Modification of plant height via RNAi suppression of OsGA20ox2 gene in rice

GA 20-oxidase (GA20ox) is a regulatory enzyme for the syntheses of biologically active GAs in plants. The loss-of-function mutations in OsGA20ox2 of rice (Oryza sativa L.) generate the well-known Green Revolution gene sd-1, which cause the semi-dwarfism phenotype. In our present investigation, semi-dwarf plants were generated from a taller rice variety QX1 by RNAi suppression on the expression of OsGA20ox2. The 531bp-fragment of OsGA20ox2 was amplified by PCR from genomic DNA of QX1 and used to construct the hairpin RNAi vector pCQK2. The wild type QX1 was transformed with pCQK2 by Agrobacterium-mediated transformation and some independent transgenic RNAi lines exhibited semi-dwarfism. RT-PCR and Northern blot analyses showed that the expression of OsGA20ox2 was specifically suppressed in the RNAi semi-dwarf lines. Endogenous GA assays revealed that the contents of the GA20ox2-catalyzed products GA₁₉, GA₂₀ and the down-stream biologically active GA₁ were drastically reduced in the RNAi semi-dwarf lines. We further showed that the RNAi semi-dwarf lines could be restored to normal plant height by applying exogenous GA₃. The results indicated that the semi-dwarfism of the RNAi semi-dwarf lines was associated with the decreased expression of OsGA20ox2 gene and the reduced content of endogenous biologically active GA₁. Analyses of panicle length, seeds per panicle and 1000-grain weight suggested that the RNAi semi-dwarf lines showed stable grain yield compared with the wild type plants. It is demonstrated that the RNAi approach could be useful for plant breeding purposes in the future. Qing Yang, Chun-Lian Wang have contributed equally to this work.     

Quantification of endogenous gibberellins in two flax (Linum usitatissimum L.) cultivars during seed development

Eight gibberellins (GAs) were identified and quantified in extracts of seed of the flax, Linum usitatissimum L. cv. Hwanamama, Hwabaekama by GC-MS-SIM and Kovats retention indices. Four of these gibberellins are members of the early-C13-hydroxylation pathway (GA₅₃, GA₁₉, GA₂₀, and GA₁), and four are members of the non-C13-hydroxylation pathway (GA₁₂, GA₂₄, GA₉, and GA₄). Of these eight gibberellins, GA₂₄ was more abundant in yellow-seeded flax (cv. Hwanamama) than in brown-seeded flax (cv. Hwabaekama), whereas GA₉ was 8.3-fold more abundant in brown-seeded flax (cv. Hwabaekama) than in yellow-seeded flax (cv. Hwanamama). In both flax cultivars, non-C13-hydroxylation pathway was the main route leading to endogenous accumulation of GA₄ in the seed.     

Effect of Prohexadione Calcium on Growth Components and Endogenous Gibberellins Contents of Rice (Oryza sativa L.)

Prohexadione calcium (Pro‐Ca) is a novel plant growth regulator that inhibits the late stages of gibberellins (GAs) biosynthesis in plants, thus reducing the vegetative growth of the plant. Pro‐Ca was applied to field‐grown rice cultivar Ilpumbyeo to evaluate its effect on growth and yield components and endogenous GAs, when applied at different rates and timings. This study illustrated that Pro‐Ca application decreased the internode length, stem length, panicle length, shoots fresh weight and stem lodging index, while stem breaking force was increased in all treatments. The effect of Pro‐Ca was more pronounced in 20 ppm concentration than 10 ppm. The number of panicles m⁻², ripened grain ratio and 1000 grain weight and milled rice yield parameters, increased with the application of Pro‐Ca, though the number of grains per panicle decreased insignificantly. The endogenous bioactive GA₁ contents significantly decreased with the application of Pro‐Ca, as the GA biosynthesis was interrupted at later stages, when GA₂₀ conversion to GA₁ was blocked, thus drastically reducing GA₁ and GA₈ levels, but only insignificant changes were observed for intermediates from GA₁₂ to GA₂₀. The levels of the endogenous GAs in rice shoots were measured by GC‐MS‐SIM using ²H₂‐labelled GAs as internal standards. Present results also illustrated that early application of Pro‐Ca was more effective than its late application for effective control of different growth parameters and thus reducing lodging in rice plants.     

不同脱水剂对杂交水稻制种种子质量及基因表达的影响

种子脱水干燥是杂交水稻制种过程的一个重要环节,种子自然脱水速率慢,遇雨天还易引起成熟期推后和穗发芽等问题。利用脱水剂快速降低种子水分对种子安全生产有重要意义。本研究以Y两优689杂交水稻品种为材料,比较不同脱水剂对其制种种子脱水及种子质量的影响。11种脱水剂均能加快种子成熟后期脱水,7号和9号脱水剂喷施后5 d,种子水分分别下降4.6%和3.6%,且对种子千粒重、发芽和幼苗生长无不良影响,种子可溶性蛋白、可溶性糖、ABA和GA3含量则显著高于对照。种子室温贮藏6个月后,与对照相比,7号、8号和9号脱水剂处理对种子发芽和幼苗生长无显著影响,7号和9号脱水剂处理的种子可溶性蛋白含量、α-淀粉酶活性、ABA含量与对照无显著差异。7号比9号的脱水效果好,可作为Y两优689的脱水剂。种子贮藏前,7号、8号和9号脱水剂处理能提高种子中OsNECD1和OsNCED2基因的表达,降低OsGA2ox1的表达;而种子贮藏6个月后,OsNECD1和OsNCED2表达量下降,OsCYP707A5和OsGA2ox1表达量上升。这表明经过6个月贮藏,脱水剂处理对种子ABA合成相关基因和GA3分解相关基因表达的影响减弱,同时OsCYP707A5的高表达导致种子ABA含量减少,可能是种子贮藏后萌发力提高的主要原因。     

沙生针茅种子吸水特性及其萌发特征研究

以民勤沙生针茅种子为试验材料,研究不同温度蒸馏水处理、GA₃处理、PEG处理对沙生针茅种子萌发的影响。研究结果表明：在不同水温处理下的沙生针茅种子发芽率随着水温的升高其吸胀率呈递增趋势,且不同水温浸泡下沙生针茅种子的吸水率差异极显著;GA₃处理对沙生针茅种子的萌发具有促进作用,在80 mg/L的GA₃溶液处理下的发芽率、发芽势最高,分别为54%、48%,第7天达萌发高峰期,有效得缩短了萌发周期;随着PEG溶液的增加,沙生针茅种子的萌发得到不同程度的诱发,PEG溶液浓度越低,诱发作用越明显,PEG溶液浓度在5%时,种子发芽率、发芽势较高,说明干旱胁迫对沙生针茅种子的萌发具有不同程度的诱发作用,沙生针茅种子能适应一定程度的干旱土壤生境。     

外源ABA抑制水稻种子发芽的生理机制

比较研究了外源植物激素ABA（脱落酸）与植物生长抑制剂MH（马来酰肼）对水稻种子发芽的抑制作用,以期阐明ABA抑制水稻种子发芽的生理学作用与机制。研究结果表明ABA与MH虽然都具有抑制水稻种子发芽的作用,但两者生理作用性质不同,ABA抑制水稻种子发芽是延缓种子发芽,而MH是降低种子活性,对种子发芽具遏制、伤害作用。MH     

莲雾催花过程中叶片内源激素的变化

为探明莲雾产期调节的内在生理生化机理,以‘粉红种’莲雾为材料,研究莲雾反季节遮阳催花过程中叶片内源激素IAA、GA₃、ABA、ZT含量的动态变化和激素平衡（比值）的变化。结果表明：在遮阳处理后,叶片的IAA、ZT、GA₃、ABA含量均降低;喷施催花药剂后,IAA、ZT、ABA含量上升,GA₃含量下降。遮阳抑制了莲雾的营养生长。在花芽分化期,高水平的ABA、ZT,低水平的GA₃,IAA/ABA、GA₃/ABA、(IAA GA₃)/ABA低,ZT/GA₃高有利于莲雾成花。其中,高水平的ABA、GA₃/ABA低、ZT/GA₃高可能在莲雾的花芽分化中起主导作用。     

Exogenous Abscisic Acid Application During Grain Filling in Winter Wheat Improves Cold Tolerance of Offspring's Seedlings

Low temperature seriously depresses seed germination and seedling growth in winter wheat (Triticum aestivum L.). In this study, wheat plants were sprayed with abscisic acid (ABA) and fluridone (inhibitor of ABA biosynthesis) at 19 days after anthesis (DAA) and repeated at 26 DAA. The seeds of those plants were harvested, and seed germination and offspring's seedling growth under low temperature were evaluated. The results showed that exogenous ABA application decreased seed weight and slightly reduced seed set and seed number per spike. Under low temperature, seeds from ABA‐treated plants showed reduced germination rate, germination index, growth of radicle and coleoptile, amylase activity and depressed starch degradation as compared with seeds from non‐ABA‐treated plants; however, activities of the antioxidant enzymes in both germinating seeds and seedling were enhanced from those exposed to exogenous ABA, resulting in much lowered malondialdehyde (MDA) and H₂O₂ concentrations and production rate. In addition, the maximum quantum efficiency of photosystem II was also enhanced in ABA‐treated offspring's seedlings. It is concluded that exogenous ABA treatment at later grain‐filling stage could be an effective approach to improve cold tolerance of the offspring during seed germinating and seedlings establishment in winter wheat.     

不同小麦品种(系)的存放种子活力比较研究

为发掘利用高活力小麦种质材料,揭示不同小麦品种间(系)的种子活力差异及其内在生理原因,本研究以置于常温种子库老化6年的180个小麦品种(系)的种子为材料进行了种子活力比较研究。结果表明:不同小麦品种(系)间发芽率存在显著差异,其中35个小麦品种(系)的种子完全失活、不能发芽,种子活力最高的3个品种分别为豫麦18(92%),西农3517(76%)和浏虎98(72%);测定了种子浸出液电导率、种子含水量和籽粒品质性状并与发芽率进行了相关分析,发现发芽率与电导率和籽粒淀粉含量呈极显著负相关,与含水量呈显著负相关,但与籽粒总蛋白质含量相关不显著;进一步对种子活力高低差异显著的6个品种(系)进行了种子萌发关键激素赤霉素(GA3)和脱落酸(ABA)的测定,发现3个高活力小麦品种(系)(豫麦18、西农3517、浏虎98)种子中GA3/ABA的比值均显著高于3个低活力品种(系)(西农2003、洛麦21和陕麦107)。本研究表明,种子活力性状在小麦种质资源中存在广泛变异,植物内源激素GA3和ABA的相对含量是影响种子活力高低的关键因素。     

滇重楼种子休眠机理研究

探讨滇重楼种子休眠机理,为打破种子休眠促进滇重楼实生育苗提供理论基础。采用重量法、胚率测定、ELISA等方法对种子吸水特性、萌发抑制物及种胚休眠与发育的内源机理进行了研究。结果表明,滇重楼种子休眠属于形态学-生理学休眠类型。滇重楼种胚发育不完全、种子中存在萌发抑制物质是种子休眠的主要原因。种子不存在吸水障碍,与休眠无关。内源激素ABA和GA3在滇重楼种子休眠与萌发过程中起着重要作用,ABA是引起休眠的关键因素,ABA含量的降低是滇重楼种胚发育的始动因子,GA3有助于种子的生理后熟。萌发抑制物的存在可能抑制了种胚发育,而与生理学后熟过程无关。在自然条件下,滇重楼生境中较长的低温时期可能是滇重楼种子具有较长时间休眠的外界原因。     

花生种子休眠解除过程中相关基因的表达分析

种子休眠性是花生重要的农艺性状,外源乙烯利能诱导花生种子休眠的解除,为了阐明乙烯利作用下花生种子休眠解除的分子机制,设置吸胀的休眠种子为对照,100 mg L–1乙烯利处理吸胀休眠种子后不同时间的样品(AE1、AE2、AE3)进行转录组分析,比较了花生种子休眠解除过程中ABA、GA、ETH、auxin相关基因的表达。结果表明,15个与GA、40个与ABA、60个与ETH、56个与auxin相关的unigenes在花生种子休眠解除过程中表现显著差异表达。荧光定量PCR结果显示,ABA合成关键基因Ah NCED2和代谢关键基因Ah CYP707A1在种子休眠解除过程中均受外源乙烯利诱导,表达差异显著;在休眠和无休眠种子吸胀萌发过程中,Ah NCED2和Ah CYP707A1的表达趋势不同,Ah NCED2对于种子休眠的维持发挥积极作用,而Ah CYP707A1对于种子休眠解除发挥积极作用。     

硝酸铈促进水稻种子萌发的生理效应

用2.5～30 mg/L的Ce(NO3)3处理水稻种子能加快萌发, 明显提高种子活力, 促进植株根系生长, 增加叶绿素含量。 这是由于在用Ce(NO3)3处理期间加快了种子的吸水速率, 增加了膜的通透性, 增强了呼吸作用, 增加了种子萌发期间IAA、 GAs和CTK等植物激素的含量, 而ABA含量变化不大。 其中用10 mg/L Ce(NO3)3处理效果最佳,     

杂交水稻制种过程中籽粒内源激素含量的动态变化

穗发芽对粮食生产造成巨大的经济损失,为研究穗发芽前杂交水稻制种中杂交种子籽粒内源激素含量变化关系。采用HPLC等方法,以2个杂交水稻组合‘Y两优1号’（亲本为Y58S× R9311×）和‘II优416’（ Ⅱ-32A×R416）为材料,在水稻制种过程中进行穗发芽实验。发现组合‘Y两优1’号种子比‘II优416’易于穗发芽。同时测定了两组合在授粉15 ~27 天期间种子内源激素（IAA﹑GA1﹑ZR和ABA）含量的变化,易穗发芽的‘Y两优1号’内源激素ZR﹑IAA和GA1含量高于‘II优416’,ABA含量低于‘II优416’,且‘Y两优1号’籽粒内源GA1/ABA比值在整个过程中均大于‘II优416’。由此可见,杂交水稻制种过程中出现穗发芽现象的最主要原因是穗发芽前期种子内GA1的含量高于ABA且在发生穗发芽时GA1/ABA比值比较大。     

转反义trxs基因小麦籽粒中ABA和GAs含量与穗发芽的关系

以转反义trxs基因两个纯合株系及其未转基因对照小麦品种‘豫麦18’为材料,采用荧光定量RT-PCR、整穗发芽试验和酶联免疫吸附（ELISA）的方法,对小麦灌浆过程中反义trxs基因表达、穗发芽特性及内源激素ABA和GAs含量进行了测定,探讨转基因小麦籽粒中内源激素ABA和GAs含量与穗发芽的关系。结果表明,在种子形成过程中,反义trxs基因能够正常表达,两个转基因株系中内源trxh基因表达量都明显低于对照;转基因株系籽粒中ABA和ABA/GAs含量显著高于对照,平均分别比对照升高了18.39%和23.47%,而GAs含量在花后15～30d较对照有所降低,平均比对照降低了9.14%;花后20、25、30d,转基因株系的穗粒发芽率显著或极显著低于对照,平均分别比对照下降了49.65%、28.2%、27.23%;相关分析表明,穗发芽率与ABA和ABA/GAs含量均呈显著或极显著负相关,与GAs含量呈正相关。由此推断,反义trxs基因导入抑制了小麦内源同源基因的表达,改变了小麦体内ABA和GAs的平衡,使ABA合成量增加,是导致转基因小麦穗发芽率降低的原因之一。     

人工老化过程中柔枝松种子内源激素含量变化的研究

本研究为柔枝松(Pinus flexilis James.)种子内源激素含量的变化实验。经过人工老化处理,5个柔枝松种源种子内源IAA、ZR、GA3和ABA含量发生明显的变化,在整个发芽过程中,各内源激素含量的变化均呈先增加后降低的趋势。经过老化处理内源激素IAA、ZR、GA3和ABA含量都增加。其中ZR的含量增加最多。通过实验判断引进的5个柔枝松种源Rl、Rg和Rsi种子的品质较好,种源Rs最差,种源Rsd变化较大。同时也为国内林木种子活力的研究提供借鉴。     

羊柴种子去荚、低温层积处理后萌发特性及HPLC法测定内源激素ABA、IAA含量变化

为了研究羊柴种子自身萌发力对其飞播过程的影响,笔者对羊柴种子去荚处理后含水量、吸水规律、萌发能力以及羊柴种子不同天数低温层积处理后萌发能力进行相关研究,同时对2种处理后内源激素ABA、IAA分别进行生物测定。结果表明：带荚种子含水量为9.16%,去荚种子为8.92%;两者吸水过程均分为急剧吸水期、稳定吸水期、饱和吸水期3个阶段,去荚种子的最大吸水速率为带荚种子的4.79倍,达到饱和期所用时间约为带荚种子的1/2;羊柴种子去荚处理后发芽率、发芽指数最高,分别为81.67%、33.27,为对照组的8.167、38.39倍,带荚种子低温层积处理60天后发芽率、发芽指数次之,为46.67%、4.19;CK组抑制物ABA含量最高,达7921.12 ng/mL,经过80天低温层积处理后ABA含量降至最低,为4590.85 ng/mL。综合去荚处理以及低温层积处理后的发芽率、发芽指数、ABA含量,IAA/ABA比值,最后确定羊柴种子的最适发芽条件为去荚处理以提高其吸水性能或对带荚种子进行60天,4℃的低温层积处理。     

赤霉素对盐胁迫抑制水稻种子萌发的缓解作用的蛋白质组分析

用粳稻日本晴(Oryza sativa L. cv.Nipponbare),研究了盐胁迫对水稻种子萌发的抑制作用和赤霉酸(GA₃)对盐胁迫的缓解作用;分别以H₂O (对照）,5 g L^-1 NaCl (处理I),5 g L^-1 NaCl + 100 μmol L^-1 GA₃(处理II)培养水稻种苗48 h,提取芽中的蛋白质,利用双向电泳(2-DE)和基质辅助激光解吸电离飞行时间质谱(MALDI-TOF MS)技术分析了水稻蛋白质组的变化。结果表明,在盐胁迫条件下,日本晴种子的萌发显著受到抑制,而GA₃能显著缓解这种抑制作用;用ImageMaster软件分析2-DE凝胶,发现有4个蛋白质斑点表现出显著的变化,在盐胁迫下斑点S1、S2和S3表达下调而斑点S4消失,在GA₃与盐共处理时,这4个蛋白质点的表达均有不同程度的恢复;经MALDI-TOF MS分析,其中2个蛋白质斑点(S1,S3)分别被鉴定为isoflavone reductase-like蛋白与葡萄糖磷酸变位酶,这些蛋白可能与GA₃提高水稻耐盐性途径相关。     

IAA、GA3和ABA对稻根负向光性和生长的影响

为探明稻根负向光性与内源植物激素含量的关系,以及外源激素对水稻根系生长的影响,以扬稻6号(籼稻)、日本晴(粳稻)和中花11 (粳稻)的OsPIN1a超表达转基因水稻为材料,观察了根负向光性生长的过程,分析了负向光弯曲部位内源生长素(IAA)、赤霉素(GA₃)和脱落酸(ABA)的含量,以及外源激素对水稻根系形态和解剖结构的影响。结果表明, 扬稻6号和日本晴的根都具有负向光性,扬稻6号侧根和不定根的负向光性大于日本晴;2种材料中,IAA、GA₃和ABA在发生负向光弯曲过程中的含量变化基本一致。光照引起3种激素的含量下降;向光侧的含量低于背光侧。单侧光照和外施IAA共同促进了籼稻根毛的大量发生;与普通水稻相比,OsPIN1a超表达稻株的根负向光性角度增大。外源GA₃和ABA处理与稻根负向光性间无直接关系;外源ABA处理显著抑制了根的生长,单株根系总吸收表面积与根数均减少。10 µmol L^-1 ABA处理的水稻根尖生长异常,分生区和根冠发育不良,细胞伸长受抑,成熟区细胞变形使根局部膨胀,并改变了中央维管组织的发育。     

不同药剂对一把伞南星种子萌发特性的影响

利用GA 3、高锰酸钾、双氧水、甲醛、硫酸、多菌灵浸泡一把伞南星种子,探讨不同浓度药剂处理对种子萌发的影响。结果表明,使用不同浓度的GA 3、硫酸、甲醛、过氧化氢、多菌灵、高锰酸钾处理一把伞南星种子,对其发芽特性具有不同程度的影响。200倍的多菌灵溶液浸泡2 h,一把伞南星种子的萌发效果最好,此时发芽率、发芽势、发芽指数达到最大值,分别为(86.79±2.73)%、(80.40±2.67)%、(9.83±1.21)%,且与对照差异极显著;其次是浓度为5%的甲醛溶液浸泡5 min,种子发芽率为(86.40±1.13)%。使用200 mg·L^-1 GA 3溶液浸泡24 h,种子根长、芽长均最大,分别为(29.76±6.04)mm、(89.46±4.18)mm,长势最好。综合考虑各萌发性状,各药剂对一把伞南星种子萌发的促进效果依次为多菌灵>GA 3>高锰酸钾>过氧化氢>甲醛>硫酸。     

甘蓝型油菜成熟角果内源激素对带壳种子发芽的影响

以甘蓝型油菜品系832、780、1-1、1-13、8-6为材料,进行了田间带壳种子发芽调查、室内模拟降雨带壳种子发芽试验、种子发芽试验、不同浓度ABA处理的种子发芽试验,并利用HPLC测定了鲜果壳和种子的吲哚乙酸(IAA)、细胞分裂素(CK)和脱落酸(ABA)的含量。结果表明,各供试甘蓝型油菜品系带壳种子发芽率存在极显著差异,而不带壳种子的发芽率在第5 天以后没有显著差异。油菜成熟角果的果壳ABA含量与带壳种子发芽存在显著负相关（r＝－0.8941）,带壳种子发芽主要受果壳所含ABA的抑制;外源ABA能明显抑制种子发芽。