驯鹿干扰素β1基因的克隆及遗传进化分析
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摘要
试验旨在从驯鹿鹿茸顶端组织中克隆干扰素β1(interferon beta 1,IFNβ1)基因全长序列,分析其分子特性,为研究其生物学活性及实际应用奠定基础。根据GenBank数据库中牛、羊等近缘物种IFNβ1基因的保守序列设计1对引物,以驯鹿鹿茸顶端间充质层组织基因组DNA为模板,采用PCR技术扩增得到驯鹿IFNβ1基因并克隆、测序,利用相关生物信息学软件对该序列进行分析。结果发现,驯鹿IFNβ1基因全长561bp,共编码186个氨基酸,含有3个糖基化位点;其编码蛋白含有4个α螺旋、4个β折叠区、7个β转角。将驯鹿IFNβ1基因推导的氨基酸序列与其他14种哺乳动物进行遗传进化分析发现,其同源性为45.1%~92.0%;驯鹿与其他动物IFNβ1基因序列分析和系统进化树分析表明,IFNβ1基因存在种属差异性,亲缘关系越近,同源性越高。本研究结果为驯鹿IFNβ1基因的进一步研究和应用提供了基础试验数据。
This study was aimed to clone the full-length sequence of interferon beta 1(IFNβ1)gene from the top of reindeer antler,and analyze its molecular characteristics,to lay a foundation for its biological activity and practical application.One pair of primers was designed based on the conserved sequence of IFNβ1 gene of cattle and sheep published in GenBank.IFNβ1 gene of reindeer was amplified by PCR from genome DNA extracted from mesenchyme of reindeer antler tip,and the product was cloned and sequenced.The result showed that the full length sequence of reindeer IFNβ1 gene was 561 bp,encoded 186 amino acids and contained 3 glycosylation sites.The encoded protein contained 4 alpha helixes,4 beta fold zones and 7 beta turn angles.Compared the amino acids of IFNβ1 gene with other mammal species,the homology was 45.1% to 92.0%.The results of sequence and phylogenetic tree analysis showed that IFNβ1 gene had species diversity,the relationship was proportional to homology.This result laid the foundation for further research on the expression,biological activity and application of reindeer IFNβ1 gene.
This study was aimed to clone the full-length sequence of interferon beta 1(IFNβ1)gene from the top of reindeer antler,and analyze its molecular characteristics,to lay a foundation for its biological activity and practical application.One pair of primers was designed based on the conserved sequence of IFNβ1 gene of cattle and sheep published in GenBank.IFNβ1 gene of reindeer was amplified by PCR from genome DNA extracted from mesenchyme of reindeer antler tip,and the product was cloned and sequenced.The result showed that the full length sequence of reindeer IFNβ1 gene was 561 bp,encoded 186 amino acids and contained 3 glycosylation sites.The encoded protein contained 4 alpha helixes,4 beta fold zones and 7 beta turn angles.Compared the amino acids of IFNβ1 gene with other mammal species,the homology was 45.1% to 92.0%.The results of sequence and phylogenetic tree analysis showed that IFNβ1 gene had species diversity,the relationship was proportional to homology.This result laid the foundation for further research on the expression,biological activity and application of reindeer IFNβ1 gene.
引文
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[15]夏伦斌.绵羊γ干扰素(IFN-γ)基因的克隆、表达与抗病毒活性初步研究[D].石河子:石河子大学,2007.
[16]焦道忠,王云龙,李晨阳,等.鸡α干扰素基因的克隆与表达[J].生物学杂志,2008,25(5):48-51.
[17]邓成钢.犬IFN-α基因的克隆、表达及其生物学活性的研究[D].扬州:扬州大学,2008.
[18]苏凤艳,李哲,刘存发,等.梅花鹿IFN-β1基因的克隆与分子进化分析[J].畜牧与兽医,2014,46(1):14-19.
[19]Li C,Clark D E,Lord E A,et al.Sampling technique to discriminate the different tissue layers of growing antler tips for gene discovery[J].Anatomical Record,2002,268(2):125.
[20]钟立成,朱立夫,卢向东.我国驯鹿起源、历史变迁与现状[J].经济动物学报,2008,12(2):102-105.
[21]翟健程.使鹿部落驯鹿种群遗传多样性研究[D].哈尔滨:东北林业大学,2015.
[22]唐戈.鄂温克族驯鹿饲养业的困境与对策[J].黑龙江民族丛刊,2008,6:129-134.
[23]邢秀梅,苏伟林,杨福合.浅谈中国驯鹿种群[J].经济动物学报,2001,5(4):57-60.
[24]孙亚红.驯鹿发展管理模式探讨[J].中央民族大学学报(自然科学版),2013,22(2):42-44.
[25]曾治,孟凡露,王朋,等.我国驯鹿(Rangifer tarandus)的种群分布、数量及致危原因[J].西南大学学报(自然科学版),2016,38(7):19-24.
[26]殷亚杰.敖鲁古雅驯鹿(Rangifer tarandus)食性及其栖息地容纳量与种群生存力分析[D].哈尔滨:东北林业大学,2016.
[27]夏春,刘津,杨琪,等.猪干扰素β基因的分子克隆与测序[J].中国兽医杂志,2000,6:6-7.
[28]Hansen K,Prabakaran T,Laustsen A,et al.Listeria monocytogenes induces IFNβexpression through an IFI16-,cGAS-and STING-dependent pathway[J].Embo Journal,2014,33(15):1654.
[29]Basler C F,Garcíasastre A.Viruses and the typeⅠinterferon antiviral system:Induction and evasion[J].International Reviews of Immunology,2009,21(4-5):305.
[30]吴杨,杨苏河.β-干扰素的特性与临床应用[J].世界医学杂志,2001,3:72-74.
[31]Kunzi M S,Pitha P M.Interferon research:A brief history[J].Methods in Molecular Medicine,2005,116:25.
[32]Staples K J,Nicholas B,Mckendry R T,et al.Viral infection of human lung macrophages increases PDL1expression via IFNβ[J].PLoS One,2015,10(3):e0121527.
[33]Cheon H J,Holveybates E G,Schoggins J W,et al.IFNβ-dependent increases in STAT 1,STAT 2,and IRF9mediate resistance to viruses and DNA damage[J].Embo Journal,2013,32(20):2751-2763.
[2]韩玉杰,王倩,张竞.干扰素β的研究现状[J].化学工程与装备,2016,8:255-257.
[3]禤雄标.猪IFN-α1、IFN-β1、白细胞介素1α和白细胞介素1β四个细胞因子基因的克隆和序列分析[D].南宁:广西大学,2006.
[4]孙瑛,刘永乐,王应玉.干扰素在兽医临床的应用[J].山东畜牧兽医,2016,4:46-47.
[5]Rogez C,Martin M,Dereuddre-Bosquet N,et al.Anti-human immunodeficiency virus activity of tau interferon in human macrophages:Involvement of cellular factors andβ-chemokines[J].Journal of Virology,2003,77(23):12914-12920.
[6]Fernandez-Sainz I,Ramanathan P,O’Donnell V,et al.Treatment with interferon-alpha delays disease in swine infected with a highly virulent CSFV strain[J].Virology,2015,483:284-290.
[7]Simpson J,Miles K,Trüb M,et al.Plasmacytoid dendritic cells respond directly to apoptotic cells by secreting immune regulatory IL-10 or IFN-α[J].Frontiers in Immunology,2016,7(73):1-13.
[8]Li M,Liu X Y.Interferon-lambdas:the modulators of antivirus,antitumor,and immune responses[J].Journal of Leukocyte Biology,2009,86(1):23-32.
[9]卡丽娜.驯鹿鄂温克人文化研究[D].北京:中央民族大学,2004.
[10]李玉萍,景志忠,蒲训,等.牦牛β-干扰素基因的克隆与表达[J].中国兽医科学,2006,36(1):40-45.
[11]曲嘉琪,李洋,钟一鸣,等.β干扰素及其研究进展[J].安徽农业科学,2013,41(9):3792-3793.
[12]Henig N,Avidan N,Mandel I,et al.Interferon-beta induces distinct gene expression response patterns in human monocytes versus T cells[J].PLoS One,2013,8(4):e62366-e62366.
[13]Guijarro C,Benito-León J,Bermejo-Pareja F.Widespread urticaria due to intramuscular interferon beta-1atherapy for multiple sclerosis[J].Neurological Sciences,2011,32(2):309-311.
[14]李玉萍,景志忠,蒲训,等.牦牛β-干扰素基因的克隆与表达[J].中国兽医科学,2006,36(1):40-45.
[15]夏伦斌.绵羊γ干扰素(IFN-γ)基因的克隆、表达与抗病毒活性初步研究[D].石河子:石河子大学,2007.
[16]焦道忠,王云龙,李晨阳,等.鸡α干扰素基因的克隆与表达[J].生物学杂志,2008,25(5):48-51.
[17]邓成钢.犬IFN-α基因的克隆、表达及其生物学活性的研究[D].扬州:扬州大学,2008.
[18]苏凤艳,李哲,刘存发,等.梅花鹿IFN-β1基因的克隆与分子进化分析[J].畜牧与兽医,2014,46(1):14-19.
[19]Li C,Clark D E,Lord E A,et al.Sampling technique to discriminate the different tissue layers of growing antler tips for gene discovery[J].Anatomical Record,2002,268(2):125.
[20]钟立成,朱立夫,卢向东.我国驯鹿起源、历史变迁与现状[J].经济动物学报,2008,12(2):102-105.
[21]翟健程.使鹿部落驯鹿种群遗传多样性研究[D].哈尔滨:东北林业大学,2015.
[22]唐戈.鄂温克族驯鹿饲养业的困境与对策[J].黑龙江民族丛刊,2008,6:129-134.
[23]邢秀梅,苏伟林,杨福合.浅谈中国驯鹿种群[J].经济动物学报,2001,5(4):57-60.
[24]孙亚红.驯鹿发展管理模式探讨[J].中央民族大学学报(自然科学版),2013,22(2):42-44.
[25]曾治,孟凡露,王朋,等.我国驯鹿(Rangifer tarandus)的种群分布、数量及致危原因[J].西南大学学报(自然科学版),2016,38(7):19-24.
[26]殷亚杰.敖鲁古雅驯鹿(Rangifer tarandus)食性及其栖息地容纳量与种群生存力分析[D].哈尔滨:东北林业大学,2016.
[27]夏春,刘津,杨琪,等.猪干扰素β基因的分子克隆与测序[J].中国兽医杂志,2000,6:6-7.
[28]Hansen K,Prabakaran T,Laustsen A,et al.Listeria monocytogenes induces IFNβexpression through an IFI16-,cGAS-and STING-dependent pathway[J].Embo Journal,2014,33(15):1654.
[29]Basler C F,Garcíasastre A.Viruses and the typeⅠinterferon antiviral system:Induction and evasion[J].International Reviews of Immunology,2009,21(4-5):305.
[30]吴杨,杨苏河.β-干扰素的特性与临床应用[J].世界医学杂志,2001,3:72-74.
[31]Kunzi M S,Pitha P M.Interferon research:A brief history[J].Methods in Molecular Medicine,2005,116:25.
[32]Staples K J,Nicholas B,Mckendry R T,et al.Viral infection of human lung macrophages increases PDL1expression via IFNβ[J].PLoS One,2015,10(3):e0121527.
[33]Cheon H J,Holveybates E G,Schoggins J W,et al.IFNβ-dependent increases in STAT 1,STAT 2,and IRF9mediate resistance to viruses and DNA damage[J].Embo Journal,2013,32(20):2751-2763.