating differentially regulated or tissue-specific c DNA probes and libraries[J].Pro Natl Acad Sci USA,1996,93(6):6025-6030.
[8]WANG J,LIU ZZ,PAN HF,et al.Analysis,pollution characteristics and health risk of carbamate pesticides in municipal drinking water sources in Zhejiang Province[J].Environmental Chemistry,2010,29(4):623-628.(in Chinese)
[9]VAN SCOY AR,YUE M,DENG X,et al.Environmental fate and toxicology of methomyl[J].Reviews of environmental contamination and toxicology,2013,222:93-109.
[10]EL-SAEID MH,AL-TURKI AM,AL-WABLE MI,et al.Evaluation of pesticide residues in Saudi Arabia ground water[J].Research journal of environmental sciences,2011,5(2):171-178.
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[13]ZHANG B,FU XQ,XU NB,et al.Determination of carbamates and their metobolites in water using liquid chromatography-mass spectrometry[J].Environmental Protection of Chemical Industry,2010,30(4):364-367.(in Chinese)
[14]WANG ZM,LU ZL,GUO AL,et al.Advances in research on the synthesis of adrenal corticosteroid hormone[J].Foreign Medical Sciences:Section of Endocrinology,1997,17(4):167-173.(in Chinese)
[15]YU HS,CHENG HH,GUO YQ,et al.Alternative splicing and differential expression of P450c17(CYP17)in gonads during sex transformation in the rice field eel[J].Biochemical and biophysical research communication,2003,307(1):165-171.
[16]BORG B.Androgens in teleost fishes[J].Comparative biochemistry and physiology part C,1994,109(3):219-245.
[17]MENG SL.Effect of environmental estrogen methomyl on hypothalamuspituitary-gonadal axis and antioxidant defense system of male tilapia Oreochromis[D].Nanjing:Nanjing Agricultural University,2014.(in Chinese)
[18]CHENG CY,MRUK DD.Cell junction dynamics in the testis:Sertoligerm cell interactions and male contraceptive development[J].Physiol Rev,2002,82(4):825-874.
[19]SALANOVA M,STEFANINI M,DE CURTIS I,et al.Integrin receptorα6β1 is localized at specific sites of cell-to-cell contact in rat seminiferous epitheium[J].Biol Reprod,1995,52(1):79-87.
[20]LEE NP,CHENG CY.Ectoplasmic specialization,a testis-specific cellcell actin-based adherens junction type:Is this a potential target for male contraceptive development[J].Hum Reprod Update,2004,10(4):349-369.
[21]HCKER G,PASCHEN SA.The rapeutic targets in the mitochondrial apoptotic pathway[J].Expert Opin Ther Targets,2007,11(4):515-526.
[22]ZHA JP,HARADA H,YANG E,et al.Serine phosphorylation of death agonist BAD in response to survival factor results in binding to 14-3-3 not BCL-XL[J].Cell,1996,87(4):619-628.
[23]YANG E,ZHA JP,JOCKEL J,et al.Bad,a heterodimeric partner for Bcl-XL and Bcl-2,displaces Bax and promotes cell death[J].Cell,1995,80(2):285-291.
[24]LI HY.Effect of ribosomal protein L22 on the development of Drosophila and its mechanism[J].Journal of Hebei University:Natural Science E-dition,2014,34(1):64-69.(in Chinese)
[25]SUN K,XUE H,XIE WP,et al.Effect of silencing of ribosomal protein L22 on the expression of cyclin D1[J].Jiangsu Medical Journal 2015,41(23):2791-2793.(in Chinese)
[2]SUN L,LEE HK.Stability studies of propoxur herbicide in environmental water samples by liquid chromatography-atmospheric pressure chemical ionization ion-trap mass spectrometry[J].Journal of chromatography,2003,1014(1/2):153-163.
[3]HE RL,QI JK,ZHANG YF,et al.Investigation on pesticide residue quantity of rice in Tongliao area[J].Chinese Journal of Public Health Management,2007,23(5):464-466.(in Chinese)
[4]MENG SL,QIU LP,HU GD,et al.Effects of methomyl on steroidogenic gene transcription of the hypothalamic-pituitary-gonad-liver axis in male tilapia[J].Chemosphere,2016,165:152-162.
[5]MENG SL,QIU LP,HU GD,et al.Responses and recovery pattern of sex steroid hormones in testis of Nile tilapia(Oreochromis niloticus)exposed to sublethal concentration of methomyl[J].Ecotoxicology,2016,25(10):1805-1811.
[6]MENG SL,QIU LP,HU GD,et al.Effect of methomyl on sex steroid hormone and vitellogenin levels in serum of male tilapia(Oreochromis niloticus)and recovery pattern[J].Environmental toxicology,2017,32(7):1869-1877.
[7]DIATCHENKO L,LAU YFC,CAMPBELL AP,et al.Suppression subtractive hybridization:A method for generating differentially regulated or tissue-specific c DNA probes and libraries[J].Pro Natl Acad Sci USA,1996,93(6):6025-6030.
[8]WANG J,LIU ZZ,PAN HF,et al.Analysis,pollution characteristics and health risk of carbamate pesticides in municipal drinking water sources in Zhejiang Province[J].Environmental Chemistry,2010,29(4):623-628.(in Chinese)
[9]VAN SCOY AR,YUE M,DENG X,et al.Environmental fate and toxicology of methomyl[J].Reviews of environmental contamination and toxicology,2013,222:93-109.
[10]EL-SAEID MH,AL-TURKI AM,AL-WABLE MI,et al.Evaluation of pesticide residues in Saudi Arabia ground water[J].Research journal of environmental sciences,2011,5(2):171-178.
[11]U.S.EPA.2012 edition of the drinking water standards and health advisories:EPA 822-S-12-001[S].Washington,DC:Office of Water U.S.Environmental Protection Agency,2012.
[12]ZHANG XJ,CHENG YH,LI SR.Determination of carbamate pesticides in drinking water by solid phase extraction/high performance liquid chromatography[J].Chinese Journal of Health Laboratory Technology,2014,24(9):1243-1244,1247.(in Chinese)
[13]ZHANG B,FU XQ,XU NB,et al.Determination of carbamates and their metobolites in water using liquid chromatography-mass spectrometry[J].Environmental Protection of Chemical Industry,2010,30(4):364-367.(in Chinese)
[14]WANG ZM,LU ZL,GUO AL,et al.Advances in research on the synthesis of adrenal corticosteroid hormone[J].Foreign Medical Sciences:Section of Endocrinology,1997,17(4):167-173.(in Chinese)
[15]YU HS,CHENG HH,GUO YQ,et al.Alternative splicing and differential expression of P450c17(CYP17)in gonads during sex transformation in the rice field eel[J].Biochemical and biophysical research communication,2003,307(1):165-171.
[16]BORG B.Androgens in teleost fishes[J].Comparative biochemistry and physiology part C,1994,109(3):219-245.
[17]MENG SL.Effect of environmental estrogen methomyl on hypothalamuspituitary-gonadal axis and antioxidant defense system of male tilapia Oreochromis[D].Nanjing:Nanjing Agricultural University,2014.(in Chinese)
[18]CHENG CY,MRUK DD.Cell junction dynamics in the testis:Sertoligerm cell interactions and male contraceptive development[J].Physiol Rev,2002,82(4):825-874.
[19]SALANOVA M,STEFANINI M,DE CURTIS I,et al.Integrin receptorα6β1 is localized at specific sites of cell-to-cell contact in rat seminiferous epitheium[J].Biol Reprod,1995,52(1):79-87.
[20]LEE NP,CHENG CY.Ectoplasmic specialization,a testis-specific cellcell actin-based adherens junction type:Is this a potential target for male contraceptive development[J].Hum Reprod Update,2004,10(4):349-369.
[21]HCKER G,PASCHEN SA.The rapeutic targets in the mitochondrial apoptotic pathway[J].Expert Opin Ther Targets,2007,11(4):515-526.
[22]ZHA JP,HARADA H,YANG E,et al.Serine phosphorylation of death agonist BAD in response to survival factor results in binding to 14-3-3 not BCL-XL[J].Cell,1996,87(4):619-628.
[23]YANG E,ZHA JP,JOCKEL J,et al.Bad,a heterodimeric partner for Bcl-XL and Bcl-2,displaces Bax and promotes cell death[J].Cell,1995,80(2):285-291.
[24]LI HY.Effect of ribosomal protein L22 on the development of Drosophila and its mechanism[J].Journal of Hebei University:Natural Science E-dition,2014,34(1):64-69.(in Chinese)
[25]SUN K,XUE H,XIE WP,et al.Effect of silencing of ribosomal protein L22 on the expression of cyclin D1[J].Jiangsu Medical Journal 2015,41(23):2791-2793.(in Chinese)
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