Wheat, triticale, tritordeum, barley, oat and rye are the most important crops in human consumptions and industry in the world. Transformation technology supplies a new source of improving Triticeae crops. In the past decade, transformation of wheat crops has considerably progressed. Many transgenic plants of Triticeae crops with various genes were produced via microprojectile bombardment,Agrobacterium mediated transformation, PEG uptake DNA technique, electroporation, microinjection, injection inflorescence and silicone carbide. Integration and expression of transgenes, inheritance and variation of transgenic plants have been studied. Technical improvements of genetic transformation for wheat crops will be extensively useful in commerce and benefit significantly to human being in the world.
李义文1* 徐龙源2 李振声1 贾旭1
麦类作物包括小麦(Triticum aestivum L.)、硬粒小麦(Triticum turgidum con v.durum Dest.e.m)、大麦(Hordeum vulgare L.)、黑麦(Secale cereal L.)、燕麦(Avena sativa L.)及小大麦(×Tritordeum Ascherson et Graebuer.).自从基因枪被发明以来,科学家们已经利用来自麦类作物的幼胚、 盾片、成熟种子胚、花粉粒、花药、幼穗、叶基组织、发芽种子幼苗的顶端分生组织及其愈伤组织或培养物作为外植体,通过基因枪、农杆菌介导、 PEG法、电激法、微注射法、硅化纤维素介导、幼穗注射法等技术先后将一些选择标记基因、报告基因和有用的目的基因如抗真菌、抗虫、 籽粒品质、抗干旱基因等转化到麦类作物中.转基因植物表现为抗性增强或籽粒的加工品质提高和营养成份增加.被转化的基因通常以单位点多拷贝的形式随机整合到受体细胞的基因组中,并以孟德尔规律遗传.整合位点一般分布在染色体的近端粒区域,整合的拷贝数大多为5～10个拷贝,最高可达到50个拷贝.在转化过程中,被转化的质粒上的片段包括选择标记基因、目标基因、甚至质粒的抗生素基因和其他无关序列,随机地连接并形成多个分子量大小不等,组成成分不同的分子簇,或首先由其中一个分子簇整合到植物基因组中,这会导致在整合位点附近产生"热点",易于其他分子簇在此处整合,从而完成两期整合;或被转化的质粒上的选择标记基因、目标基因、质粒的抗生素基因和其他无关序列、植物基因组DNA等片段共同形成各种不同类型的分子簇,当植物细胞染色体复制时,在复制叉处整合到植物基因组中.转基因可以在各种水平上表达,也会时常发生基因沉默,这会导致转基因植物DNA水平上表达但在蛋白质水平上不表达,后代偏向分离,沉默的转基因重新表达.转基因的位置效应、甲基化和启动子都会诱发转基因沉默.在麦类作物中,35S启动子易于导致转基因沉默,应尽量减少使用.转基因还导致被转化麦类作物在农艺性状和细胞学上的变异.目前,麦类作物遗传转化已经成为一种常规的技术,转基因麦类作物正开始进入商业应用阶段.相信多种转化新技术的应用和发展将会培育出高产、稳产、优质、低投入的各类品种和种质.
关键词 麦类作物 转化技术 转基因植株 基因表达和变异
*通讯作者 Email: email@example.com
Ficus hispida L. (Moraceae) is a remarkable species in the ecosystem of tropical rainforests in Xishuangbanna, China. The figs and fig pollination wasps (Chalcidoidae: Agaonidae) are highly co evolved mutualists that depend completely on each other for propagating descendants. Pollination of all fig species is do ne by fig wasps; their unique symbiotic associates, the fig wasps, cannot develop in anywhere except in the fig syconia. The present paper reports on the biology and flowering phenology of F. hispida, as well as the propagation character and pollination behavior of the fig wasps (Ceratosolen solmsi marchali Mayr) based on our observations in the rainforests of Xishuangbanna, southern Yunnan of China. F. hispida is a dioecious tree that annually blossoms and bears fruits 6-8 times, with four to five fruit bearing peaks. The male trees produce pollen and provide fig wasps with reproductive havens, while the female trees produce fig seeds after pollination by the female wasps. Pollen of F. hispida cannot escape from the dehiscent anthers until they are disturbed by fig wasps. The female wasps open the anthers and collect pollen with their antennal scrapes, mandibles and legs, and then carry pollen to the female receptive syconia where fertilization takes place. Meanwhile, some of the female wasps lay eggs in the male receptive syconia. It takes about 3-67 min to search for the receptive syconia for pollination, and 15-23 h to enter the female receptive syconia. The number of female wasps entering a syconium has close relation with the impregnation and seed bearing rate of female flowers, as well as the oviposition and reproduction rate of the fig wasps themselves. F. hispida is endowed with a relatively high level of seed bearing (54.1%-82.5%, average 73.8 % ). The wasp oviposition rate on the male flowers is between 72.3% and 93.8% with a mean of 84.4%.
杨大荣* 彭艳琼 宋启示 张光明 王瑞武 赵庭周 王秋艳
研究了西双版纳热带雨林地区雌雄异株植物对叶榕(Ficus hispida L.)的生物学、传粉物候学以及榕小蜂(Ceratosolen solmsi marchali Mayr)的传粉和繁殖行为.研究结果表明:雌雄异株的对叶榕与其他雌雄同株的榕属植物不同,它的种子形成与传粉者有着更为复杂的相互关系.对叶榕一年结隐花果6～8次,结果高峰期4～5次,其中雄性植株仅产生花粉和孕育榕小蜂,而雌性植株(无雄蕊)则需榕小蜂带花粉进入隐花果内,进行传粉授精,使之发育成种子.对叶榕的成熟花粉不能从花药开裂处自行散发出来,必须由榕小蜂采集才能散落.榕小蜂雌蜂羽化、交配后,找到雄花区,用足、触角、口器在推动中采集花粉.雌蜂飞出熟榕果找寻雌株或雄株榕树上的幼嫩隐花果,一般需3～67 min;一部分雌蜂在雄株中寻找幼嫩的隐花果,进去产卵繁殖,另一部分雌蜂则寻找雌株雌花期嫩隐花果进去传粉.雌蜂在雌株榕树的隐花果内传粉时间为15～23 h,在雄株榕树的隐花果内产卵时间为6～9 h.对叶榕小蜂在雌株上进入单个隐花果的数量多少关系到雌花结实率;观察表明,每个隐花果最佳进蜂数为2头;榕小蜂传粉后榕树成熟种子形成率在54.1%～82.5%之间,平均为73.8%;而在雄株上雌蜂进蜂数量则关系到榕小蜂在隐花果内的产卵率,每个隐花果最佳进蜂数为3～4头,产卵率在72.3%～93.8%之间,平均为84.4%.
关键词 叶榕 对叶榕小蜂 传粉生物学 热带雨林 西双版纳
*通讯作者 Email: firstname.lastname@example.org
In order to investigate the morphological response of freshwater green algae toelevated CO2 concentration, Chlamydomonas reinhardtii Dang and Scenedesmus obliquus Kütz were cultured with enriched CO2, and their microstructure and ultrastructure were examined by microscopy and electron microscopy. The effect of CO2 enrichment to 186 μmol/L was insignificant on the shape and size of C. reinhardtii, but significant in reducing the volume of S. obliquus. High CO2 increased the amount of chloroplast. The pyrenoids occurred in low CO2 grown cells but not in high CO2 grown ones and more starch granules were observed in the former.
为了探讨淡水绿藻在适应CO2浓度变化过程中细胞形态和结构的变化,通过普通显微镜和电子显微镜观察了在不同CO2浓度培养下的莱因衣藻(Chlamydomonas reinhardtii Dang)和斜生栅藻(Scenedesmus obliquus Kütz)细胞.结果表明,CO2浓度变化对莱因衣藻细胞体积没有明显的影响,但斜生栅藻在低浓度CO2培养下细胞体积明显增大,并可见细胞内含有大量颗粒.两种绿藻细胞的超微结构显示,在低浓度CO2培养下,细胞内叶绿体数目明显减少,并可见明显的淀粉盘包围的蛋白核;细胞内还可见大量的淀粉粒.而在高浓度CO2培养下,这两种绿藻细胞内均未见明显的蛋白核和大量淀粉粒出现.
关键词 莱因衣藻 CO2 显微结构 蛋白核 斜生栅藻 淀粉粒 超微结构
Auxin influences a variety of developmental and physiological processes. Early reports suggested that auxin might affect plant stress response. We have identified a number of auxin responsive genes in Arabidopsis thaliana (L.) Heynh. by using cDNA array and found that stress responsive genes, such as Arabidopsis homolog of MEK kinase 1 (ATMEKK1), RelA/SpoT homolog 3 (At-RSH3), Catalase 1 (Cat1) and Ferritin 1 (Fer1),were down regulated by auxin, indicating that auxin regulates the expression of stress responsive genes. We also demonstrated that nitrilase genes, nitrilase 1 (NIT1) and nitrilase 2(NIT2) involving in indole 3 acetic acid (IAA) biosynthesis, were induced by salinity stress, suggesting that the level of IAA might increase in response to salinity stress. To dissect the signal pathway involved in the interaction, two auxin insensitive mutants, auxin resistant 2 (axr2) and auxin resistant 1-3 (axr1-3) were used. Stress responsive genes were induced by salt stress in wild type and axr2, but not in axr1 3. The result suggests that the interaction between auxin and stress responses may be linked in the ubiquitin pathway.
包 方 李家洋*
摘要： 生长素影响植物多种生理过程,有报道显示生长素可能影响植物对逆境胁迫的反应.我们利用cDNA阵列技术鉴定拟南芥(Arabidopsis thaliana (L.) Heynh.)的生长素应答基因,发现多个胁迫应答基因受生长素抑制,包括Arabidopsis homolog of MEK kinase1 (ATMEKK1),RelA/SpoT homolog 3 (At-RSH3),Catalase 1 (Cat1) 和Ferritin 1 (Fer1),说明生长素可调节胁迫应答基因的表达.此外,我们还证明吲哚乙酸(IAA)合成途径中的腈水解酶基因nitrilase 1 (NIT1) 和nitrilase 2 (NIT2) 受盐胁迫诱导,提示在逆境条件下IAA的合成可能随之增加.我们利用生长素不敏感突变体研究生长素与逆境反应相互作用的信号转导,发现胁迫应答基因在野生型和生长素不敏感突变体auxin resistant 2 (axr2) 中可被盐胁迫诱导,而在auxin resistant 1-3 (axr1-3)中则不被诱导,说明生长素与逆境胁迫反应的相互作用可能发生在泛素途径.
通讯作者。E-mail:jyli @ genetics.ac.cn.
The accumulation of citric acid and associated correlative strain indexes were investigated in the seedlings of Puccinellia tenuiflora (Griseb.) Scribn. et Merr. stressed with 0-175 mmol/L Na2CO3. The results showed that the citric acid accumulation is a specific physiological respond of this plant to alkali stress. On the contrary, the citric acid content decreased slightly in this plant stressed with 0-400 mmol/L neutral salt NaCl. The accumulation of citric acid increased with increasing strength of alkali stress, the citric acid content increased gently when the strength was lower than 100 mmol/L Na2CO3, but increased obviously when the strength was higher than 100 mmol/L Na2CO3. The citric acid rapidly accumulated at early alkali stress, an obvious raise can be mensurated after 4 h. About 48 h after treatment, the amount of citric acid accumulated nearly reached the maximum. In various parts of P. tenuiflora seedlings alkali stressed for 144 h, the order of citric acid content from high to low is: old leaf, mature leaf, old leaf sheath, young leaf sheath, young stem, old stem, and young leaf. In the mature leaf, the citric acid content gradually increased with increasing strength of alkali stress, while the citric acid content increased sharply in old leaf and sheath just for strength higher than 125 mmol/L. There was little change of citric acid content in stem, but no change in young leaf. The results of the experiment showed that citric acid was outstanding among accumulated organic acid caused by alkali stress. The contribution of other organic acids was negligible.
石德成1 尹尚君2 杨国会1 赵可夫3
（1. 东北师范大学生命科学学院，长春130024；2. 浙江农业技术师范专科学校，宁波315001；
摘要：对碱胁迫(0-175 mmol/L Na2CO3)下星星草(Puccinellia tenuiflora (Griseb.) Scribn.et Merr.)体内柠檬酸的积累规律及其相关胁变指标进行分析测定.实验结果证明:积累柠檬酸是星星草对碱胁迫特有的生理反应.盐胁迫(0-400 mmol/L NaCl)下,柠檬酸含量反而稍有下降.柠檬酸积累量随碱胁迫强度增大而增大,低胁迫强度时积累量上升缓慢,当胁迫强度大于100 mmol/L Na2CO3时,积累量明显上升.柠檬酸积累与胁迫时间之间呈直角曲线关系,一定胁迫强度下胁迫4 h后即可测出柠檬酸含量明显上升,约48 h后渐趋最大值.碱胁迫144 h后星星草各部位中柠檬酸含量从高到低的顺序依次是老叶、成熟叶、老叶鞘、幼叶鞘、幼茎、老茎和幼叶.成熟叶中柠檬酸随碱胁迫强度增大而逐渐上升,老叶和叶鞘中的柠檬酸在碱胁迫强度大于125 mmol/L后急剧上升,茎中柠檬酸含量无明显增高,幼叶中柠檬酸含量基本不变.实验证明,碱胁迫下积累的主要是柠檬酸,其他有机酸无明显变化.
R phycoerythrin, a light harvesting protein in some marine algae, and can be widely used in medicine, was isolated and purified from a red alga, Palmaria palmata (Lannaeus) Kuntze, using the streamline column (expanded bed adsorption) combined with ion exchange chromatography. Because the crude extract was applied to the column upwardly, the column would not be blocked by polysaccharides usually very abundant in the extract of marine alga, this kind of blockage could hardly be overcome in ordinary chromatographic column. After applying the crude extract containing 0.5 mol/L (NH4)2SO4, (NH4)2SO4 solution of different concentrations (0.2 mol/L, 0.1 mol/L and 0.05 mol/L) was used to elute the column downwardly and the eluates were collected and desalted. The desalted eluates were then applied onto an ion exchange chromatographic column loaded with Qsepharose for further purification of the R phycoerythrin. Through these two steps, the purity ( OD565/OD280) of the R phycoerythrin from P． palmata was up to 3.5, more than 3.2, the commonly accepted criterion for purity, and the yield of the purified R phycoerythrin could reach 0.122 mg/g of frozen P. palmata, much higher than that of phycobiliproteins purified with the previous methods. The result indicated that the cost of R phycoerythrin will drop down with the method reported in this article.
王广策 孙海宝 范晓 曾呈奎
以Phenyl-sepharose作为柱填料,用streamline柱层析技术从红藻(Palmaria palmata (Lannaeus) Kuntze)中规模分离捕光色素蛋白--R-藻红蛋白.由于不是传统的从层析柱上方进样,而是用泵将样品从streamline层析柱的下方加样(从下至上),因而解决了用一般层析柱分离R-藻红蛋白时海藻抽提液中大量的粘性多糖堵塞层析柱的难题.用P.palmata粗提液上样后,分别用0.2 mol/L、 0.1 mol/L和0.05 mol/L的(NH4)2SO4溶液从相反的方向(即从上到下)洗脱层析柱,发现这些洗脱液中的藻红蛋白纯度已经较高.然后将洗脱液透析去盐,用阴离子交换柱层析(Q-sepharose)进一步纯化.经过这两次柱层析后,R-藻红蛋白的纯度(OD565/OD280)超过3.5,高于一般认可的R-藻红蛋白的纯度标准3.2;产率为每克冷冻P.palmata可纯化0.122 mg高纯度的R-藻红蛋白,比使用一般分离方法的产率要高10倍.这些结果表明,使用本文报道的方法纯化藻红蛋白,将会使作为生化检测试剂的藻红蛋白市场价格大幅度下降.
关键词 红藻 R-藻红蛋白 streamline柱层析 离子交换柱层析
Fourier transform infrared spectroscopy (FTIR) was used to detect the spectral difference among leaf tips from rice (Oryza sativa L.) plants with different nitrogen fertilizer rates. The proposed spectral index, (A3400-A1653)/(A3400+A1653) (A denotes absorption valuet certain frequency (cm-1)), was found to decline with the increase of nitrogen fertilizer rates and the results suggested that FTIR may be useful to diagnose nitrogen status in crops.
周启发1 沈掌泉2 王人潮2
（1. 浙江大学生命科学学院，杭州310012；2. 浙江大学农业遥感与信息技术应用研究所，杭州310029）
摘要： 四个氮素水平处理的盆栽水稻(Oryza sativa L.)的叶尖在不同生育期均表现出明显的傅里叶转换红外光谱差异.新定义的光谱指数((A3400-A1653)/(A3400+A1653),A为某频率处的吸收值)随着施氮水平的提高而降低.结果表明,傅里叶转换红外光谱可用于诊断植物的氮素状况.
通讯作者。E-mail:qifaz @ mail.hz.zj.cn.
During the course of mungbean ( Phaseolus radiatus L.) germination, the rate of ethylene production and the activity of ACC synthase (1 aminocyclopropane 1 carboxylic acid synthase, EC22.214.171.124) began to increase in the 5th day of germination, and reached its peak in the 10th day and then decreased. The ethylene production and the activity of ACC synthase were obviously promoted by 10 μmol/L exogenous IAA (indole 3 acetic acid).The production of superoxide radical (O[SX(B-*3]-·)]2) and hydrogen peroxide (H2O2) were also promoted by exogenous IAA, suggesting that there was some relationship between active oxygen production and the activity of ACC synthase induced by exogenous IAA. The production of ethylene and the activity of ACC synthase increased dramatically when the seedlings were treated with exogenous O[SX(B-*3]- ·[SX)]2, whereas the exogenous H2O2 had no effects on the production of ethylene and the activity of ACC synthase. Exogenous SOD (superoxide dismutase, one scavenger of O[SX(B-*3]-·[SX)]2) could inhibit the production of ethylene and the activity of ACC synthase, but exogenous CAT (catalase) could not. So it was possible that IAA would stimulate the activity of ACC synthase by inducing the production of O[SX(B-*3]- ·[SX)]2 in germinating mungbean seedlings, and this might be one of the regulating mechanism of ethylene synthesis in higher plants; the production of H2O2 induced by IAA was not the cause of the increase of the activity of ACC synthase and the production of ethylene.
柯德森1 王爱国1 孙谷畴1 董良峰2
（1. 中国科学院华南植物研究所，广州510650；2.Department of Biological Sciences,Unixersity of Essex, Colchester CO4 3SQ, UK
摘要： 本文试图从活性氧的角度阐明外源IAA诱导ACC合酶活性的机制.绿豆(Phaseolus radiatus L.)幼苗的乙烯产生及ACC合酶活性从萌发的第5天开始上升,到第10天达到高峰,接着下降.10 μmol/L的外源IAA能明显促进绿豆幼苗乙烯的产生及ACC合酶的活性,同时也促进了超氧阴离子自由基(O2-)、过氧化氢(H2O 2)的产生.显示外源IAA诱导的ACC合酶的活性与其诱导的活性氧的产生具有某种相关性.外源O2-)处理能明显提高绿豆幼苗的乙烯产生速率及ACC合酶的活性,而外源H2O2无论对乙烯产生或ACC合酶的活性均没有明显的作用.外加(O2-)的清除剂SOD对绿豆幼苗乙烯的产生及ACC合酶活性的提高有一定的抑制作用,而外源过氧化氢酶却没有明显的作用.为此我们可以得出结论:外源IAA诱导的绿豆幼苗ACC合酶活性的提高可能是由于其诱导的O(O2-)产生的升高引起的,这可能也是高等植物中调控乙烯生物合成的机制之一;而IAA诱导的H2O2产率的升高并不是其诱导ACC合酶活性升高的原因.
Comparative studies of absorption spectra of phycobiliproteins of Gracilaria lemaneiformis Greville and its pigmental mutants were conducted in this study. The results showed that the absorption spectra of phycoerythrins (PE) from different material changed significantly, while those of phycocyanins(PC)and allophycocyanins (APC) were basically similar. In order to disclose the essence of the difference, partial sequences of the subunit genes of PE of Qingdao strain of G. lemaneiformis (qd) and its pigmental mutants were determined. The amino acid sequences were deduced and used to explain spectral shifts of PE from the pigmental mutants. The amino acid sequences of PE resembled each other, and several residues changed among qd and its pigmental mutants. Residue substitutions were found in a region consisting of amino acids which determined the secondary structure and subunits interactions, thus might influence the confirmation and interaction of subunits, and further caused spectral deviation.
隋正红 张学成* 程晓杰
摘要：对龙须菜(Gracilaria lemaneiformis Greville)及其色素突变体藻胆蛋白吸收光谱进行了比较研究,结果显示不同藻株藻红蛋白的吸收光谱有显著的变化,而藻蓝蛋白和别藻蓝蛋白的基本相同.我们克隆了龙须菜及其色素突变体的藻红蛋白亚基的部分基因序列,用该基因序列推导出的氨基酸序列进行分析以揭示这一变化的分子机理,结果显示除几个氨基酸残基的替换外,几株藻间的藻红蛋白的氨基酸序列十分相似,一些氨基酸的替换发生在决定藻红蛋白二级结构及亚基间相互作用的区域,可能会影响藻胆蛋白的构型及相互作用,导致光谱性质的变化.
!通讯作者。E-mail:xczhamg @ ouqd.edu.cn.
A gene homologous to the human putative tumor suppressor gene QM, designated OSQM1, was isolated from rice (Oryza sativa L.) genomic DNA library using through homology screening. It contained 4 exons and 3 introns, encoding a protein of 219 amino acids with 46 basic amino acids, leading to a high isoelectric point of 11.02. Homology search showed that this gene existed in eukaryotes and highly conserved throughout eukaryotes, suggesting an essential role of this gene. Northern blot analysis showed that it was expressed in various rice organs, but at lower level in developing flower and callus tissue than in other vegetative organs. Its expression levels in roots and leaves were influenced by different environmental factors.
葛晓春* 宗 晖 詹树萱 陈继超 孙崇荣 曹凯鸣
摘要： 用同源筛选方法 ,从水稻 (OryzasativaL .)基因组文库中分离到一个与人类肿瘤抑制基因QM具有同源性的基因 ,命名为OSQM1。该基因包括 4个外显子和 3个内含子 ,编码 2 19个氨基酸 ,其中有 4 6个碱性氨基酸 ,其等电点高达 11.0 2。同源性搜寻发现此基因存在于真核生物中而且保守性较强 ,表明它可能具有重要的作用。North ern分析结果表明 ,它在不同的水稻器官中都有表达 ,但在花和愈伤组织中的表达水平明显低于其他营养器官。它在根和叶中的表达水平受环境因素的影响。
Powdery mildew is a serious disease of wheat in China. As part of ITEC (International Triteace EST cooperation), EST （expressed sequence tags） technique was used to explore the gene expression in leaf induced by Erysiphe graminis DC. A conventional cDNA library was constructed, and a total of 1 500 clones picked randomly from the library were sequenced, three hundred and eighty seven ESTs of them were unique, which got the Accession Number in GenBank. About 49.4% ESTs showed significant similarity to functions of known sequences in GenBank. There are 196 ESTs' with functions not able to be determined, and eighty four ESTs were demonstrated to be novel sequences. High density dot membranes from unique clones were produced, and several disease resistance related genes were screened by differential hybridization.
骆蒙 孔秀英 姜涛 贾川 周荣华 贾继增*
白粉病是我国小麦的主要病害之一.尝试用表达序列标签(expressed sequence tags, EST)技术,研究了经白粉病菌诱导后的小麦基因表达.从构建的普通cDNA文库中随机挑取约1 500个阳性克隆并进行测序, 获不重复ESTs序列387条.不重复序列均获GenBank的存储号.其中49.4%的序列与已知基因同源,196条序列功能未知, 84条序列为新ESTs.将不重复序列制备成高密度点阵膜,用差示杂交法筛选到几个抗病相关序列.
关键词 小麦 白粉病 表达序列标签 抗病相关基因
*通讯作者 Email: email@example.com
A vacuolar ATPase (V ATPase) B subunit gene has been cloned and characterized from a phosphorus starvation induced rice root subtractive cDNA library by suppression subtractive hybridization (SSH) method and RT-PCR amplification. This gene encodes a polypeptide of 487 amino acid residues, containing a conservative ATP binding site and with a molecular weight of 54.06 kD and an isoelectric point of 4.99. Southern analysis of the genomic DNA indicates that V-ATPase B subunit is encoded by a single gene in rice genome. The amino acid homologies of V-ATPase B subunits among different organisms range from 76% to 97% and reveals that the evolution of V-ATPase B subunit is accompanied with the biological evolution. Expression pattern analysis indicated that the maximal expression of V-ATPase B subunit gene occurred at an early stage (6-12 h) after phosphorus starvation in roots, and lately stage (24-48 h) in leaves. Under phosphorus deficiency, the up regulated expression of V-ATPase gene was presumed to strengthen the proton transport and provide the required energy to maintain an electrochemical gradient across the tonoplast to facilitate phosphorus transport.
水稻液泡ATPase B 亚基基因的克隆及其在低磷
夏 铭 王小兵 李海波 吴平*
摘要： 利用抑制性扣除杂交(SSH)技术构建水稻(Oryza sativa L.)根系磷饥饿诱导cDNA文库,获得编码液泡ATPase (V-ATPase) B亚基的克隆,通过反转录PCR方法获得该基因的完整序列.该基因编码487个氨基酸,含有一个保守的ATP结合位点,其蛋白分子量为54.06 kD,等电点为4.99.Southern印迹表明,V-ATPase B亚基基因在水稻基因组中以单拷贝形式存在.氨基酸同源性分析发现,V-ATPase B亚基是一个较为保守的蛋白亚基,其序列变化伴随生物的进化过程同步进行.Northern印迹表明,V-ATPase B亚基在水稻根系中受到磷饥饿诱导表达,磷饥饿6～12 h出现表达高峰,而在叶片中表达高峰有所滞后(24～48 h).在缺磷环境条件下,ATPase B亚基可能通过提高其表达量,进而提高质子转运活性,形成跨膜的电化学梯度,为体内储备磷跨液泡膜运输提供能量,从而提高植物体内磷的利用效率及其耐低磷的能力。
关键词： 水稻；液泡ATPase B 亚基；磷饥饿；抑制性扣除杂交（SSH）；基因克隆
通讯作者。E-mail: docpwu @ cls.zju.edu.cn,Tel:0571-86971130
Strictosidine synthase (STR) is a key enzyme involved in the biosynthesis of terpenoid indole alkaloids (TIA) by condensing tryptamine and secologanin into strictosidine. The transgenic tobacco plants targeting STR to subcellular compartments were established to express STR in chloroplast, vacuole and endoplasmic reticulum (ER) by the tobacco stable transformation. It was shown that STR was effectively expressed in the above subcellular compartments by Western blot analysis and STR enzymatic assay. In vitro, STR enzymatic assay was measured indirectly by fluorimetrically detecting depletion of tryptamine feeding on secologanin in the reaction mixture. The tryptamine were completely depleted by STR in the crude extract of leaves of transgenic tobacco plants targeting and expressing STR in the chloroplast, vacuole and ER, which ascertained the STR functionally targeted to the three subcellular compartments. To confirm STR correct targeting and expressing in chloroplast, the chloroplasts were isolated and the fractions of purified chloroplasts were analyzed by Western blot. The hypothesis of STR correct targeting to the chloroplast was tested. The results have implications on our understanding of the complex intracellular trafficking in metabolic intermediates of TIA biosynthesis.
王淼* 李秋荣2 Stefano Di FIORE3 Rainer FISCHER 3
（1. 中国科学院沈阳应用生态研究所，沈阳110016；2. 沈阳军区总医院，沈阳110016；
3.Botanical Institute of Rheinisch-Westfalisch Technische Hochschule Aachen,D-52074 Aachen,Germany)
A FISH procedure was adopted to physical mapping rice RFLP marker RZ69 and the BAC clone 38J9 screened by RZ69 which is linked to gene Bph3 in Oryza sativa L. and O. officinalis Well ex Watt. The FISH results showed that both 38J9 and RZ69 were located in the middle of 4S in O. officinalis and the centromere area of 4S in O. sativa. In O. officinalis the percentage distances from the centromere to the hybridization sites were 20.00±5.40 and 22.12±3.44, the detection rates were 50.00% and 6.14%, but in O.sativa they were 0 and 0,56.10% and 6.29% correspondingly. The results obtained from the BAC and RFLP clone were almost the same in the cultivated rice and O. officinalis. It was suggested that the marker RZ69 of the cultivated rice and its homologous sequence in O.officinalis were in the same BAC clone, the homologous sequence of Bph3 in O. officinalis should be located at the sites hybridized by both RZ69 and 38J9. Many signals on different chromosomes of O. officinal is were observed under no blocking with Cot-1 DNA, showing that the repetitive sequences were also homologous between O. sativa and O. officinalis. The identification of chromosome 4 of O. officinalis is based on comparative map with RG214 and BAC clone screened by RG214. The feasibility of comparative physical mapping performed between O. sativa and O. officinal is with rice BAC clones was discussed.
鄢慧民1 覃 瑞1,2 金危危1 何光存1 宋运淳1*
（1. 武汉大学植物发育生物学教育部重点实验室，武汉430072；2. 中南民族学院化学与生命科学学院，武汉430074）
摘要： 用栽培稻(Oryza sativa L.)遗传图第四连锁群中与抗褐稻虱基因Bph3紧密连锁的RFLP标记RZ69及筛选出来的BAC克隆38J9作探针,对药用野生稻(O.officinalis Well ex Watt)和栽培稻荧光原位杂交,供试标记RZ69及38J9均被定位于药用野生稻和栽培稻第4染色体的短臂上,药用野生稻杂交信号的百分距分别为22.12±3.44和20.00±5.40,而栽培稻均为0.在栽培稻中,信号检出率相应地为6.29%和56.10%,在药用野生稻中则为6.14%和50.00%.BAC克隆和RFLP标记探针杂交信号的百分距十分接近,说明在栽培稻和野生稻中RFLP标记RZ69都在同一BAC克隆的大插入片段中.由此推知,药用野生稻与抗性基因Bph3的同源顺序就在第4染色体信号出现的相应位置.在未封阻的情况下,药用野生稻的BAC杂交在多条染色体上具有信号,这表明它和栽培稻的Cot-1 DNA重复顺序也在一定程度上具有同源性.药用野生稻第4染色体是根据栽培稻与药用野生稻的比较遗传图选用与Gm-6连锁的RG214通过FISH确定的.讨论了栽培稻BAC克隆对药用野生稻比较原位杂交物理作图的可行性问题.
通讯作者。E-mail:songyc @ whu.edu.cn.
The infrageneric classification currently in use for Cymbidium is based on gross morphology, with emphasis on the number of pollinia and state of fusion between lip and column. The sequences of nrDNA regions of 27 species and 3 cultivars of Cymbidium and 3 outgroup species (Eulophia graminea, Geodorum densiflorum, Amitostigma pinguiculum) were analyzed using PCR amplification and direct DNA sequencing. The phylogenetic trees generated from maximum parsimony analysis, however, show that the existing division among three subgenera (subgen. Cymbidium, subgen.Cyperorchis and subgen. Jensoa) should be evaluated with more data. Subgenus Cyperorchis was not a monophyletic group, with the unexpected nesting of C.dayanum (subgen.Cymbidium) within it; subgenus Jensoa also appeared paraphyletic, with C. lancifolium being the sister group to the remainder of the genus; species of subgen. Cymbidium appeared polyphyletic, being split into several clades and intermixed with the main subgen. Cyperorchis and subgen. Jensoa clades, respectively. However, because of the insufficiency of informative characters of ITS sequences, some of the clades identified, especially the major lineages of Cymbidium, received relatively low support; sectional delimitations were also not clear within each subgenus. Further study is needed for achieving a robust phylogeny of Cymbidium.
基于nrDNA ITS 序列数据的兰属系统发育关系的初步分析
张明永1 孙彩云1 郝 刚1 叶秀 1 梁承邺1 朱光华2
（1. 中国科学院华南植物研究所，广州510650; 2. 密苏里植物园，圣·路易斯，MO 63166-0299，美国）
摘要： 现存的兰属分类系统是基于宏观形态学性状、尤其是花粉块的数目以及唇瓣与蕊柱的愈合程度而建立的.兰属因此而划分为3个亚属:兰亚属 (subgenus Cymbidium),大花亚属(subgenus Cyperorchis) 和建兰亚属 (subgenus Jensoa).本文运用PCR扩增和直接测序的方法分析兰属 (Cymbidium) 27种、3个栽培品种以及3个外类群的核DNA ITS 区段序列.通过最简约性分析产生的ITS系统发育树表明,兰属的3个亚属均可能为不自然的类群.大花亚属表现为一复系群,兰亚属的冬凤兰 (C.dayanum) 隐藏于其中;建兰亚属为一并系群,它的成员之一兔耳兰 (C.lancifolium) 偏离出去而成为兰属一最基部的分支;兰亚属为一复系群,它分为几支而分别与另两个亚属组合在一起.由于兰属ITS序列位点变异率较低,最简约性分析产生的几支主要分支均得不到Bootstrap分析的高度支持,各亚属内组之间的关系也不明确.研究兰属的系统发育关系还需要新的数据.
通讯作者。E-mail:xiulinye @ scib.ac.cn.
This paper reports on two years of measurement of soil respiration and canopy root biomass in a Leymus chinensis community in the Xilin River basin of Inner Mongolia. Correlations between components of plant biomass and soil respiration rates were examined. From respiration data based on CO2 uptake by NaOH and corresponding root biomass values for each run of 10 plots, a linear regression of CO2 evolution rates on root dry weights has been achieved for every ten days. By applying the approach of extrapolating the regressive line to zero rootbiomass, the proportion of the total soil respiration flux that is attributableto live root respiration was estimated to be about 27% on average, ranging from 14% to 39% in the growing season in 1998. There were no evident relations between the total canopy biomass or root biomass and CO2 evolution rates, but a significant exponential relation did exist between the live canopy biomass and CO2 evolution rates.
李凌浩 韩兴国 王其兵 陈全胜 张焱 杨晶 闫志丹 李鑫 白文明 宋世环
摘要：采用根系生物量梯度上土壤呼吸变化趋势线外推法对锡林河流域一个羊草(Leymus chinensis (Trin.) Tzvel.)群落中根系呼吸占土壤总呼吸的比例进行了估计,对生物量各组分(地上、地下部分)之间以及它们与土壤呼吸间的相关性进行了分析.结果表明:在测定年度(1998年)整个生长季的不同月份,该群落中根系呼吸量占土壤呼吸总量的比例在14%～39%之间,平均为27%;地上总生物量及根系生物量与土壤呼吸间的相关性较差,但地上活生物量与土壤呼吸间存在着显著的乘幂关系.上述结果与国外同类研究结果相比,具有很好的一致性.
Taxol production of Taxus chinensis（Pilger） Rehd. var.mairei (Lemeeet Lévl.) Cheng et L. K. Fu induced by oligosaccharide from Fusarium oxysporum f.vasinfectum (Atkinson) Snyder et Hansen was studied in suspension cultures, and it was found that oligosaccharide triggered cell apoptosis. Under transmission electron microscope the following morphological changes were observed: cell shrinkage, condensation of cytoplasm, nuclear fragmentation, and the increase of high electron density bodies in vacuole in great quantity. In oligosaccharide treated cells, agarose gel electrophoresis revealed that DNA was digested into oligonucleosomal fragments that were times of 200 bp appearing as DNA ladders. Control cells were in normal physiological state, they were intact, abundant in organelle and with integral nucleus DNA, and the rate of taxol biosynthesis in these cells was very low. After the oligosaccharide to the culture system, the defense system of cells was elicited and the secondary metabolism was strengthened, i.e. phenolics were accumulated in the medium, the activity of polyphenol oxidase (PPO) was increased quickly and secondary wall of cells was thickened. The activity of L phenylalanine ammonia lyase (PAL), the critical enzyme of the phenylpropanoid pathway, was increased promptly 1 h after elicitation. The rate of taxol production was improved sharply and the maximal taxol concentration at 72 h was six times that of control. Appearance of cell apoptosis was accompanied with the highest concentration of taxol in suspension cultures.
李 春1,2 元英进1 马忠海1 胡宗定1 孙安慈3 胡昌序3
（1. 天津大学化工学院制药工程系，天津300072；2. ( 清华大学化工系生物化工研究所，北京100084;3.( 中国科学院植物研究所，北京100093）
摘要：在真菌(Fusarium oxysporum f.vasinfectum (Atkinson) Snyder et Hansen)寡聚糖诱导悬浮培养南方红豆杉(Taxus chinensis (Pilger) Rehd.var.mairei (Lemee et Lévl.) Cheng et L.K.Fu)细胞生产紫杉醇的体系中发现细胞出现凋亡,次生代谢增强.电镜观察到细胞核质和原生质出现凝集现象,液泡内出现大量的高电子致密体.核DNA经琼脂糖凝胶电泳,呈200 bp的整数倍的梯状条带(ladders);而对照组细胞核DNA完整,呈大片段,细胞完整,细胞器发达,但紫杉醇合成速率很低.加入寡聚糖后,细胞防御系统开启,细胞生长停止,次生代谢物酚类物质大量积累且次生壁加厚,多酚氧化酶活性迅速提高,苯丙烷类代谢途径的关键酶苯丙氨酸解氨酶的活性在1 h后急速提高,目的产物紫杉醇在诱导后72 h达到峰值,比对照组提高了6倍,且细胞凋亡的出现与紫杉醇合成的峰值具有时间上的一致性.
Suspension cell cultures of Maytenus hookeri Loes. (Celastraceae) in SH media were established from the calli induced from the leaves and young stems of M. hookeri on MS media with the supplement of 2 mg/L 2,4-D and 0.1 mg/L KIN (kinetin). Ethyl acetate extract of the cultures showed inhibitory activities against Penicillium avellaneum UC-4376 which was sensitive to maytansinoids. Exhaustive isolation of natural products from a large scale of suspension cell cultures did not yield maytansine instead of affording nine compounds including one novel triterp enoid, named 2,3-diacetoxyl maytenusone (1), and eight known ones including squalene (2), β-sitosterol (3), 2′,3′,4′-triacetyl-sitoindoside Ⅰ (4), salaspermic acid (5), maytenonic acid (6), 2α-hydroxy-maytenonic acid (7), 6,11,12-trihydroxy 8,11,13-abietrien-7-one (8) and 11,12 dihydroxy 8,11,13 abietatrien 7 one (9) elucidated on the basis of 1D and 2D-NMR data. The 1H-NMR and 13C-NMR assignments were made for 1, 5, 6 and 7, while the 13C-NMR assignments for 5 and 6 were revised. The chemical results suggested that the suspension cell cultures of M. hookeri did not produce maytansinoids under the reported experiment conditions.
鲁春华1 张建新2 甘烦远1 沈月毛1，2
（1. 中国科学院昆明植物研究所，昆明650204；2. 贵州省、中国科学院天然产物化学重点实验室，贵阳550002）
利用组织培养方法以云南美登木(Maytenus hookeri Lose.)未木质化的茎和嫩叶为材料诱导出愈伤组织,经过继代培养建立了悬浮细胞培养系.培养物的乙酸乙酯提取物显示抗橙色青霉(Penicillium avellaneum UC-4376)生长的生物活性.对培养物进行化学成分研究,分离鉴定了9个化合物:2,3-diacetoxyl maytenusone (1)、角鲨烯 (squalene,2)、β-谷甾醇 (β-sitosterol,3)、2′,3′,4′-triacetylsitoindoside Ⅰ (4)、salaspermic acid (5)、美登酮酸 (maytenonic acid,6)、2α-羟基美登酮酸(2α-hydroxy-maytenonic acid,7)、6,11,12-trihydroxy-8,11,13-abietrien-7-one (8)和11,12-dihydroxy-8,11,13-abietrien-7-one (9),其中化合物1为新化合物.通过2D NMR对化合物5-7的NMR数据进行了全指定,并修正了化合物5和6的部分碳谱数据指定.
关键词： 云南美登木；卫矛科；悬浮细胞培养；美登木素2，3-diacetoxyl maytenusone
通讯作者。E-mail:<yshen@ public.km.yn.cn>;Tel.:086-871-5219300;Fax: 086-871-5150227.
Vegetation changes and human influences on natural environments since 5 000 aBP were discussed based on the pollen data supported by radiocarbon dating from Cangumiao and Yujiawa profiles in Qian'an Basin, the southern area of Yanshan Mountain. The research indicates that (1) from 4 600 to 3 100 aBP, deciduous broad leaved forests dominated by Quercus grew in the Basin, (2) after 3 100 aBP, broad leaved species reduced and Pinus increased, in response to the cooling climate, (3) since 1 600 aBP, forests had been destroyed seriously due to the increase of agricultural activities, and (4) pollen analysis showed that Castanea trees had been cultivated for 1 600 years, and Oryzahad been planted for 700 years in Qian’an Basin.
许清海1,2孔昭宸1*阳小兰2 梁文栋3 孙黎明4
（1. 中国科学院植物研究所，北京100093:2. 河北省科学院地理研究所，石家庄050011；
3. 中国科学院地质与地球物理研究所，北京100029；4. 石家庄经济学院，石家庄050031）
摘要： 依据位于燕山南部迁安盆地蚕姑庙村和余家洼村具放射性碳年代测定支持的埋藏泥炭剖面的孢粉分析资料 ,揭示了迁安盆地 5 0 0 0aBP以来自然植被变化和人类活动对自然环境的影响。在 4 6 0 0～ 310 0aBP期间 ,迁安盆地曾经生长着以栎为主的落叶阔叶林 ,至 310 0aBP以后 ,森林中阔叶树成分显著下降 ,喜凉耐干的松树比例增加 ,16 0 0aBP后 ,迁安盆地人类农耕活动进一步加剧 ,森林遭受到大规模的破坏。孢粉分析资料揭示迁安盆地人类栽培板栗的历史可以追溯到 16 0 0aBP左右 ,种植水稻的历史可以追溯到 70 0aBP左右
Black bedded, nodular, and lenticular chert and silicified parts of stromatolites from the approximately 1 400-1 600 million year old Gaoyuzhuang Formation, North China, contain remarkably well preserved microflora composed of diverse prokaryotic and possible eukaryotic microbes. Macroscopic algae have also been found. This Formation provides important paleobiological information from which to assess status of Mesoproterozoic life, and to evaluate paleoenvironmental conditions and pre Phanerozoic biostratigraphy. The Gaoyuzhuang Formation has yielded over 100 taxa described by over 10 investigators. Most of the microbial taxa can be divided into three different assemblages according to their paleobiological and paleoenvironmental characteristics.1.The oscillatoriacean/chroococcacean assemblage which occurs in the first member of the Formation and this is interpreted to represent an assemblage from shallow water mat building and mat dwelling taxa with some possible planktonic taxa; 2. the nostocacean assemblage found only in the second member, and includes benthic mat building or mat dwelling taxa; and 3. the chroococcacean assemblage of the fourth member, which represents intertidal to supratidal mat building and allochthonous taxa.Proterozoic microfossils found in chert, especially cyanobacteria, are possibly more valuable as paleoenvironmental indicators than as chronostratigraphic tools because of their morphological conservation, although trends in diversity appear to exist for cyanobacteria in the Proterozoic. The general characteristics of Gaoyuzhuang microfossils, and the fact that most nostocacean chains occur and are preserved with the chain perpendicular to bedding indicate that: (1) the individual size of Gaoyuzhuang microfossils are part of a general trend of increase with time; (2) Gaoyuzhuang deposits are possibly in a fresh water environment and the rate of sedimentation might be equal to or less than the microbial growth rate; (3) the sedimentary model of Gaoyuzhuang might be from intertidal to subtidal or deep sea, and then to intertidal or supratidal; and (4) the eukaryotic organisms had densely occurred when Gaoyuzhuang Formation deposited.
（1. 中国科学院植物研究所，北京100093；2. 美国加州大学圣巴巴拉分校地质系克劳德研究室，CA93106）
摘要： 距今14～16亿年的华北地区高于庄组黑色层状、结核状和透镜状燧石与叠层石的黑色硅质部分中保存着极丰富的原核和真核生物微化石.宏观藻类在该组的页岩中亦已发现.本组为评估中元古代生命状况、古环境和前显生宙生物地层提供了重要的生物信息.迄今为止在高于庄组地层中已有百余个化石种被人们认识.根据古植物和古环境的特征这些生物种类可分为三个不同的组合,即:1.颤藻和色球藻组合,出现在高于庄组一段,代表浅水藻席建造者和居住者与一些可能的外来浮游生物的种类;2.念珠藻组合,仅发生在该组的二段,还含一些底栖藻席建造者或居住者的种类;3.第四段的色球藻组合,它代表了潮间带至亚潮带的藻席建造者和外来的种类.元古代燧石中的微化石,尤其是蓝藻化石,尽管在元古代它们就已不断趋向于多样化,但由于它们形态上的保守性,对环境的指示比对地质时代的指示更有价值.高于庄组微化石的特点和大多数链状念珠藻垂直层理保存的事实表明:1.高于庄组微化石的个体大小随时间趋向于增大;2.高于庄组的沉积可能是处于一个淡水环境,且沉积率可能等于或少于微生物的生长率;3. 高于庄组织沉积模式可能是从潮间带至亚潮带或深海,然后再至潮间带或潮上带;4.当高于庄组沉积时真核生物亦已出现.
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