唐菖蒲赤霉素受体基因克隆及表达分析

以唐菖蒲子球为材料,采用RT PCR技术,克隆了1个赤霉素（GA）受体基因,命名为GhGID1a（GenBank登录号为KU525107）。其开放阅读框为1 032 bp,编码343个氨基酸。序列比对结果表明,GhGID1a推导氨基酸序列与百子莲、油棕和海枣等植物的GID1的氨基酸序列相似性较高,分别为82%、78%和77%。50 mg/L GA₃对子球萌发具有促进作用,150 mg/L GA₃会抑制其萌发。实时荧光定量PCR结果显示,GA3处理对GhGID1a基因表达具有反馈抑制作用,GhGID1a表达量随着子球休眠解除而逐渐降低,推测唐菖蒲子球休眠解除可能与GA及其受体基因有关,GA及其受体基因GhGID1a可能参与了调控唐菖蒲的子球休眠与萌发过程。     

海岛棉GbHCT13基因的功能验证

该研究利用海岛棉‘新海21’和陆地棉ND203以及模式植物拟南芥,通过转基因及荧光定量检测等方法探究海岛棉GbHCT13基因(GenBank 登录号MW048849)在纤维发育中的功能。结果显示：(1)成功构建重组载体pCAMBIA3301 GbHCT13,经农杆菌介导法转化、除草剂抗性基因筛选、荧光定量检测方法鉴定获得转GbHCT13基因拟南芥T₃代植株4株;qRT PCR检测表明,转基因植株中GbHCT13基因表达量较野生型极显著增加。(2)转基因拟南芥过表达GbHCT13基因使植株同一时期的生长较野生型旺盛,株形、叶片数、抽薹数和茎秆表皮毛数量均与野生型存在差异;组织化学分析发现,转GbHCT13基因的拟南芥较野生型茎秆初生木质部生长活跃,导管增粗,次生木质部导管细胞壁横截面积变大,但髓质细胞无明显变化;过表达GbHCT13使拟南芥中木质素合成途径基因发生不同程度改变,其中CAD、CCoAOMT、PAL和4CL与GbHCT13基因的表达呈正相关。(3)经大田筛选、分子鉴定,成功获得转GbHCT13基因棉花植株3株;转GbHCT13基因棉花的棉纤维伸长率增加,纤维强度增大;沉默GbHCT13基因使棉花植株木质素含量降低,茎秆表皮毛数量减少,木质部导管细胞数量减少,导管细胞壁中木质素沉积量降低,而棉株并未发生株高上的明显矮化现象,且木质素合成通路中的CAD、CCoAOMT、CCR、PAL 4个基因的表达均呈降低趋势,说明抑制GbHCT13使得棉花生长代谢受阻,影响纤维发育起始。研究表明,GbHCT13基因能影响棉花植株中木质素合成从而调控纤维的生长发育,其功能与GbHCT13基因在模式植物拟南芥中的基本一致。     

滇水金凤花距发育相关基因ABP的克隆及表达分析

为探究滇水金凤(Impatiens uliginosa)ABP基因的结构和表达特征,该研究以滇水金凤为材料,采用RT-PCR 技术对滇水金凤ABP基因进行克隆,运用DNAMAN和MEGA对其所编码的蛋白序列进行同源性分析和系统进化分析,并利用qRT-PCR分析ABP基因的时空表达模式。结果表明:(1)滇水金凤ABP基因的cDNA 全长为627 bp,编码208 aa,命名为IuABP基因,其蛋白具有Cupin超家族蛋白的典型结构。(2)同源性分析表明滇水金凤ABP基因的氨基酸序列与喜马拉雅凤仙花(I. glandulifera)、月季(Rose chinensis)、木薯(Manihot esculenta)等物种的同源性均达71%; 系统进化分析表明IuABP与喜马拉雅凤仙花(Impatiens glandulifera)聚为一支,亲缘关系最近。(3)qRT-PCR分析表明IuABP基因在滇水金凤花距发育的3个时期及2个部位均有表达。随着花距的发育,IuABP基因在滇水金凤花距檐部的表达量呈先下降后上升的趋势,在盛花期时达最高,而在花距距部的表达量逐渐下降。以上结果为进一步研究滇水金凤ABP基因在花距发育中的功能及其表达调控机制提供了一定的理论参考。     

甜荞花青素合成相关基因FeR2R3-MYB的克隆与表达分析

MYB 是一类常见的转录因子,广泛参与植物花青素生物合成的调控。为探究 MYB转录因子在甜荞花青素生物合成中的调控作用,该研究从红花甜荞和白花甜荞转录组学数据中筛选并克隆出一个和花青素生物合成相关的MYB基因,将其命名为 FeR2R3-MYB,GenBank 登录号为 MT151381.1,并对该序列进行生物信息学分析,以及利用 qRT-PCR 分析FeR2R3-MYB基因在白花甜荞和红花甜荞中的表达特征。结果表明:(1)FeR2R3-MYB基因全长 831 bp,编码 276 个氨基酸,蛋白的相对分子质量为 30.95 kD,理论等电点(pI)为 8.73,蛋白的不稳定指数为 69.64,属于不稳定蛋白,总疏水值为-0.679,整条肽链呈现亲水特性。(2)FeR2R3-MYB 具有典型的 R2R3-MYB 结构域,属于 R2R3-MYB 亚家族。(3)FeR2R3-MYB 与同属蓼科的苦荞和虎杖亲缘关系比较近。(4)FeR2R3-MYB 的启动子序列共含有 9 个光照响应元件、17 个转录因子结合位点、4 个非生物响应元件和 2 个激素响应元件。(5)亚细胞定位发现 FeR2R3-MYB 只在细胞核中表达。(6)FeR2R3-MYB 基因的表达量在叶片和花序中红花甜荞均高于白花甜荞,推测 FeR2R3-MYB 基因可以正向调节甜荞花青素生物合成。综上所述,该研究结果为进一步深化 FeR2R3-MYB 基因在甜荞花青素生物合成途径中的功能及表达调控方面的研究提供了基础。     

猪SOCS-2基因的克隆及序列分析

从中国地方猪品种八眉猪（BaMei）肾脏组织中提取总RNA,采用RT-PCR方法克隆了猪SOCS-2(suppressor of cytokine signaling -2,细胞因子信号转导抑制因子-2)基因的cDNA序列,经T/A克隆,插入到pMD19-T载体上,导入大肠杆菌DH-5α,阳性克隆经PCR鉴定后进行测序,将测序结果与GenBank中已登录的人、大鼠和小鼠SOCS-2基因的序列进行同源性比较,利用生物信息学和分子生物学软件对猪SOCS-2基因编码的蛋白进行结构预测。结果表明：首次成功克隆了猪SOCS-2基因的cDNA序列（GenBank登录号为EF121242）,其长度为822 bp,该基因ORF区核苷酸序列与其他物种相比同源性达到93%以上,氨基酸同源性则达到89%以上,生物信息学分析表明该蛋白分子量为22.25kD,等电点pI=8.30,包含199个氨基酸残基。该基因cDNA序列的克隆,有利于进一步研究SOCS-2调节机体发育的分子机理。     

猪SOCS-2基因的克隆及序列分析

从中国地方猪品种八眉猪（BaMei）肾脏组织中提取总RNA,采用RT-PCR方法克隆了猪SOCS-2(suppressor of cytokine signaling -2,细胞因子信号转导抑制因子-2)基因的cDNA序列,经T/A克隆,插入到pMD19-T载体上,导入大肠杆菌DH-5α,阳性克隆经PCR鉴定后进行测序,将测序结果与GenBank中已登录的人、大鼠和小鼠SOCS-2基因的序列进行同源性比较,利用生物信息学和分子生物学软件对猪SOCS-2基因编码的蛋白进行结构预测。结果表明：首次成功克隆了猪SOCS-2基因的cDNA序列（GenBank登录号为EF121242）,其长度为822 bp,该基因ORF区核苷酸序列与其他物种相比同源性达到93%以上,氨基酸同源性则达到89%以上,生物信息学分析表明该蛋白分子量为22.25kD,等电点pI=8.30,包含199个氨基酸残基。该基因cDNA序列的克隆,有利于进一步研究SOCS-2调节机体发育的分子机理。     

油茶DELLA基因的克隆和表达分析

为了解油茶(Camellia oleifera)中DELLA基因功能及其表达特性,采用PCR技术从‘长林4号’油茶中克隆了5个DELLA基因,命名为CoDELLA1~CoDELLA5,对其编码的5个CoDELLA蛋白进行生物信息学分析,并对5个DELLA基因的表达模式以及激素响应活性进行了分析。结果表明,5个CoDELLA基因的编码区长度分别为1 791、1 875、1 848、1 593和1 581 bp,分别编码597、625、616、531和527个氨基酸。5个CoDELLA蛋白的氨基酸序列相似度较高,丝氨酸残基为主要的潜在磷酸化位点,CoDELLA蛋白N端均含有典型的DELLA结构域。不同物种中DELLA蛋白的系统发育存在差异,CoDELLA与茶树的CsDELLA同源性最高。CoDELLA基因在油茶不同组织中的表达也存在差异,且赤霉素和独脚金内酯等多种激素和非生物胁迫对其表达具有调控作用。CoDELLA基因可能在油茶的生长发育和非生物胁迫响应中发挥重要作用。     

三倍体枇杷花期调控基因EjAGL6的表达特性和功能分析

以三倍体枇杷(Eriobotrya japonica) ‘华玉无核1号’的花芽为材料,采用基因克隆技术获得EjAGL6基因,分析其序列、亚细胞定位特性以及在二倍体和三倍体枇杷早晚花品种中的表达水平。采用花序浸染转化拟南芥,并利用实时荧光定量PCR分析转基因拟南芥植株的EjAGL6基因表达量,进一步观察野生型与EjAGL6转基因拟南芥的表型差异,分析EjAGL6基因的功能,为解析EjAGL6基因参与三倍体枇杷花期调控机制提供理论依据。结果显示：(1)成功获得MADS box基因EjAGL6;该基因的编码区序列(CDS)为732 bp,编码243个氨基酸,分子质量为27.88 kD,等电点为 9.05,脂溶指数为 79.05;系统进化树分析表明,枇杷EjAGL6与苹果MdAGL6蛋白质的相似性较高,聚在同一分支。(2)蛋白序列比对发现,EjAGL6的M区有57个氨基酸,I区有30个氨基酸,K区有82个氨基酸,C区有74个氨基酸,其中C区包含高度保守的AGL6基序Ⅰ和AGL6基序Ⅱ。(3)亚细胞定位分析表明,EjAGL6蛋白定位在细胞核,具有典型的MADS box转录因子亚细胞定位特性。(4)实时荧光定量PCR分析表明,EjAGL6基因在二倍体和三倍体枇杷早、晚花品种中均有表达,主要集中于小花分化期(S6)、花蕾露白期(S7)和盛花期(S8),且EjAGL6基因在二倍体和三倍体早花品种中的花蕾露白期的表达量均较高。(5) 转基因拟南芥株系的EjAGL6基因表达量显著高于野生型拟南芥;转EjAGL6基因植株表型观察显示,EjAGL6基因在拟南芥中过量表达能够使转EjAGL6基因拟南芥的开花时间提前1周左右。研究认为,EjAGL6基因可促使枇杷开花时间提前,推测EjAGL6基因在花蕾露白期发挥调控花期的关键作用。     

地黄C3H基因的克隆及生物信息学分析

香豆酸-3-羟化酶属于植物中最大的蛋白酶细胞色素P450家族之一,在植物生命活动中发挥着重要作用。为了解地黄香豆酸-3-羟化酶基因RgC3H合成毛蕊花糖苷的功能,该研究基于地黄代谢组学分析获得KEGG途径中的C3H,采用多重比对在NCBI中获得同源基因的一个保守序列,并基于该保守序列和地黄SRA数据库,采用电子克隆和RT-PCR克隆技术获得地黄C3H基因全长CDS(RgC3H),对其进行生物信息学分析。结果表明:RgC3H基因全长为1 530 bp,且编码一个含509个氨基酸、分子量为57.91 kD、无信号肽的蛋白质; 基于氨基酸序列的结构分析显示,RgC3H有一个保守区域-P450结构域; 系统进化分析结果显示,RgC3H与芝麻和猴面花的C3H基因具有很高的同源性。上述结果为进一步研究RgC3H基因在地黄毛蕊花糖苷生物合成途径中的作用奠定了基础。     

彩斑突变体菊花CmF3′Ha基因及启动子的克隆与分析

该研究以菊花彩瓣突变体CQ17 mu为材料,利用RT PCR克隆获得了类黄酮 3′ 羟化酶基因(F3′H),该基因开放阅读框全长1 527 bp,编码508个氨基酸,与已知的菊花CmF3′H相似性达到99%,故将其命名为CmF3′Ha。氨基酸序列分析表明,CmF3′Ha编码的蛋白具有保守的F3′H结构域,属于P450超家族。多重序列比对和系统发育分析表明,CmF3′Ha与其他菊花品种的F3′H亲缘关系最近。实时定量PCR分析显示,CQ17 mu花瓣中CmF3′Ha基因的表达水平显著高于CQ17的粉色花瓣,表明CmF3′Ha参与了CQ17 mu花瓣紫色条斑部位的花色素代谢。进一步利用染色体步移法克隆得到了CQ17 mu的F3′H基因上游1 086 bp的启动子区序列,经分析显示,该序列中除包含TATA box等核心启动子元件外,还包括多个MYB、MYC结合位点及多个光响应元件和激素应答元件。研究结果证明了菊花CmF3′Ha基因与植物花青素积累呈正相关,参与彩色条斑部分的花青素合成过程,为菊花的分子育种提供了理论依据。     

Expression of novel rice gibberellin 2-oxidase gene is under homeostatic regulation by biologically active gibberellins

We have cloned two genes for gibberellin (GA) 2-oxidase from rice (Oryza sativa L.). Expression of OsGA2ox2 was not observed. The other gene, OsGA2ox3, was expressed in every tissue examined and was enhanced by the application of biologically active GA. Recombinant OsGA2ox3 protein catalyzed the metabolism of GA₁ to GA₈ and GA₂₀ to GA₂₉-catabolite. These results indicate that OsGA2ox3 is involved in the homeostatic regulation of the endogenous level of biologically active GA in rice. Electronic Publication     

Function and Expression Analysis of Gibberellin Oxidases in Apple

Three cDNAs, encoding gibberellin (GA) 20-oxidase (MdGA20ox1, identical to AB037114), 3-oxidase (MdGA3ox1), and 2-oxidase (MdGA2ox1), were isolated from apple cv. Fuji (Malus x domestica). Southern blot analysis indicated that each of these genes belongs to a gene family. Standard enzyme assays show that the MdGA20ox1-MBP fusion protein can sequentially oxidize three times at C-20 position of GA₁₂ and GA₅₃ and generate GA₉ and GA₂₀; the MdGA3ox1-MBP fusion protein converts GA₂₀ and GA₉ to GA₄ and GA₁, and the MdGA2ox1-MBP fusion protein converts GA₄ and GA₁ to GA₃₄ and GA₈, respectively. In addition, we confirmed that MdGA20ox1 is strongly expressed in immature seeds and scarcely detected in other tissues, whereas MdGA3ox1 and MdGA2ox1 are mainly expressed in flowers. Therefore, all the three cDNAs are localized in reproductive tissues. Functional and expression analysis of the three GA oxidases would provide fundamental molecular information to analyze GA metabolic regulation in apple.     

Characterization of <Emphasis Type="Italic">GA20ox</Emphasis> genes in tall and dwarf types coconut (<Emphasis Type="Italic">Cocos nucifera</Emphasis> L.)

Coconuts (Cocos nucifera L.) are divided by the height into tall and dwarf types. In many plants the short phenotype was emerged by mutation of the GA20ox gene encoding the enzyme involved in gibberellin (GA) biosynthesis. Two CnGA20ox genes, CnGA20ox1 and CnGA20ox2, were cloned from tall and dwarf types coconut. The sequences, gene structures and expressions were compared. The structure of each gene comprised three exons and two introns. The CnGA20ox1 and CnGA20ox2 genes consisted of the coding region of 1110 and 1131 bp, encoding proteins of 369 and 376 amino acids, respectively. Their amino acid sequences are highly homologous to GA20ox1 and GA20ox2 genes of Elaeis guineensis, but only 57% homologous to each other. However, the characteristic amino acids two histidines and one aspartic acid which are the two iron (Fe²⁺) binding residues, and arginine and serine which are the substrate binding residues of the dioxygenase enzyme in the 20G-FeII_Oxy domain involved in GA biosynthesis, were found in the active site of both enzymes. The evolutionary relationship of their proteins revealed three clusters in vascular plants, with two subgroups in dicots and three subgroups in monocots. This result confirmed that CnGA20ox was present as multi-copy genes, and at least two groups CnGA20ox1 and CnGA20ox2 were found in coconut. The nucleotide sequences of CnGA20ox1 gene in both coconut types were identical but its expression was about three folds higher in the leaves of tall coconut than in those of dwarf type which was in good agreement with their height. In contrast, the nucleotide sequences of CnGA20ox2 gene in the two coconut types were different, but the expression of CnGA20ox2 gene could not be detected in either coconut type. The promoter region of CnGA20ox1 gene was cloned, and the core promoter sequences and various cis-elements were found. The CnGA20ox1 gene should be responsible for the height in coconut, which is different from other plants because no mutation was present in CnGA20ox1 gene of dwarf type coconut.     

New semi-dwarfing alleles with increased coleoptile length by gene editing of gibberellin 3-oxidase 1 using CRISPR-Cas9 in barley (Hordeum vulgare L.)

The green revolution was based on genetic modification of the gibberellin (GA) hormone system with “dwarfing” gene mutations that reduces GA signals, conferring shorter stature, thus enabling plant adaptation to modern farming conditions. Strong GA-related mutants with shorter stature often have reduced coleoptile length, discounting yield gain due to their unsatisfactory seedling emergence under drought conditions. Here we present gibberellin (GA) 3-oxidase1 (GA3ox1) as an alternative semi-dwarfing gene in barley that combines an optimal reduction in plant height without restricting coleoptile and seedling growth. Using large-scale field trials with an extensive collection of barley accessions, we showed that a natural GA3ox1 haplotype moderately reduced plant height by 5–10 cm. We used CRISPR/Cas9 technology, generated several novel GA3ox1 mutants and validated the function of GA3ox1. We showed that altered GA3ox1 activities changed the level of active GA isoforms and consequently increased coleoptile length by an average of 8.2 mm, which could provide essential adaptation to maintain yield under climate change. We revealed that CRISPR/Cas9-induced GA3ox1 mutations increased seed dormancy to an ideal level that could benefit the malting industry. We conclude that selecting HvGA3ox1 alleles offers a new opportunity for developing barley varieties with optimal stature, longer coleoptile and additional agronomic traits.     

Physiological genetics of the dominant gibberellin-nonresponsive maize dwarfs,Dwarf8 and Dwarf9

Maize (Zea mays L.) Dwarf8-1 (D8-1) is an andromonoecious dwarf mutant proposed to be involved in gibberellin (GA) reception (Fujioka et al. 1988b; Harberd and Freeling 1989). The mutant D8-1 is dominant and GA-nonresponsive (Phinney 1956). We show by map position and similarity of phenotype that five additional dwarf mutants are D8 alleles. We show by map position and similarity of phenotype that a second andromonoecious dwarf mutant, D9-1, defines a duplicate gene. Maize D9-1 and each dominant D8 allele specify a different plant stature, from very mild to very severe dwarfism. Plants of D9-1 and all dominant D8 alleles, except D8-1591, were GA-nonresponsive when treated with 7500 nmol GA₃. The behavior of the mild dwarf D8-1591 was unique in that a small but significant growth response was detected (37% for D8-1591 vs. 130% for the wild type) when treated with 7500 nmol GA₃. These results establish that all dwarf genotypes, except D8-1591, in one dose set a maximum limit on plant growth and block the normal response to GA. When treated with the GA-synthesis inhibitor paclobutrazol, plants of all dwarf genotypes and wild-type siblings were severely dwarfed. Plants of all dwarf genotypes treated with the GA-synthesis inhibitor paclobutrazol and GA₃ were returned to their normal dwarf phenotype. Dominant dwarfing, delayed flowering, increased tillering, and anther development in the ear are characteristic features of D9-1 and all D8 alleles. The GA-synthesis-deficient dwarfs also have these characteristic features. We discuss the function of the wild-type gene product in the context of the observed results.Abbreviations D8 Dwarf8 - D9 Dwarf9 - GA_(n) gibberellin A_(n) - GA₃ gibberellic acid - MNL Maize Genetics Cooperation Newsletter - NIL near-isogenic lines - RFLP restriction fragment length polymorphism - WT wild type This work was supported, in part, by a National Science Foundation Plant Postdoctoral Fellowship to R.G.W., by grants from NIH and ICI Seeds to M.F., the NSF Center for Plant Developmental Biology and the California Agriculture Experiment Station. Much of the work was done in the laboratory of Tim Helentjaris and was supported by a grant from Pioneer Hi-Bred Int'l. The generous gifts of the dominant dwarfing mutants from M.G. Neuffer and O.E. Nelson Jr. are gratefully acknowledged.     

2,4‐D‐induced parthenocarpy in pear is mediated by enhancement of GA4 biosynthesis

Parthenocarpy, the productions of seedless fruit without pollination or fertilization, is a potentially desirable trait in many commercially grown fruits, especially in pear, which is self‐incompatible. Phytohormones play important roles in fruit set, a process crucial for parthenocarpy. In this study, 2,4‐dichlorophenoxyacetic acid (2,4‐D), an artificially synthesized plant growth regulator with functions similar to auxin, was found to induce parthenocarpy in pear. Histological observations revealed that 2,4‐D promoted cell division and expansion, which increased cortex thickness, but the effect was weakened by paclobutrazol (PAC), a gibberellin (GA) biosynthesis inhibitor. Phenotypic differences in pear may therefore be due to different GA contents. Hormone testing indicated that 2,4‐D mainly induced the production of bioactive GA₄, rather than GA_3. Three key oxidase genes function in the GA biosynthetic pathway: GA20ox, GA3ox and GA2ox. In a pear group treated with only 2,4‐D, PbGA20ox2‐like and PbGA3ox‐1 were significantly upregulated. When treated with 2,4‐D supplemented with PAC, however, expression levels of these genes were significantly downregulated. Additionally, PbGA2ox1‐like and PbGA2ox2‐like expression levels were significantly downregulated in pear treated with either 2,4‐D only or 2,4‐D supplemented with PAC. We thus hypothesize that 2,4‐D can induce parthenocarpy by enhancing GA₄ biosynthesis.     

Developmental and Hormonal Regulation of Gibberellin Biosynthesis and Catabolism in Pea Fruit

In pea (Pisum sativum), normal fruit growth requires the presence of the seeds. The coordination of growth between the seed and ovary tissues involves phytohormones; however, the specific mechanisms remain speculative. This study further explores the roles of the gibberellin (GA) biosynthesis and catabolism genes during pollination and fruit development and in seed and auxin regulation of pericarp growth. Pollination and fertilization events not only increase pericarp PsGA3ox1 message levels (codes for GA 3-oxidase that converts GA₂₀ to bioactive GA₁) but also reduce pericarp PsGA2ox1 mRNA levels (codes for GA 2-oxidase that mainly catabolizes GA₂₀ to GA₂₉), suggesting a concerted regulation to increase levels of bioactive GA₁ following these events. 4-Chloroindole-3-acetic acid (4-Cl-IAA) was found to mimic the seeds in the stimulation of PsGA3ox1 and the repression of PsGA2ox1 mRNA levels as well as the stimulation of PsGA2ox2 mRNA levels (codes for GA 2-oxidase that mainly catabolizes GA₁ to GA₈) in pericarp at 2 to 3 d after anthesis, while the other endogenous pea auxin, IAA, did not. This GA gene expression profile suggests that both seeds and 4-Cl-IAA can stimulate the production, as well as modulate the half-life, of bioactive GA₁, leading to initial fruit set and subsequent growth and development of the ovary. Consistent with these gene expression profiles, deseeded pericarps converted [¹⁴C]GA₁₂ to [¹⁴C]GA₁ only if treated with 4-Cl-IAA. These data further support the hypothesis that 4-Cl-IAA produced in the seeds is transported to the pericarp, where it differentially regulates the expression of pericarp GA biosynthesis and catabolism genes to modulate the level of bioactive GA₁ required for initial fruit set and growth.     

Molecular Cloning, Characterization, and Expression Analysis of Genes Encoding Gibberellin 20-Oxidase in Dasypyrum villosum Dwarf Mutant

Thirty-six gene sequences encoding the gibberellin (GA) 20-oxidase were obtained from Dasypyrum villosum and its dwarf mutant. Sequence alignment showed that there were 21 SNPs and 4 InDels among these sequences which could be divided into three haplotypes??haplotype I, II, and III with 1,293, 1,297, and 1,294?bp in length, respectively. They contained a CDS with 1,080?bp in length encoding a putative polypeptide of 359 amino acids. Two haplotypes were found in wild type (I and II) and dwarf mutant (II and III), respectively. Q-PCR analysis showed that in the whole growing stages, the majority expression levels of haplotypes from wild types were higher than that of dwarf mutant, suggesting that wild types could synthesize more active GA substrates than dwarf mutant. The expression level in stem nodes and internodes between wild type and dwarf mutant were not significantly different, whereas their expression levels in roots were distinctly distinguished from each other in seedling, stem elongation, and heading stages, implying that most active GAs were synthesized in the root, and some were consumed by the root itself, and the others might be transported to other organs.