油菜素内酯合成酶(Steroid 5α-Reductase)基因的超量表达对毛白杨生长的影响

将棉花Steroid 5α—reductase基因（DET2）超量表达载体导入毛白杨中,观察其对毛白杨生长和芽休眠的影响。结果表明,超量表达DET2基因的毛白杨茎高度、直径生长和不定根的生长增强,芽的休眠破除提前。     

棉花赤霉素氧化酶同源基因GhGA20ox1在烟草中的功能表达

为研究棉花GA20-氧化酶同源基因GhGA20ox1的功能,将该基因转入本明烟（N．benthamiana）中进行超量表达。RT-PCR分析表明GhGA20ox1基因在转基因植株中得到了不同水平的表达。GhGA20ox1基因的超量表达促进了本明烟中的GA4＋7合成,并导致赤霉素过量的表型出现。转基因本明烟的表型变化程度与GhGA20ox1基因的表达水平和GA4＋7的含量一致。这些结果表明,GhGA20ox1基因编码一个有功能的GA20-氧化酶,能够在转基因烟草中促进活性GA（GA4＋7）的合成,可以用作目的基因来提高棉花纤维和其他植物的内源GA水平。     

高等植物赤霉素生物合成关键组分GA20-oxidase氧化酶基因的研究进展北大核心CSCD

GA-20氧化酶(GA-20 oxidase)是重要的GA生物合成和调控酶,直接催化生成有生物活性的GAs,是一种多功能酶,最显著的特点就是负反馈调节。GA20-氧化酶在植物发育和生理过程中起着重要的调控作用。综述了高等植物体内GA20氧化酶基因的克隆及表达调控研究及其对株高、纤维、开花、产量性状等影响,重点阐述了GA20氧化酶基因与激素、光周期、抗性等之间的相互作用,以便更好地揭示GA-20氧化酶信号网络系统及其作用机制。     

高等植物赤霉素生物合成关键组分GA20-oxidase氧化酶基因的研究进展

GA-20氧化酶(GA-20 oxidase)是重要的GA生物合成和调控酶,直接催化生成有生物活性的GAs,是一种多功能酶,最显著的特点就是负反馈调节。GA20-氧化酶在植物发育和生理过程中起着重要的调控作用。综述了高等植物体内GA20氧化酶基因的克隆及表达调控研究及其对株高、纤维、开花、产量性状等影响,重点阐述了GA20氧化酶基因与激素、光周期、抗性等之间的相互作用,以便更好地揭示GA-20氧化酶信号网络系统及其作用机制。     

高等植物GA 20-氧化酶研究进展

GA20-氧化酶（GA 20-oxidase）是高等植物体内GA生物合成途径中最重要的限速酶,它能够催化从GA12到GA9及GA53到GA20-系列的氧化反应。作者对近年来有关GA20一氧化酶的特征,GA20一氧化酶基因及编码蛋白、GA20一氧化酶基因表达调控进行了综合评述,并对GA20一氧化酶基因转化在农业、林业、;园艺业方面的应用前景进行了展望。     

拟南芥突变体cka3-2的筛选及其功能初步研究

CKA3-2基因属于拟南芥CK基因家族成员,该基因编码一条691个氨基酸残基的多肽.利用三引物法和实时定量PCR法鉴定获得CKA3-2基因对应的T-DNA插入纯合突变体cka3-2,并对其表T型变化进行了观察.实时定量PCR分析表明:CKA3-2基因在茎生叶中表达最高,花和茎中次之,在根、莲座叶和果荚中表达量较低.赤霉素使CKA3-2基因的表达升高,6 h时表达量明显增加,8 h时表达量下降.用50μmol/L GA3处理Col-0和cka3-2幼苗6 h,结果发现赤霉素信号转导通路相关基因如KS、KAO1、KAO2、GA2ox1、GA3OX1和GA20OX1等在两种幼苗中的相对表达量发生明显变化.此外,突变体cka3-2相对野生型Col-0来说,具有较明显的早花现象.     

荔波连蕊茶GA20氧化酶基因的克隆及表达分析

根据植物GA20ox基因编码区的保守序列设计引物,以山茶属荔波连蕊茶幼嫩茎段为材料,提取总RNA,进行RT-PCR。采用RACE技术扩增获得1 567 bp的GA20氧化酶基因全长cDNA序列,命名为ClGA20ox2(GenBank登录号KF823787)。序列分析表明,ClGA20ox2开放阅读框(ORF)为1 146 bp,编码382个氨基酸,5'非编码区115 bp,3'非编码区303 bp。预测的蛋白质分子量为43.56 kD,等电点为7.02,所推导的蛋白氨基酸序列与夹竹桃和杨树GA20ox蛋白的同源性分别为73%和72%。ClGA20ox2与其它植物GA20ox蛋白比较,构建系统进化树,结果显示山茶GA20ox蛋白与夹竹桃和杨树的GA20ox蛋白的亲缘关系最为密切。实时定量PCR结果显示,该基因在荔波连蕊茶的根、茎、叶和种子中均有表达,其表达模式却不同:ClGA20ox2基因在二年生茎段中的表达丰度最高,在顶端分生组织中表达丰度最低,在嫩叶和根中表达丰度较高,成熟叶片和种子表达丰度较低。     

高等植物赤霉素2-氧化酶基因的克隆、表达及其调控

赤霉素(GA)是一类重要的植物激素,对高等植物整个生命周期的生长发育起关键作用。调控赤霉素生物合成和代谢途径中的关键酶基因的表达可以控制植物体内赤霉素的含量。GA2-氧化酶是调节赤霉素合成和代谢的关键酶之一,使活性GA失活。本文主要对GA2-氧化酶基因的克隆、表达调控及其在植物基因工程中的应用等方面进行综述,为通过基因工程技术调控植物体内活性赤霉素的含量从而得到改良品种提供思路。     

小桐子赤霉素20氧化酶的基因克隆及低温下表达分析

赤霉素20氧化酶是植物赤霉素生物合成的限速酶,决定有生物活性的GA1与GA4的合成量。基于先前获得的小桐子低温锻炼转录组数据,以小桐子幼苗的根为材料,采用RT-PCR技术克隆到小桐子赤霉素20氧化酶基因Jc GA20ox的c DNA序列(Gen Bank登录号KJ670150.1)。该c DNA全长1 307 bp,含有完整的开放阅读框(1 131 bp),编码376个氨基酸,分子量为43 k D,理论等电点为6.7。其推导蛋白属于2-ODD家族,包含2-酮戊二酸双加氧酶结构域(Fe2OG_OXY)。半定量RT-PCR表达分析显示,Jc GA20ox在小桐子各组织中都有表达,但表达水平具有组织特异性,在茎中表达量较高,且受低温诱导表达最显著,而在叶中表达量相对较低。     

烟草GA合成调控转录因子RSG应答甲醇和乙醇刺激的分子机理初探

为了考察甲醇或乙醇促进植物生长与赤霉素(GA)的合成关系,该研究在MS固体培养基中培养并添加外源GA和GA合成抑制剂多效唑(PAC),分析其对2mmol/L甲醇或乙醇促进烟草生长的影响及GA合成调控转录因子RSG(for repression of shoot growth)应答甲醇或乙醇刺激的分子机理。结果显示:(1)外源添加GA可增强甲醇或乙醇对烟草生长的刺激作用,而添加PAC却抑制甲醇和乙醇对烟草生长的刺激作用。(2)14-3-3蛋白与RSG结合抑制RSG进入细胞核及其转录调控活性;甲醇和乙醇诱导烟草14-3-3基因的转录和表达,对RSG蛋白表达也有诱导作用。(3)甲醇和乙醇可降低14-3-3蛋白与RSG的相互作用,同时增强RSG与GA20ox1启动子的结合。研究表明,甲醇和乙醇刺激烟草的生长可能通过增加RSG表达,且减弱RSG与14-3-3蛋白的结合来增加RSG细胞核定位作用,从而增强RSG与GA20ox1启动子的结合,最终增加GA的合成,从而促进烟草的生长,这可能是甲醇和乙醇促进烟草生长的一种重要的分子机制。     

Expression of a gibberellin 2-oxidase gene around the shoot apex is related to phase transition in rice

A major catabolic pathway for gibberellin (GA) is initiated by 2beta-hydroxylation, a reaction catalyzed by GA 2-oxidase. We have isolated and characterized a cDNA, designated Oryza sativa GA 2-oxidase 1 (OsGA2ox1) from rice (Oryza sativa L. cv Nipponbare) that encodes a GA 2-oxidase. The encoded protein, produced by heterologous expression in Escherichia coli, converted GA(1), GA(4), GA(9), GA(20), and GA(44) to the corresponding 2beta-hydroxylated products GA(8), GA(34), GA(51), GA(29), and GA(98), respectively. Ectopic expression of the OsGA2ox1 cDNA in transgenic rice inhibited stem elongation and the development of reproductive organs. These transgenic plants were deficient in endogenous GA(1). These results indicate that OsGA2ox1 encodes a GA 2-oxidase, which is functional not only in vitro but also in vivo. OsGA2ox1 was expressed in shoot apex and roots but not in leaves and stems. In situ hybridization analysis revealed that OsGA2ox1 mRNA was localized in a ring at the basal region of leaf primordia and young leaves. This ring-shaped expression around the shoot apex was drastically decreased after the phase transition from vegetative to reproductive growth. It was absent in the floral meristem, but it was still present in the lateral meristem that remained in the vegetative phase. These observations suggest that OsGA2ox1 controls the level of bioactive GAs in the shoot apical meristem; therefore, reduction in its expression may contribute to the early development of the inflorescence meristem.     

Daylength and spatial expression of a gibberellin 20-oxidase isolated from hybrid aspen (Populus tremula L. × P. tremuloides Michx.)

Physiologically active gibberellins (GAs) are key regulators of shoot growth in trees. To investigate this mechanism of GA-controlled growth in hybrid aspen, we cloned cDNAs encoding gibberellin 20-oxidase (GA 20-oxidase), a key, highly regulated enzyme in the biosynthesis of GAs. Clones were isolated from leaf and cambium cDNA libraries using probes generated by polymerase chain reaction, based on conserved domains of GA 20-oxidases. Upon expression in Escherichia coli, the GST-fusion protein was shown to oxidise GA12 as well as oxidising the 13-hydroxylated substrate GA53, successively to GA9 and GA20, respectively. The gene PttGA20ox1 was expressed in meristematic cells and growing tissues such as expanding internodes, leaves and roots. The expression was negatively regulated by both GA4 and overexpression of phytochrome A. RNA analysis also showed that the expression was down-regulated in late-expanding leaf tissue in response to short days (SDs). Actively growing tissues such as early elongating internodes, petioles and leaf blades had the highest levels of C19-GAs. Upon transfer to SDs an accumulation of GA19 was observed in early elongating internodes and leaf blades. The levels of C19-GAs were also to some extent changed upon transfer to SDs. The levels of GA20 were down-regulated in internodes, and those of GA1 were significantly reduced in early expanding leaf blades. In roots the metabolites GA19 and GA8 decreased upon shifts to SDs, while GA20 accumulated slightly. The down-regulation of GA 20-oxidase activity in response to SDs was further indicated by studies of [14C]GA12 metabolism in shoots, demonstrating that the substrate for GA 20-oxidase, [14C]GA53, accumulates in SDs.     

Dissection of GA 20-oxidase members affecting tomato morphology by RNAi-mediated silencing

GA 20-oxidase is a key enzyme involved in gibberellin (GA) biosynthesis. In tomato, the GA 20-oxidase gene family consists of three members: GA20ox1, GA20ox2, and GA20ox3. To investigate the roles of these three genes in regulating plant growth and development, we used RNA interference technology to generate three kinds of transgenic tomato plants with suppressed expression of each three individual genes. Suppression of GA20ox1 or GA20ox2 resulted in shorter stems, a decreased length of internodes, and small dark green leaves while plants with decreased expression of GA20ox3 had no visible changes on stems and leaves. The plants of the three transgenic lines can flower and set fruits normally, but the seeds from these plants germinated slower than that from the normal plants. Decreased levels of endogenous GAs were detected in the apex of the three transgenic lines. These results demonstrate that the three GA 20-oxidase genes play different roles in the control of plan vegetative growth, but show no effects on flower and fruit development.Equal contribution authors: J. Xiao and H. Li.     

Changes in GA 20-oxidase gene expression strongly affect stem length, tuber induction and tuber yield of potato plants

Gene StGA20ox1 encoding potato GA 20-oxidase is expressed to relatively high levels in leaves and regulated by daylength. To investigate whether this gene is involved in photoperiodic regulation of tuber formation, we have obtained transgenic potato plants expressing sense and antisense copies of the StGA20ox1 cDNA. Over-expression of this cDNA resulted in taller plants that required a longer duration of a short day photoperiod (SD) to tuberize. Tubers from these plants had a decreased time of dormancy and developed sprouts with elongated internodes. Plants expressing antisense copies of the StGA20ox1 cDNA had shorter stems, a decreased length of the internodes and tuberized earlier than control plants, showing increased tuber yields. Antisense inhibition of this gene had no visible effect on the time of dormancy of the tubers, although at the end of dormancy these formed sprouts with shortened internodes. Decreased levels of endogenous GA20 and GA1 were detected in the apex and first leaves of the antisense lines. These results demonstrate the involvement of the GA 20-oxidase activity encoded by StGA20ox1 in the control of stem elongation and in tuber induction but not in tuber dormancy, indicating that the latter may be regulated by another member of the gene family.     

Production of Dwarf Lettuce by Overexpressing a Pumpkin Gibberellin 20-Oxidase Gene

We investigated the effect of overexpressing a pumpkin gibberellin (GA) 20-oxidase gene encoding an enzyme that forms predominantly biologically inactive products on GA biosynthesis and plant morphology in transgenic lettuce (Lactuca sativa cv Vanguard) plants. Lettuce was transformed with the pumpkin GA 20-oxidase gene downstream of a strong constitutive promoter cassette (El2-35S-Omega). The transgenic plants in which the pumpkin gene was detected by polymerase chain reaction were dwarfed in the T(2) generation, whereas transformants with a normal growth phenotype did not contain the transgene. The result of Southern-blot analysis showed that the transgene was integrated as a single copy; the plants segregated three dwarfs to one normal in the T(2) generation, indicating that the transgene was stable and dominant. The endogenous levels of GA(1) and GA(4) were reduced in the dwarfs, whereas large amounts of GA(17) and GA(25), which are inactive products of the pumpkin GA 20-oxidase, accumulated in these lines. These results indicate that a functional pumpkin GA 20-oxidase is expressed in the transgenic lettuce, resulting in a diversion of the normal pathway of GA biosynthesis to inactive products. Furthermore, this technique may be useful for controlling plant stature in other agricultural and horticultural species.     

Modification of gibberellin production and plant development in Arabidopsis by sense and antisense expression of gibberellin 20-oxidase genes

Gibberellin (GA) 20-oxidase catalyses consecutive steps late in GA biosynthesis in plants. In Arabidopsis, the enzyme is encoded by a gene family of at least three members (AtGA20ox1, AtGA20ox2 and AtGA20ox3) with differential patterns of expression. The genes are regulated by feedback from bioactive GAs, suggesting that the enzymes may be involved in regulating GA biosynthesis. To investigate this, we produced transgenic Arabidopsis expressing sense or antisense copies of each of the GA 20-oxidase cDNAs. Over-expression of any of the cDNAs gave rise to seedlings with elongated hypocotyls; the plants flowered earlier than controls in both long and short days and were 25% taller at maturity. GA analysis of the vegetative rosettes showed a two- to threefold increase in the level of GA4, indicating that GA 20-oxidase normally limits bioactive GA levels. Plants expressing antisense copies of AtGA20ox1 had short hypocotyls and reduced rates of stem elongation. This was reflected in reduced levels of GA4 in both rosettes and shoot tips. In short days, flowering was delayed and the reduction in the rate of stem elongation was greater. Antisense expression of AtGA20ox2 had no apparent effects in long days, but stem growth in one transgenic line grown in short days was reduced by 20%. Expression of antisense copies of AtGA20ox3 had no visible effect, except for one transgenic line that had short hypocotyls. These results demonstrate that GA levels and, hence, plant growth and development can be modified by manipulation of GA 20-oxidase expression in transgenic plants.