文心兰生物钟相关基因PRR7克隆与表达分析
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摘要
利用转录组文库中的PRR7基因序列,从文心兰盛开期花瓣中扩增得到文心兰PRR7基因的2个成员,分别命名为OnPRR7-1(GenBank登录号:MG543993)和OnPRR7-2(GenBank登录号:MG543994)。OnPRR7-1和OnPRR7-2基因全长分别为2091 bp和2 154 bp,分别编码696和717个氨基酸大小的蛋白质序列。蛋白质二级结构分析表明,OnPRR7-1和OnPRR7-2均为疏水性蛋白。BlastX比对和系统发育分析表明,OnPRR7-1和OnPRR7-2基因和铁皮石斛和小兰屿蝴蝶兰的PRR基因同源性比较高。实时荧光定量PCR分析结果表明,OnPRR7-1和OnPRR7-2基因均呈现昼夜节律性表达格式,并且在正午12点时表达量达到峰值;在花发育和衰老过程中,OnPRR7-1和OnPRR7-2基因在花瓣中的表达量均在绽口期和衰老初期达到峰值,表明其有可能在花发育的开花和花衰老的过程中发挥作用。
Two members of Oncidium PRR7 gene family, OnPRR7-1 and OnPRR7-2, were isolated from the petals of the full-opened flowers based on the predicted PRR7 gene sequence through transcriptome analysis. The full-length of OnPRR7-1 and OnPRR7-2 was 2 091 bp and 2 154 bp in length, encoding peptides with 696 and 717 amino acids, respectively. Secondary structure modeling showed that these proteins were hydrophobic. BLASTX alignment and phylogenetic analysis showed that the OnPRR7-1 and OnPRR7-2 had high homology with the the counterparts of Dendrobium officinale and Phalaenopsis equestris. Real-time fluorescence quantitative PCR analysis showed that OnPRR7-1 and OnPRR7-2 genes both had a circadian expression pattern, with the expression levels peaked at 12:00 within 24 hours. During the flower development and senescence, the expression levels of OnPRR7-1 and OnPRR7-2 in petals reached the peak at the bud burst stage and the initial stage of senescence, suggesting that they play a role in flowering and flower senescence in Oncidium.
Two members of Oncidium PRR7 gene family, OnPRR7-1 and OnPRR7-2, were isolated from the petals of the full-opened flowers based on the predicted PRR7 gene sequence through transcriptome analysis. The full-length of OnPRR7-1 and OnPRR7-2 was 2 091 bp and 2 154 bp in length, encoding peptides with 696 and 717 amino acids, respectively. Secondary structure modeling showed that these proteins were hydrophobic. BLASTX alignment and phylogenetic analysis showed that the OnPRR7-1 and OnPRR7-2 had high homology with the the counterparts of Dendrobium officinale and Phalaenopsis equestris. Real-time fluorescence quantitative PCR analysis showed that OnPRR7-1 and OnPRR7-2 genes both had a circadian expression pattern, with the expression levels peaked at 12:00 within 24 hours. During the flower development and senescence, the expression levels of OnPRR7-1 and OnPRR7-2 in petals reached the peak at the bud burst stage and the initial stage of senescence, suggesting that they play a role in flowering and flower senescence in Oncidium.
引文
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[2]Harmer S L. The circadian system in higher plant[J]. Annual Review of Plant Biology, 2009, 60(1):357-377.
[3]徐小冬,谢启光.植物生物钟研究的历史回顾与最新进展[J].自然杂志, 2013, 35(2):118-126.
[4]王晗.生物钟生物学及其研究进展[J].生命科学,2015, 27(11):1 313-1 319.
[5]BendixC,MarshallCM,HarmonFG.Circadianclock genes universally control key agricultural traits[J]. Molecular Plant, 2015, 8(8):1 135-1 152.
[6]BeckwithEJ,YanovskyMJ.Circadianregulationofgene expression:atthecrossroadsoftranscriptionaland post-transcriptionalregulatorynetworks[J].CurrentOpinion in Genetics&Development, 2014, 27:35-42.
[7]Matsushika A, Makino S, Kojima M, et al. Circadian waves ofexpressionoftheAPRR1/TOC1familyofpseudo-responseregulatorsinArabidopsisthaliana:insightinto theplantcircadianclock[J].Plant&CellPhysiology,2000,41(9):1 002-1 012.
[8]FarréE M, Liu T. The PRR family of transcriptional regulatorsreflectsthecomplexityandevolutionofplantcircadian clocks[J].CurrentOpinioninPlantBiology, 2013,16(5):621-629.
[9]Mizuno T. Two-component phosphorelay signal transduction systemsinplants:fromhormoneresponsestocircadian rhythms[J]. Bioscience Biotechnology&Biochemistry, 2005,69:2 263-2 276.
[10]Mizuno T, Nakamichi N. Pseudo-response regulators(PRRs)or true oscillator components(TOCs)[J]. Plant&Cell Physiology, 2005, 46:677-685.
[11]LiY,XuM.CCTfamilygenesincerealcrops:Acurrent overview[J]. The Crop Journal, 2017, 5(6):449-458.
[12]KaczorowskiKA,QuailPH.ArabidopsisPSEUDO-RESPONSEREGULATOR7isasignalingintermediateinphytochrome-regulatedseedlingdeetiolationand phasingofthecircadianclock[J].PlantCell,2003,15(11):2 654-2 665.
[13]SaloméP A, Weigel D, Mcclung C R. The role of the Arabidopsis morning loop components CCA1, LHY, PRR7, and PRR9intemperaturecompensation[J].PlantCell,2010,22(11):3 650-3 661.
[14]刘晓荣,王碧青,朱根发.文心兰研究进展[J].亚热带植物科学, 2007, 36(3):85-90.
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[16]石乐松,孙慧,田晓岩,等.文心兰南茜种花发育进程与分期[J].热带生物学报, 2016, 7(4):440-443, 449.
[17]勾昕,胡薇薇,范亚飞,等.文心兰切花不同开放阶段花被的生理生化变化[J].热带生物学报, 2016, 7(1):70-75.
[18]黄浩宇,肖云骁,石乐松,等.文心兰切花花被衰老过程中酶活性变化研究.热带生物学报, 2016, 7(2):199-203
[19]YangGH,LiuJP.Isolationofan1-aminocyclopropane-1-carboxylatesynthasegenefromOncidiumGowerRamsey[J].GeneticsandMolecularResearch,2014,13(4):8 480-8 488
[20]杨光华,刘进平.文心兰ACC氧化酶基因OnACO1克隆与表达分析[J].热带作物学报, 2014, 35(4):693-699.
[21]Shi L S, Liu J P. Molecular cloning and expression analysis ofan1-aminocyclopropane-1-carboxylatesynthasegene fromOncidiumGowerRamsey[J].BiochemicalandBiophysical Research Communications, 2016, 469:203-209.
[22]田晓岩,石乐松,潘英文,等.文心兰OnACO2基因的克隆及表达分析[J].分子植物育种, 2015, 13(7):1 602-1 610.
[23]田晓岩,杨翠萍,胡进,等.文心兰OnERS1全长基因克隆及表达分析[J].分子植物育种,2017,15(4):1 265-1 272.
[24]徐张红,赵晓刚,何奕昆.拟南芥生物钟分子机制研究进展[J].植物学通报, 2005, 22(3):341-349.
[25]JohanssonM,StaigerD.Timetoflower:interplaybetween photoperiodandthecircadianclock[J].JournalofExperimental Botany, 2015, 66(3):719-730.
[26]SatbhaiSB.ThefamilyofPseudoResponseRegulator genes(PRR)in the moss Physcomitrella patens[D]. Nagoya:Nagoya University, 2011.
[27]MurakamiM,MatsushikaA,AshikariM,etal.Circadian-associated rice pseudo response regulators(OsPRRs):insight into the control of flowering time[J]. Bioscience, Biotechnology, and Biochemistry, 2005, 69(2):410-414.
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[29]MurakamiM,TagoY,YamashinoT,etal.Comparative overviews of clock-associated genes of Arabidopsis thalianaandOryzasativa[J].Plant&CellPhysiology,2007,48(1):110-121.
[30]Takata N, Saito S, Saito C T, et al. Phylogenetic footprint of theplantclocksysteminangiosperms:evolutionaryprocessesofpseudo-responseregulators[J].BMCEvolutionary Biology, 2010, 10:126.
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[32]MakinoS,KibaT,ImamuraA,etal.Genesencoding pseudo-responseregulators:insightintoHis-to-AspphosphorelayandcircadianrhythminArabidopsisthaliana[J].Plant&Cell Physiology, 2000, 41(6):791-803.
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[34]FarréEM,KaySA.PRR7proteinlevelsareregulatedby lightandthecircadianclockinArabidopsis[J].PlantJournal, 2007, 52(3):548-560.