RNAi沉默烟草GA 20-氧化酶基因研究

赤霉素是一种高效能的广谱植物生长调节剂，为五大植物激素之一，具有重要的生物学功能。目前利用赤霉素突变体研究生物合成途径和信号转导已经成为热点。 GA 20-氧化酶是GA生物合成中的一类关键酶，它位于GA合成途径的中心位置。本研究根据烟草（Nicotiana tabacum���GA 20-氧化酶基因序列，设计2对分别含有特定酶切位点的特异引物，以烟草基因组DNA为模板，扩增目的基因(约250 bp)片段。将正、反向目的片段分别插入中间载体的内含子两侧，再经BamH I和Sac I双酶切回收约700 bp的目的片段，插入到双元载体质粒p2355中，成功构建了含GA 20-氧化酶基因片段反向重复序列的植物表达载体p23700。分别将p2355质粒和p23700质粒导入根癌农杆菌（Agrobacterium tumefaciens）EHA105中并转化烟草叶片细胞，经卡那霉素选择培养，PCR及GUS组织染色鉴定，获得转基因烟草植株。以EHA105-p2355转化的烟草，获得41株转基因植株，均没有矮化表型；而以EHA105-p23700转化的烟草，获得转基因植株14株，其中具有矮化表型的烟草10株，表明反向重复序列转录产物能形成发夹RNA(hpRNA)，产生小分子干扰RNA（small interferring RNA，简称siRNA），干扰目的基因的表达。 赤霉素含量测定表明矮化植株中赤霉素合成途径的最终产物GA3总含量明显低于野生型烟草植株。荧光定量PCR结果表明，矮化转基因烟草的GA 20-氧化酶基因表达量受到明显抑制，表达量明显低于野生型对照。同时对上游内根-贝壳杉合成酶（Ent-kaurene synthase，KS）基因，下游的GA-3β羟化酶基因进行了RT-PCR分析，结果显示上游基因的表达没有规律性变化，而下游基因表达量亦降低。上述结果表明，GA 20-氧化酶基因的表达被有效地干扰了，表达受到抑制，从而影响植株体内GA3的合成，影响植株的生长发育，导致植株矮化。并推测，GA 20-氧化酶基因受到抑制，可能影响下游基因的表达。并且通过干旱胁迫测试，发现矮化植株相对于野生型植株及不含干扰片段的转基因植株，对干旱的耐受力有了很大的提高，具有更强的耐受力。 研究结果为进一步进行相关研究奠定基础。 Gibberellin(GA) is an efficient plant growth regulator. As one of five major plant hormones, it plays an important biological function. Using GA mutant for investigating biosynthetic pathways and signal transduction has become high lights. GA 20-oxidase is a crucial enzyme involved in gibberellin biosynthesis. According to tobacco (Nicotiana tabacum) GA 20-oxidase enzyme gene sequence and based on binary vector p2355, we constructed a plant expression vector p23700, which habors an inverted repeat DNA fragment of GA 20-oxidase gene drivered by Cauliflower mosaic virus promtor (CaMV 35Sp). Binary plasmid p2355 had no inverted repeat DNA fragment of GA 20-oxidase gene. The vector p2355 and p23700 were introduced into Agrobacterium tumefaciens EHA105 and tobacco leaf transformation was conducted. After selected by kanamycin and characterized by PCR and GUS hischemical reaction, transsgenic plants were obtained. Fourtheen transgenic plants, which were transformed by EHA105-p23700, were obtained. Among them, 10 were dwarf mutants. However, 41 transgenic plants with the same normal phenotype as wild type,which were transformed by EHA105-p2355, were obtained. Analysis of Gibberellin contents showed that it was lower in dwarf mutants than in normal phenotype plants. Moreover, comparing to normal phenotype plants including wild type and transgenic plants with no interference fragment, the drought tolerance of dwarf plants have greatly increased. And their proline content increased obviously after drought test. Fluorescence quantitative real time PCR (RT-PCR) showed that GA 20-oxidase gene expression was significantly inhibited in dwarf transgenic tobacco. Meanwhile, the expression of the upstream gene ent-kaurene synthase (KS) gene and downstream gene GA-3β hydroxylase gene was also detected by RT-PCR. The results presented that KS gene expression had no regular change while GA-3β hydroxylase gene expression reduced. It implied that inhibiting GA 20-oxidase gene probably reduce the expression of downstream genes. The results showed that the transcriptional products of the foreign inverted repeat fragment can form hairpin RNA (hpRNA) to induce RNAi. It presented that GA 20-oxidase gene expression was effectively interfered, resulting in reducing GA3 synthesis and inhibiting plant growth and development, then dwarf plants were produced. However, the dwarf plants had higher tolerance of drought.

80MeV/u~(20)Ne轰击~9Be碎裂产物反应总截面的测量

介绍了用透射法测量中能区2 0 Ne打9Be靶碎裂产生的次级束与Si靶作用的核反应总截面的方法 ,以及实验的探测器布局、实验过程和实验结果 .并对理论上预言有奇异结构的核1 2 N ,1 7Ne和1 7F的实验结果与其相邻核进行了比较

~(16)O(α,γ)~(20)Ne反应的实验研究

讨论了目前16O(α ,γ) 2 0 Ne反应的实验研究工作 ,并对2 0 Na β+—→2 0 N →16O +α的β+ 缓发衰变α能谱的相对强度值同文献进行了比较 ,讨论了低能激发能级、共振强度及其对共振截面的贡献、S因子的理论研究 .

β~+延发粒子衰变的一种在束测量方法

报道了一种在束测量β+ 延发粒子的方法。由新建成的兰州放射性次级束流线 (RIBLL )提供的2 0 Na次级束 ,利用飞行时间 (TOF)和能损 (ΔE)符合的方法实现 2 0 Na次级束流的在束鉴别与调制。将数据获取过程分为有束和停束两个获取时段 ,分别完成对次级束流和β+ 延发粒子的记录。同时利用脉冲发生器和计数器实现2 0 Na延发粒子衰变半衰期的测量

~(20)Na的β~+延发α粒子测量

利用兰州放射性次级束流线提供的2 0 Na束流 ,通过2 0 Na β+2 0 Ne →16 O+α过程 ,测量了2 0 Na的衰变半衰期T1/2 及衰变α粒子能谱 .结果表明 ,除了Ed≥ 2 .688MeV的 9条较高激发能级的衰变α粒子外 ,实验中还观察到衰变能量Ed 为 0 .890和 1 .0 54MeV ,1 .991MeV ,2 .4 2 4和 2 .4 57MeV的2 0 Ne低激发能级的 3条α谱线 .

50MeV/u~(20)Na+~(208)Pb→p+~(19)Ne+~(208)Pb反应的探测效率模拟

用半经典模型考虑20Ｎａ＋２０８Ｐｂ→ｐ＋１９Ｎｅ＋２０８Ｐｂ的Ｃｏｕｌｏｍｂ解离过程。用ＭｏｎｔｅＣａｒｌｏ方法计算前角放置的望远镜阵列对Ｃｏｕｌｏｍｂ解离碎片的探测效率。讨论了天体环境下ｐ＋１９Ｎｅ→２０Ｎａ俘获反应道的低能级共振截面测量的可行性。

Double ionization of helium in collision with 20-500-keV/amu Cq+ and Oq+ (q=1-3) ions

Target ionization and projectile charge changing were investigated for 20-500 keV/u Cq+, Oq++He (q=1-3) collisions. Double- to single-ionization ratios R-21 of helium associated with no projectile charge change (direct ionization), single-electron capture, and single-electron loss were measured. The cross-section ratio R-21 depends strongly on the collision velocity v, the projectile charge state q, and the outgoing reaction channel. Meanwhile, a model extended from our previous work [J. X. Shao, X. M. Chen, and B. W. Ding, Phys. Rev. A 75, 012701 (2007)] is presented to interpret the above-mentioned dependences. Good agreement is found between the model and the experimental data.