紫色番茄SlWD40-like基因克隆与表达分析
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摘要
WD-重复蛋白(WD-repeat protein)是一类含有WD40基序的蛋白,在植物响应非生物逆境胁迫的过程中具有重要调控作用。为揭示紫色番茄中WD40重复蛋白的功能,从富含花青素紫色番茄品种‘砚紫1号’中克隆 SlWD40-like基因,并对该基因在不同组织部位和不同胁迫条件下的表达模式进行分析。结果表明, SlWD40-like基因编码1个含有460个氨基酸的蛋白质。蛋白序列分析显示,SlWD40-like蛋白含有6个WD40基序,是典型的WD40重复蛋白。预测分析发现,该蛋白为亲水性蛋白,其二级结构以无规则卷曲为主;该蛋白序列在物种间具有高度的保守性,系统发生进化树分析表明紫番茄SlWD40-like氨基酸序列与潘那利番茄蛋白亲缘关系最近,其次是胡萝卜。应用qRT-PCR分析 SlWD40-like基因组织特异表达发现,该基因在叶和紫色果实中表达量最高。qRT-PCR分析表明, SlWD40-like基因受到盐胁迫、干旱胁迫脱落酸(ABA)和茉莉酸甲酯(MeJA)诱导表达,推测该基因可能参与紫色番茄逆境胁迫应答。该研究为后续进一步分析紫色番茄 SlWD40-like基因功能奠定了基础。
WD-repeat proteins contain WD40 motifs and play an important regulatory role in plant response to abiotic stress. In order to reveal the function of WD40 repeating proteins in purple tomato,the SlWD40-like gene was cloned from the variety ‘Yan Purple 1',and the expression pattern of SlWD40-like gene in different tissues and under different stress conditions was analyzed. The results showed that the SlWD40-like gene encodeed a protein containing 460 amino acids. Protein sequence analysis showed that the SlWD40-like protein contained 6 WD40 motifs and was a typical WD40 repeat protein. Prediction implied that the protein was hydrophilic protein and its secondary structure was dominated by irregular curl; the protein sequence was highly conserved among species. Phylogenetic tree analysis showed that the SlWD40-like amino acid sequence of purple tomato was closest to pennali tomato protein,followed by carrot. qRT-PCR analysis showed that the expression level of SlWD40-like gene was highest in leaves. qRT-PCR analysis showed that SlWD40-like gene was induced by drought,salt stress,abscisic acid and methyl jasmonate,and the gene might be involved in the response to the stress. So this paper have laid a foundation for further study in the function of SlWD40-like gene.
WD-repeat proteins contain WD40 motifs and play an important regulatory role in plant response to abiotic stress. In order to reveal the function of WD40 repeating proteins in purple tomato,the SlWD40-like gene was cloned from the variety ‘Yan Purple 1',and the expression pattern of SlWD40-like gene in different tissues and under different stress conditions was analyzed. The results showed that the SlWD40-like gene encodeed a protein containing 460 amino acids. Protein sequence analysis showed that the SlWD40-like protein contained 6 WD40 motifs and was a typical WD40 repeat protein. Prediction implied that the protein was hydrophilic protein and its secondary structure was dominated by irregular curl; the protein sequence was highly conserved among species. Phylogenetic tree analysis showed that the SlWD40-like amino acid sequence of purple tomato was closest to pennali tomato protein,followed by carrot. qRT-PCR analysis showed that the expression level of SlWD40-like gene was highest in leaves. qRT-PCR analysis showed that SlWD40-like gene was induced by drought,salt stress,abscisic acid and methyl jasmonate,and the gene might be involved in the response to the stress. So this paper have laid a foundation for further study in the function of SlWD40-like gene.
引文
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[21] 李洁.棉花 GhWD40基因的克隆和功能验证[D].武汉:华中农业大学,2013. LI J.Isolation and characterization of GhWD40 in cotton [D].Wuhan:Huazhong Agricultural University,2013.
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[23] 严莉,陈建伟,王翠平,等.黑果枸杞WD40编码基因 LrAN11的克隆及表达分析[J].核农学报,2017,31(11):2103-2112. YAN L,CHEN J W,WANG C P,et al.Cloning and expression analysis of LrAN11 encoding a WD40 protein in Lycium ruthenicum [J].Journal of Nuclear Agricultural Sciences,2017,31(11):2103-2112.
[24] 王国霞,张宁,杨玉珍,等.盐胁迫对红心萝卜种子萌发及幼苗生长的影响[J].西北农业学报,2016,25(5):744-749. WANG G X,ZHANG N,YANG Y ZH,et al.Effects of salt stress on seed germination and seedling growth of Raphanus sativas [J].Acta Agriculturae Boreali-occidentalis Sinica,2016,25(5):744-749.
[25] 薛泽民,薛琪,高景慧.渗透胁迫下紫花苜蓿生长及ABA受体基因的表达[J].西北农业学报,2018,27(3):331-338. XUE Z M,XUE Q,GAO J H.Growth and expression of abscisic acid receptor gene of alfalfa under osmotic stress [J].Acta Agriculturae Boreali-occidentalis Sinica,2018,27(3):331-338.
[26] 李国良,刘中华,许泳清,等.光温对叶菜型甘薯叶片花青素合成及相关酶基因表达的影响[J].核农学报,2017,31(1):8-13. LI G L,LIU ZH H,XU Y Q,et al.Effect of light and temperature on anthocyanin biosynthesis and expression of anthocyanin structural genes in leaf-vegetable sweet potato leaves [J].Journal of Nuclear Agricultural Sciences,2017,31(1):8-13.
[2] LEE J H,KIM W T.Regulation of abiotic stress signal transduction by E3 ubiquitin ligases in Arabidopsis [J].Molecules and Cells,2011,31(3):201-208.
[3] JAIN B P,PANDEY S.WD40 repeat proteins:signalling scaffold with diverse functions[J].The Protein Journal,2018,37(5):391-406.
[4] CHAO X,MIN J R.Structure and function of WD40 domain proteins [J].Protein and Cell,2011,2(3):202-214.
[5] MISHRA A K,PURANIK S,PRASAD M.Structure and regulatory networks of WD40 protein in plants [J].Journal of Plant Biochemistry and Biotechnology,2012,21(1):32-39.
[6] 华静静,陈晓静.植物体中WD40蛋白的研究进展[J].黑龙江农业科学,2015(5):153-156. HUA J J,CHEN X J.Progress of WD40 proteins in plants [J].Heilongjiang Agricultural Science,2015(5):153-156.
[7] VAN NOCKER S,LUDWIG P.The WD-repeat protein superfamily in Arabidopsis:conservation and divergence in structure and function [J].BMC Genomics,2003,4(1):50-61.
[8] BOUVERET R,SCH?NROCK N,GRUISSEM W,et al.Arabidopsis MSI1 functions in photoperiodic flowering time control [J].Frontiers in Plant Science,2014,5(3):77-87.
[9] TSUGAMA D,LIU S,TAKANO T.A bZIP protein,VIP1,interacts with Arabidopsis heterotrimeric G protein β subunit,AGB1 [J].Plant Physiology and Biochemistry,2013,71:240-246.
[10] HUANG J,WANG M M,BAO Y M,et al.SRWD:a novel WD40 protein subfamily regulated by salt stress in rice(Oryza sativa L.) [J].Gene,2008,424(1):71-79.
[11] 王佺珍,刘倩,高娅妮,等.植物对盐碱胁迫的响应机制研究进展[J].生态学报,2017,37(16):5565-5577. WANG Q ZH,LIU Q,GAO Y N,et al.Review on the mechanisms of the response to salinity-alkalinity stress in plants [J].Acta Ecologica Sinica,2017,37(16):5565-5577.
[12] ZHU Y,HUANG S,MIAO M,Tang X,et al.Genome-wide identification,sequence characterization,and protein-protein interaction properties of DDB1(damaged DNA binding protein-1)-binding WD40-repeat family members in Solanum lycopersicum[J].Planta,2015,241(6):1337-1350.
[13] 杨述章,高兰阳,孙晓春,等.过量表达 SlWD6基因增强番茄抗旱和耐盐功能[J].应用与环境生物学报,2015,21(3):413-420. YANG SH ZH,GAO L Y,SUN X CH,et al.Over-expressing SlWD6 gene to improve drought and salt tolerance of tomato[J].Chinese Journal of Applied and Environment Biology,2015,21(3):413-420.
[14] 余佳,苗敏,高兰阳,等.番茄 SlWDR141基因的克隆及功能研究[J].中国科技论文,2017,12(6):639-646. YU J,MIAO M,GAO L Y,et al.Cloning and functional study of tomato gene SlWDR141 [J].China Sciencepaper,2017,12(6):639-646.
[15] 陈林波,宋维希,李晓霞,等.茶树‘紫娟’肉桂酰辅酶A还原酶基因(CCR)克隆及序列分析[J].西北农业学报,2016,25(1):80-85. CHEN L B,SONG W X,LI X X,et al.Cloning and sequence analysis of Cinnamoyl-CoA Reductase gene from tea plant [Camellia sinensis(L.) O.Kuntz][J].Acta Agriculturae Boreali-occidentalis Sinica,2016,25(1):80-85.
[16] 屈霄霄,胡宗利,胡廷章,等.番茄转录辅激活子基因LeMBFI在非生物胁迫中抗性分析[J].生命科学研究,2010,14(4):322-326. QU X X,HU Z L,HU Y ZH,et al.Analysis of resistance to abiotic stresses of transcriptional co-activator gene LeMBF1 in tomato [J].Life Science Research,2010,14(4):322-326.
[17] 刘佳,徐秉良,薛应钰,等.美洲南瓜(Cucurbita pepo)种皮苯丙氨酸解氨酶基因克隆与表达分析[J].中国农业科学,2014,47(6):1216-1226. LIU J,XU B L,XUE Y J,et al.Cloning and expression analysis of PAL gene in seed coat of Cucurbita pepo [J].Scientia Agricultura Sinica,2014,47(6):1216-1226.
[18] 冯如,李泽宏,袁红慧,等.银杏两个WD40转录因子基因克隆及序列分析[J].北方园艺,2017(18):100-108. FENG R,LI Z H,YUAN H H,et al.Cloning and sequence analysis of two WD40 transcription factors in Ginkgo biloba L.[J].Northern Horticulture,2017(18):100-108.
[19] KRONZUCKER H J,COSKUN D,SCHULZE L M,et al.Sodium as nutrient and toxicant [J].Plant & Soil,2013,369:1-23.
[20] 李辉,李德芳,陈安国,等.盐和干旱胁迫下红麻 HcWD40-1基因的克隆及表达特征[J].农业生物技术学报,2017,25(12):1970-1978. LI H,LI D F,CHEN A G,et al.Cloning and expression characteristics of HcWD40-1 gene under salt and drought stress in Kenaf(Hibiscus cannabinus)[J].Journal of Agricultural Biotechnology,2017,25(12):1970-1978.
[21] 李洁.棉花 GhWD40基因的克隆和功能验证[D].武汉:华中农业大学,2013. LI J.Isolation and characterization of GhWD40 in cotton [D].Wuhan:Huazhong Agricultural University,2013.
[22] KONG D,LI M,DONG ZH,et al.Identification of TaWD- 40D,a wheat WD40 repeat-containing protein that is associated with plant tolerance to abiotic stresses [J].Plant Cell Reports,2015,34(3):395-410.
[23] 严莉,陈建伟,王翠平,等.黑果枸杞WD40编码基因 LrAN11的克隆及表达分析[J].核农学报,2017,31(11):2103-2112. YAN L,CHEN J W,WANG C P,et al.Cloning and expression analysis of LrAN11 encoding a WD40 protein in Lycium ruthenicum [J].Journal of Nuclear Agricultural Sciences,2017,31(11):2103-2112.
[24] 王国霞,张宁,杨玉珍,等.盐胁迫对红心萝卜种子萌发及幼苗生长的影响[J].西北农业学报,2016,25(5):744-749. WANG G X,ZHANG N,YANG Y ZH,et al.Effects of salt stress on seed germination and seedling growth of Raphanus sativas [J].Acta Agriculturae Boreali-occidentalis Sinica,2016,25(5):744-749.
[25] 薛泽民,薛琪,高景慧.渗透胁迫下紫花苜蓿生长及ABA受体基因的表达[J].西北农业学报,2018,27(3):331-338. XUE Z M,XUE Q,GAO J H.Growth and expression of abscisic acid receptor gene of alfalfa under osmotic stress [J].Acta Agriculturae Boreali-occidentalis Sinica,2018,27(3):331-338.
[26] 李国良,刘中华,许泳清,等.光温对叶菜型甘薯叶片花青素合成及相关酶基因表达的影响[J].核农学报,2017,31(1):8-13. LI G L,LIU ZH H,XU Y Q,et al.Effect of light and temperature on anthocyanin biosynthesis and expression of anthocyanin structural genes in leaf-vegetable sweet potato leaves [J].Journal of Nuclear Agricultural Sciences,2017,31(1):8-13.
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