番杏TtASR基因的克隆及其抗逆功能分析
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摘要
ASR(ABA, Stress and Ripening)蛋白是植物中特有的亲水性小分子蛋白,在植物细胞失水条件下对细胞器起保护性作用。本研究在构建番杏[Tetragonia tetragonioides (Pall.) Kuntze]全长cDNA文库中通过随机克隆测序分析基础上,获得了一个编码番杏ASR蛋白的全长cDNA序列,命名为TtASR(GenBank登录号:MH454101)。TtASR的cDNA全长序列包含一个为723 bp完整开放阅读框,编码蛋白TtASR含有240个氨基酸,等电点为5.26,分子量为25.29 ku。对TtASR与其他植物中同源蛋白的进化分析表明,与辽宁碱蓬SlASR和海蓬子SbASR-1亲缘关系较近。将TtASR的cDNA阅读框插入到原核表达载体pGEX6p-1中,通过转化大肠杆菌进行原核表达分析。结果表明,重组大肠杆菌菌株能表现出良好地抗逆性,特别是对NaCl、高渗透压和氧化胁迫的抗逆性增强。
ABA, stress and ripening(ASR) protein belongs to a plant-specific small family of proteins with low molecular weight and high hydrophobicity, and is supposed to play protective roles in plant cells under abiotic stresses. An ASR gene, designated as TtASR(GenBank accession no.: MH454101), was isolated from a cDNA library prepared from Tetragonia tetragonioides(Pall.) Kuntze and both the nucleotide and the deduced protein sequences were analyzed. The full length cDNA of TtASR contained a 723 bp long open reading frame, which would encode a peptide of 240 amino acid residues with a molecular weight of 25.29 ku and an isoelectric point of 5.26. TtASR shared a high amino acid sequence homology with the ASRs from Suaeda liaotungensis(SlASR) and Salicornia brachiata(SbASR-1). The open reading frame of TtASR was inserted into the prokaryotic expression vector of pGEX 6p-1 and transformed into E. coli.The E. coli cells under the induction of IPTG showed higher tolerance to NaCl, osmotic stress and oxidative stress than control cells.
ABA, stress and ripening(ASR) protein belongs to a plant-specific small family of proteins with low molecular weight and high hydrophobicity, and is supposed to play protective roles in plant cells under abiotic stresses. An ASR gene, designated as TtASR(GenBank accession no.: MH454101), was isolated from a cDNA library prepared from Tetragonia tetragonioides(Pall.) Kuntze and both the nucleotide and the deduced protein sequences were analyzed. The full length cDNA of TtASR contained a 723 bp long open reading frame, which would encode a peptide of 240 amino acid residues with a molecular weight of 25.29 ku and an isoelectric point of 5.26. TtASR shared a high amino acid sequence homology with the ASRs from Suaeda liaotungensis(SlASR) and Salicornia brachiata(SbASR-1). The open reading frame of TtASR was inserted into the prokaryotic expression vector of pGEX 6p-1 and transformed into E. coli.The E. coli cells under the induction of IPTG showed higher tolerance to NaCl, osmotic stress and oxidative stress than control cells.
引文
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[17]Jha B,Lal S,Tiwari V,et al.The SbASR-1 gene cloned from an extreme halophyte Salicornia brachiata enhances salt tolerance in transgenic tobacco[J].Mar Biotechnol(NY),2012,14(6):782-792.
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[19]Hu Y X,Yang X,Li X L,et al.The SlASR gene cloned from the extreme halophyte Suaeda liaotungensis K.enhances abiotic stress tolerance in transgenic Arabidopsis thaliana[J].Gene,2014,549:243-251.
[20]Li J,Dong Y,Li C,et al.SiASR4,the target gene of SiARDPfrom Setaria italica,improves abiotic stress adaption in plants[J].Frontiers in Plant Science,2017,7:2053.
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[22]Czolpinska M,Rurek M.Plant glycine-rich proteins in stress response:an emerging,still prospective story[J].Frontiers in Plant Science,2018,9:2.
[23]Mangeon A,Junqueira R M,Sachetto-Martins G.Functional diversity of the plant glycine-rich proteins superfamily[J].Plant Signaling&Behavior,2010,5(2):99-104.
[24]Kim Y O,Pan S,Jung C H,et al.A zinc finger-containing glycine-rich RNA-binding protein,at RZ-1a,has a negative impact on seed germination and seedling growth of Arabidopsis thaliana under salt or drought stress conditions[J].Plant Cell and Physiology,2007,48(8):1170-1181.
[25]Jabeen B,Naqvi SM,Mahmood T,et al.Ectopic expression of plant RNA chaperone offering multiple stress tolerance in E.coli[J].Molecular Biotechnology,2017,59(2-3):66-72.
[26]Saavedra L,Svensson J,Carballo V,et al.A dehydrin gene in Physcomitrella patens is required for salt and osmotic stress tolerance[J].The Plant Journal,2006,45(2):237-249.
[2]林更铭,骆建宁,项鹏.利用冬闲海水养殖池塘种植番杏的初步探讨[J].农业科技通讯,2014,11:117-119.
[3]王蕾,吴朝波,徐微风,等.海水胁迫对番杏生长、渗透调节物质和细胞膜透性的影响[J].江苏农业科学,2016,44(7):248-251.
[4]Wilson C,Lesch S M,Grieve C M.Growth stage modulates salinity tolerance of New Zealand spinach(Tetragonia tetragonioides,Pall.)and red orach(Atriplex hortensis L.)[J].Annals of Botany,2000,85:501-509.
[5]Mishra A,Tanna B.Halophytes:potential resources for salt stress tolerance genes and promoters[J].Frontiers in Plant Science,2017,8:829.
[6]王永泰.盐土植物和盐土改良[J].生物学通报,1991,6:12-17.
[7]Roy S J,Negr?o S,Tester M.Salt resistant crop plants[J].Current Opinion in Biotechnology,2014,26:115-124.
[8]Munns R,Tester M.Mechanisms of salinity tolerance[J].Annual Review of Plant Biology,2008,59:651-681.
[9]Zhu J K.Salt and drought stress signal transduction in plants[J].Annual Review of Plant Biology,2002,53:247-273.
[10]González R M,Iusem N D.Twenty years of research on Asr(ABA-stress-ripening)genes and proteins[J].Planta,2014,239:941-949.
[11]Frankel N,Carrari F,Hasson E,et al.Evolutionary history of the Asr gene family[J].Gene,2006,378:74-83.
[12]Golan I,Dominguez P G,Konrad Z,et al.Tomato abscisic acid stress ripening(ASR)gene family revisited[J].PLo SONE,2014,9:e107117.
[13]Canel C,Bailey-Serres J N,Roose M L.Pummelo fruit transcript homologous to ripening-induced genes[J].Plant Physiology,1995,108:323-1324.
[14]Dominguez P G,Carrari F.ASR1 transcription factor and its role in metabolism[J].Plant Signaling&Behavior,2015,10:e992751.
[15]Yang C Y,Chen Y C,Jauh G Y,et al.A Lily ASR protein involves abscisic acid signaling and confers drought and salt resistance in Arabidopsis[J].Plant Physiology,2005,139(2):836-846.
[16]Dai J R,Liu B,Feng D R,et al.MpAsr encodes an intrinsically unstructured protein and enhances osmotic tolerance in transgenic Arabidopsis[J].Plant Cell Reports,2011,30:1219-1230.
[17]Jha B,Lal S,Tiwari V,et al.The SbASR-1 gene cloned from an extreme halophyte Salicornia brachiata enhances salt tolerance in transgenic tobacco[J].Mar Biotechnol(NY),2012,14(6):782-792.
[18]Tiwari V,Chaturvedi A K,Mishra A,et al.Introgression of the SbASR-1 gene cloned from a halophyte Salicornia brachiate enhances salinity and drought endurance in transgenic groundnut(Arachis hypogaea)and acts as a transcription factor[J].PLoS ONE,2015,10:e0131567.
[19]Hu Y X,Yang X,Li X L,et al.The SlASR gene cloned from the extreme halophyte Suaeda liaotungensis K.enhances abiotic stress tolerance in transgenic Arabidopsis thaliana[J].Gene,2014,549:243-251.
[20]Li J,Dong Y,Li C,et al.SiASR4,the target gene of SiARDPfrom Setaria italica,improves abiotic stress adaption in plants[J].Frontiers in Plant Science,2017,7:2053.
[21]Glenn E P,Brown J J,Blumwald E.Salt tolerance and crop potential of halophytes[J].Critical Reviews in Plant Sciences,1999,18:227-255.
[22]Czolpinska M,Rurek M.Plant glycine-rich proteins in stress response:an emerging,still prospective story[J].Frontiers in Plant Science,2018,9:2.
[23]Mangeon A,Junqueira R M,Sachetto-Martins G.Functional diversity of the plant glycine-rich proteins superfamily[J].Plant Signaling&Behavior,2010,5(2):99-104.
[24]Kim Y O,Pan S,Jung C H,et al.A zinc finger-containing glycine-rich RNA-binding protein,at RZ-1a,has a negative impact on seed germination and seedling growth of Arabidopsis thaliana under salt or drought stress conditions[J].Plant Cell and Physiology,2007,48(8):1170-1181.
[25]Jabeen B,Naqvi SM,Mahmood T,et al.Ectopic expression of plant RNA chaperone offering multiple stress tolerance in E.coli[J].Molecular Biotechnology,2017,59(2-3):66-72.
[26]Saavedra L,Svensson J,Carballo V,et al.A dehydrin gene in Physcomitrella patens is required for salt and osmotic stress tolerance[J].The Plant Journal,2006,45(2):237-249.