FMDV vp1基因在豆科牧草百脉根中的转化与表达

通过根癌农杆菌介导法,将FMDV阿克苏(Akesu/O/58)株结构基因vp1转化豆科牧草百脉根子叶和子叶柄,其愈伤、芽和生根等过程经50 mg/L Kan筛选后,获得Kan抗性百脉根植株。对抗性植株进行vp1基因的PCR、RT-PCR检测和VP1蛋白的Western-blotting杂交。结果表明:vp1基因转入百脉根中,检测有转录活性;目的蛋白获得了正确表达;扩繁和移栽后获得了批量转基因百脉根,为下一阶段的动物试验提供了实验材料。     

FMDV vp1基因在豆科牧草百脉根中的转化与表达

通过根癌农杆菌介导法,将FMDV阿克苏(Akesu/O/58)株结构基因vp1转化豆科牧草百脉根子叶和子叶柄,其愈伤、芽和生根等过程经50 mg/L Kan筛选后,获得Kan抗性百脉根植株.对抗性植株进行vp1基因的PCR、RT-PCR检测和VP1蛋白的Western-blotting杂交.结果表明vp1基因转入百脉根中,检测有转录活性;目的蛋白获得了正确表达;扩繁和移栽后获得了批量转基因百脉根,为下一阶段的动物试验提供了实验材料.     

农杆菌介导的植物基因转化研究进展

农杆菌介导的植物基因转佛当今植物基因转化的主要方法之一,因而深受关注,本文从农力介导的基因转化机理,植物对农杆菌侵染的反应,转基因植物的遗传表达,以及农杆菌对单子叶植物的转化等方面论述了该领域的最新研究进展,并提出了进一步研究的方向。     

耐高温α-淀粉酶基因在马铃薯中的表达

我们将从地衣芽孢杆菌(Bacilluslicheniformis)克隆的约1．68kb的耐高温α-淀粉酶基因构建成表达载体,并转入根癌农杆菌。以马铃薯栽培品种“杂交荷兰7号”块茎圆盘为外植体,按本实验室建立起的再生实验系统[9]及杨美珠等的方法[8]进行转化。采取共培养、芽的诱导、芽的选择再生三步方法获得抗性芽。将抗性芽通过先诱导生根壮苗,再进行卡那霉素筛选,最后再诱导生根的方法得到可能的转基因植株。 对部分可能的转基因植株按改进的王广立等的PCR简单快速鉴定转基因植物的方法[12]进行检测,株号102001、102607、110402均可见到特异性片段的存在。参照张振清[13]及王福荣等[14]的方法对这些植株进行耐高温α—淀粉酶活力测定,这些植株具有相对较强的耐高温α—淀粉酶活性。实验结果表明,耐高温α—淀粉酶基因可能已转入上述植物基因组中,并获表达。     

苏云金芽孢杆菌(Bt)晶体毒蛋白基因在烟草叶绿体中的表达

将全长３．５ｋｂ的Ｂｔ基因３’端缺失,得到长为２．１ｋｂ、１．８ｋｂ的基因。分别将这３个长度（１．８ｋｂ、２．１ｋｂ、３．５ｋｂ）的基因置于水稻叶绿体ｐｓｂＡ基因的启动子和终止子调控之下,并与选择标记基因ａａｄＡ（编码氨基糖苷－３’－腺苷酸转移酶,具壮观霉素抗性）表达盒相连;以烟草叶绿体基因ｔｒｎＨ－ｐｓｂＡ－ｔｒｎＫ为同源片段,构建成叶绿体转化载体ｐＢＴ３、ｐＢＴ８和ｐＢＴ２２。用基因枪把Ｂｔ基     

缺失了控制肿瘤的功能之后,从烟草冠瘿瘤再生了能表达(鱼章)鱼碱合成酶基因的正常可育植株

对于植物遗传工程,用T-DNA作为载体的一个重要问题是需要再生正常的植株,这些植株要能够稳定地保持、表达和有性地传递被转移的DNA。     

多能硫杆菌RbisCO基因在大肠杆菌中表达产物分析

通过对多能硫杆菌ＲｕｂｉｓＣＯ的基因表达分析表明该基因能够在ｐＢＲ３２２的Ｐ１启动子、ｐｕＣ１９的ｌａｃ启动子以及ｐＫＫ２２３－３的ｔａｃ启动子的启动下,在大肠杆菌中表达,ＲｕｂｉｓＣＯ基因片段在ｐＵＣ１９和ｐＫＫ２２３－３载体上的表达活性较高。进一步对ＲｕｂｉｓＣＯ基因表达产物进行了非变性聚丙烯酰胺凝胶电泳,检测到了ＲｕｂｉｓＣＯ蛋白质带。     

mel基因在苏云金芽孢杆菌中的转移和表达

苏云金芽孢杆菌(Bacillus thuringhensis)是研究较深入、使用较广泛的杀虫细菌,野外使用易受阳光中紫外线的影响而失活。黑色素具有很强的抗辐射作用。将构建的含有嗜麦芽假单胞菌mel基因的质粒pWSY通过原生质体转化导入苏云金芽孢杆菌体内,使后获得了稳定产生黑色素的能力。并在转化子细胞内检测到与供体菌相同的质粒。SDS-PAGE显示该转化子体内额外表达了一分子量约为18kD的蛋白,该蛋白很可能就是转化子表达的酪氨酸酶。经测定,转化子抗辐射作用明显增强,有效杀虫时间显延长。     

高温乳糖酶基因在大肠杆菌中的高效表达

来源于嗜热脂肪芽孢杆菌（Bacillus stearothermophilus)的β-半乳糖苷酶基因bgaB经克隆,测序后,转入大肠杆菌高效表达载体pET-20(b)中,重组菌在IPTG诱导下,表达出的重组蛋白比酶活量为6.66U/mg。比出发菌株高50倍。     

Influence of root crown temperature on plant development

Summary Temperature effects on the root crown section, where most of the plant fluid translocation streams through a narrow space, were investigated. Cooled root crown section of tomato seedlings during daytime increased root weight; high temperatures enhanced shoot development. High and low temperatures day and night, on root crown, reduced root and shoot development. Wilting effects of shoots due to high temperature root environment, were delayed greatly by cooled root crown.The root crown section may, under certain temperature conditions, be the bottleneck of plant development, as it controls the up and down flow of water, nutrients, assimilated products, and hormones.     

Expression of effector gene SIX1 of Fusarium oxysporum requires living plant cells

Fusarium oxysporum is an asexual, soil inhabiting fungus that comprises many different formae speciales, each pathogenic towards a different host plant. In absence of a suitable host all F. oxysporum isolates appear to have a very similar lifestyle, feeding on plant debris and colonizing the rhizosphere of living plants. Upon infection F. oxysporum switches from a saprophytic to an infectious lifestyle, which probably includes the reprogramming of gene expression. In this work we show that the expression of the known effector gene SIX1 of F. oxysporum f. sp. lycopersici is strongly upregulated during colonization of the host plant. Using GFP (green fluorescent protein) as reporter, we show that induction of SIX1 expression starts immediately upon penetration of the root cortex. Induction requires living plant cells, but is not host specific and does not depend on morphological features of roots, since plant cells in culture can also induce SIX1 expression. Taken together, F. oxysporum seems to be able to distinguish between living and dead plant material, preventing unnecessary switches from a saprophytic to an infectious lifestyle.     

Expression of nopaline synthesis in tumorous and regenerant tobacco crown galls

Summary Tissues formed in liquid cultures of tobacco (Nicotiana tabacum cv. Wisconsin 38) crown galls incited byAgrobacterium tumefaciens C58 were of three types: unorganized callus, organized teratoma, and organized normal appearing. These tissues contained 400±12, 410±17, and 614±53 μg nopaline/g fresh weight, respectively. Using [¹⁴C]arginine, methods were developed for measuring in vivo nopaline biosynthetic rates. Tissues were incubated in a low concentration (i.e., 3 μM) of [¹⁴C]arginine to minimize disruption of the internal pool (approximately 140 μM free arginine). Radioactivity in the tissue was assayed and the specific radioactivity of free arginine, the precursor of nopaline, was determined. The linear rate of incorporation of radioactivity into nopaline was used to calculate the following biosynthetic rates (expressed as microgram nopaline per gram fresh weight per 24 h): callus, 14; teratoma, 21; normal appearing, 24. These results show conclusively that normal appearing tissues obtained from crown gall tumors can synthesize nopaline. Abnormal growth and opine biosynthesis, therfore, can be expressed independently.     

Expression of a C3 plant Rubisco SSU gene in regenerated C4 Flaveria plants

We report the successful transformation, via Agrobacterium tumefaciens infection, and regeneration of two species of the genus Flaveria: F. brownii and F. palmeri. We document the expression of a C₃ plant gene, an abundantly expressed ribulose 1,5-bisphosphate carboxylase/oxygenase small subunit gene isolated from petunia, in these C₄ plants. The organ-specific expression of this petunia gene in Flaveria brownii is qualitatively identical to its endogenous pattern of expression.     

山西濒危植物翅果油树植冠的构型分析

应用分形理论及方法,从分枝格局和冠幅扩展2个方面对山西翼城和乡宁的35株翅果油树(Elaeagnus mollis Diels)个体的植冠构型进行统计分析。结果表明,幼树和成树的总体分枝率和逐步分枝率有显著变化;一级枝的平均枝长和枝径有显著差异,但枝和叶的方位角及叶倾角差异不明显。不同发育阶段个体的冠幅变化较大,幼树冠幅的分形维数(2.0026)小于成树(2.2694)。翅果油树在不同生长发育阶段对生境变化有不同的构型策略。     

ACC deaminase from plant growth-promoting bacteria affects crown gall development

In addition to the well-known roles of indoleacetic acid and cytokinin in crown gall formation, the plant hormone ethylene also plays an important role in this process. Many plant growth-promoting bacteria (PGPB) encode the enzyme 1-aminocyclopropane-1-carboxylate (ACC) deaminase, which can degrade ACC, the immediate precursor of ethylene in plants, to alpha-ketobutyrate and ammonia and thereby lower plant ethylene levels. To study the effect of ACC deaminase on crown gall development, an ACC deaminase gene from the PGPB Pseudomonas putida UW4 was introduced into Agrobacterium tumefaciens C58, so that the effect of ACC deaminase activity on tumour formation in tomato and castor bean plants could be assessed. Plants were also coinoculated with A. tumefaciens C58 and P. putida UW4 or P. putida UW4-acdS- (an ACC deaminase minus mutant strain). In both types of experiments, it was observed that the presence of ACC deaminase generally inhibited tumour development on both tomato and castor bean plants.     

Expression of the bacterial gdhA gene encoding a NADPH glutamate dehydrogenase in tobacco affects plant growth and development

We investigated the effects of genetic modification of nitrogen metabolism via the bacterial glutamate dehydrogenase (GDH) on plant growth and metabolism. The gdhA gene from Escherichia coli encoding a NADPH-GDH was expressed in tobacco plants under the control of the 35 S promoter. The specific activity of GDH in gdhA plants was 8-fold of that in E. coli. Damage caused by spray application of 1.35 mM of phosphinothricin (PPT) herbicide, a glutamine synthetase (GS) inhibitor, was less pronounced in gdhA plants as compared with the control plants which suggests that the introduced GDH can assimilate some of the excess ammonium, at least during GS inhibition. However, gdhA plants were susceptible to 2.7 mM PPT. Biomass production was consistently increased in gdhA transgenic plants grown under controlled conditions and in the field. Total free amino acids and total carbohydrates were increased in gdhA plants grown in the greenhouse suggesting that both nitrogen and carbon metabolism were altered. We conclude that the modifications in transgenic plants may result from both increased nitrogen efficiency and altered gene expression and metabolism. This revised version was published online in June 2006 with corrections to the Cover Date.