不同启动子驱动下GL6基因表达对水稻叶表皮毛发育的影响
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摘要
【目的】研究水稻茸毛发育机制及相关基因的功能与调控模式,为深入研究相关基因功能及其在生产上的应用提供理论支撑。【方法】从不同水稻品种中克隆叶片表皮毛发育相关基因GL6的启动子序列,并将具有显著表皮毛特征的突变体品种75-1-127的GL6基因启动子与叶表无显著表皮毛特征的野生型品种相应基因的启动子序列进行比对,同时克隆突变体品种75-1-127中GL6基因的CDS序列,并分别构建以玉米泛素蛋白Ubiquitin和花椰菜花叶病毒CaMV35S为启动子驱动的过表达载体,以农杆菌介导的方法转化野生型粳稻品种Kitaake。【结果】不同品种中克隆的启动子序列区存在显著的序列差异,以玉米泛素蛋白Ubiquitin启动子驱动的过表达载体获得的转基因水稻出现了显著的表皮毛特征,以花椰菜花叶病毒CaMV35S为启动子驱动的过表达载体的转基因水稻则未出现典型的表皮毛特征。【结论】目标基因GL6的表达调控受启动子的影响,突变体品种75-1-127的叶表皮毛发育特征是因启动子区序列差异所致。
【Objective】 Being one of the important agronomic traits that reflect the yield and physiological functions of a rice plant, the growth and photosynthesis of the fine hairs on the surface of rice leaves were studied. 【Method】 The sequences of GL6 gene promoters relating to leaf epidermal hair development were obtained and cloned from different rice varieties. That of Mutant 75-1-127 showing the characteristic epidermal hair development was compared with those of the wild type rice lacking it. The CDS sequence of the mutant gene was cloned. Then, the maize Ubiquitin and cauliflower mosaic virus CaMV35 S were used as promoter-driven overexpression vectors to transform the wild-type Japonica rice, Kitaake, mediated by Agrobacterium. 【Result】There were significant differences on the sequences in regions of the genes regulated by the promoters and cloned from different cultivars. The leaves of transgenic rice plants with GL6 driven by maize Ubiquitin promoter had significant hairy phenotype of the typical velvet appearance, but not those involved CaMV35S as a promoter. 【Conclusion】It was clearly demonstrated that the promoters affected the expression of the target gene GL6 and that the epidermal hair development on leaves of Mutant 75-1-127 was derived by the promoter with a unique sequence.
【Objective】 Being one of the important agronomic traits that reflect the yield and physiological functions of a rice plant, the growth and photosynthesis of the fine hairs on the surface of rice leaves were studied. 【Method】 The sequences of GL6 gene promoters relating to leaf epidermal hair development were obtained and cloned from different rice varieties. That of Mutant 75-1-127 showing the characteristic epidermal hair development was compared with those of the wild type rice lacking it. The CDS sequence of the mutant gene was cloned. Then, the maize Ubiquitin and cauliflower mosaic virus CaMV35 S were used as promoter-driven overexpression vectors to transform the wild-type Japonica rice, Kitaake, mediated by Agrobacterium. 【Result】There were significant differences on the sequences in regions of the genes regulated by the promoters and cloned from different cultivars. The leaves of transgenic rice plants with GL6 driven by maize Ubiquitin promoter had significant hairy phenotype of the typical velvet appearance, but not those involved CaMV35S as a promoter. 【Conclusion】It was clearly demonstrated that the promoters affected the expression of the target gene GL6 and that the epidermal hair development on leaves of Mutant 75-1-127 was derived by the promoter with a unique sequence.
引文
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[13]BERNHARDT C,LEE M M,GONZALEZ ,et al.ThebHLH genes GLABRA3 (GL3) and ENHANCER OF GLABRA3 (EGL3) specify epidermal cell fate in the Arabidopsis root[J].Development,2003,130:6431-6439.
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[16]SZYMANSKI D B,JILK R A,POLLOCK S M,et al.Control of GL2 expression in Arabidopsis leaves and trichomes[J].Development,1998,125:1161-1171.
[17]SUN W Q,GAO D W ,XIONG Y,et al.Hairy Leaf 6,an AP2/ERF Transcription Factor,Interacts with OsWOX3B and Regulates Trichome Formation in Rice[J].Molecular Plant,2017,10(11):1417-1433.
[18]LI J J ,YUAN Y D,LU Z F,et al.Glabrous Rice 1,encoding a homeodomainprotein,regulates trichome development in rice[J].Rice,2012,5(32):1-10.
[19]QIN B,TANG D,HUANG J,et al.Rice OsGL1-1 is involved in leaf cuticular wax and cuticle membrane[J].Mol Plant,2011,4:985-995.
[20]曾跃辉,朱永生,连玲,等.水稻茸毛基因GL6的遗传学分析与精细定位[J].科学通报,2013,58(1):1027-1035.ZENG Y H,ZHU Y S,LIAN L,et al.Genetic analysis and fine mapping of the pubescence gene GL6 in rice (Oryzae sativa L.)[J].Chinese Science Bulletin,2013,58(1):1-8.(in Chinese)
[21]ZHENG K,TIAN H,HU Q ,et al.Ectopic expression of R3MYB transcription factor gene OsTCL1 in Arabidopsis,but not rice,affects trichome and root hair formation[J].Sci Rep,2016,6:19254.
[22]KASUGA M,LIU Q,MIURA S,et al.Improving plant drought,salt,and freezing tolerance by gene transfer of a single stress-inducible transcription factor[J].Nat biotechnol,1999,17(3):287-291
[23]乔龙飞,于延冲.LEAFY启动子中W-box对LEAFY表达及拟南芥开花的影响[J].植物生理学报,2017,53 (4):713-720.QIAO L F,YU Y C.Effect of W-box in LEAFY promoter on LEAFY expression and Arabidopsis flowering[J].Plant Physiology Journal,2017,53 (4):713-720.(in Chinese)
[24]陈兆骞,陈熙,张炜,不同启动子在水稻悬浮细胞中诱导外源基因的表达,江苏农业学报[J],2014,30(6):1208-1215.CHEN Z Q ,CHEN X,ZHANG W,et al.Promoter-induced expression of exogenous gene in rice suspension cell lines[J].Jiangsu Journal of Agricultural Sciences,2014,30(6):1208-1215.(in Chinese)
[2]LAUTER D J,MUNNS D N.Water loss via the glandular trichomes of chickpea (Cicer arietinum L.) [J].J Exp Bot,1986,37:640-649.
[3]YAN A,PAN J,AN L,et al.The responses of trichome mutants to enhanced ultraviolet-B radiation in Arabidopsis thaliana[J].J PhotochemPhotobiol B Biol,2012,113:29-35.
[4]TATTINI M,GRAVANO E,PINELLI P,et al.Flavonoids accumulate in leaves and glandular trichomes of Phillyrealatifolia exposed to excess solar radiation[J].New Phytol,2000,148:69-77.
[5]尚宏芹,刘建萍.干旱胁迫下不同茸毛性状辣椒植株抗旱性比较[J].核农学报,2010,24(4):835-839.SHANG H Q,LIU J P.Comparison of drought resistance of pepper with different hairiness character under drought stress[J].Journal of Nuclear Agricultural Sciences,2010,24(4):835-839.(in Chinese)
[6]CASTILLO-LóPEZ J L,CANO-SANTANA Z,OYAMA K.Preferences and survival of Lophoceramicapyrrha,a shelter builder gregarious noctuid,in two host plants[J].Dugesiana,2010,17:229-236.
[7]DALIN P,GREN J,BJRKMAN C,et al.Chapter 4:Leaf trichome formation and plant resistance to herbivory.Induced Plant Resistance to Herbivory[M].Netherlands:Springer,2008:89-105.
[8]MCGINNIS A J,KASTING R.Dietary cellulose:Effect on food consumption and growth of a grasshopper[J].Ca J Zool,2011,45:165-167.
[9]HANDLEY R,EKBOM B,AGREN J.Variation in trichome density and resistance against a specialist insect herbivore in natural populations of Arabidopsis thaliana[J].EcolEntomol,2005,30:284-292.
[10]AN L,ZHOU Z,SU S,et al.GLABROUS INFLORESCENCE STEMS (GIS) is required for trichome branching through gibberellic acid signaling in Arabidopsis[J].Plant Cell Physiol,2012,53:457.
[11]KHOSLA A,PAPER J M,BOEHLER A P,et al.HD-Zip proteins GL2 and HDG11 have redundant functions in Arabidopsis trichomes and GL2 activates a positive feedback loop via MYB23[J].Plant Cell,2014,26:2184-2200.
[12]MOROHASHI K,ZHAO M,YANG M,et al.Participation of the ArabidopsisbHLH factor GL3 in trichome initiation regulatory events[J].Plant Physiol,2007,145:736-746.
[13]BERNHARDT C,LEE M M,GONZALEZ ,et al.ThebHLH genes GLABRA3 (GL3) and ENHANCER OF GLABRA3 (EGL3) specify epidermal cell fate in the Arabidopsis root[J].Development,2003,130:6431-6439.
[14]CHOPRA D,WOLFF H,SPAN J,et al.Analysis of TTG1 function in Arabisalpina[J].BMC Plant Biol,2014,14:16.
[15]LARKIN J C,OPPENHEIMER D G,MARKS M D,et al.The GL1 gene and the trichome developmental pathway in Arabidopsis thaliana[J].Results Probl Cell Differ,1994,20:259-275.
[16]SZYMANSKI D B,JILK R A,POLLOCK S M,et al.Control of GL2 expression in Arabidopsis leaves and trichomes[J].Development,1998,125:1161-1171.
[17]SUN W Q,GAO D W ,XIONG Y,et al.Hairy Leaf 6,an AP2/ERF Transcription Factor,Interacts with OsWOX3B and Regulates Trichome Formation in Rice[J].Molecular Plant,2017,10(11):1417-1433.
[18]LI J J ,YUAN Y D,LU Z F,et al.Glabrous Rice 1,encoding a homeodomainprotein,regulates trichome development in rice[J].Rice,2012,5(32):1-10.
[19]QIN B,TANG D,HUANG J,et al.Rice OsGL1-1 is involved in leaf cuticular wax and cuticle membrane[J].Mol Plant,2011,4:985-995.
[20]曾跃辉,朱永生,连玲,等.水稻茸毛基因GL6的遗传学分析与精细定位[J].科学通报,2013,58(1):1027-1035.ZENG Y H,ZHU Y S,LIAN L,et al.Genetic analysis and fine mapping of the pubescence gene GL6 in rice (Oryzae sativa L.)[J].Chinese Science Bulletin,2013,58(1):1-8.(in Chinese)
[21]ZHENG K,TIAN H,HU Q ,et al.Ectopic expression of R3MYB transcription factor gene OsTCL1 in Arabidopsis,but not rice,affects trichome and root hair formation[J].Sci Rep,2016,6:19254.
[22]KASUGA M,LIU Q,MIURA S,et al.Improving plant drought,salt,and freezing tolerance by gene transfer of a single stress-inducible transcription factor[J].Nat biotechnol,1999,17(3):287-291
[23]乔龙飞,于延冲.LEAFY启动子中W-box对LEAFY表达及拟南芥开花的影响[J].植物生理学报,2017,53 (4):713-720.QIAO L F,YU Y C.Effect of W-box in LEAFY promoter on LEAFY expression and Arabidopsis flowering[J].Plant Physiology Journal,2017,53 (4):713-720.(in Chinese)
[24]陈兆骞,陈熙,张炜,不同启动子在水稻悬浮细胞中诱导外源基因的表达,江苏农业学报[J],2014,30(6):1208-1215.CHEN Z Q ,CHEN X,ZHANG W,et al.Promoter-induced expression of exogenous gene in rice suspension cell lines[J].Jiangsu Journal of Agricultural Sciences,2014,30(6):1208-1215.(in Chinese)
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