OsPHGPx基因的过量表达增强水稻抗氧化能力

植物在生物或非生物胁迫下会通过表达磷脂氢谷胱甘肽过氧化物酶(PHGPx)来抵御胁迫引起的氧化损伤,但是PHGPx在植物体内抗氧化途径中所扮演的生理角色目前尚不完全清楚。利用农杆菌遗传转化技术,构建了过表达Os PHGPx基因的转基因水稻,并对转基因水稻进行了PCR、实时定量PCR以及Western blot等检测分析,结果表明Os PHGPx基因已成功转入水稻并正常表达。与野生型水稻相比,这些过量表达Os PHGPx的转基因水稻抵御百草枯氧化伤害的能力提高。     

Photosynthetic Characteristics of Transgenic Rice Plants Overexpressing Maize Phosphoenopyruvate Carboxylase

转PEPC基因水稻的光合生理特性 迟伟¹焦德茂^1* 黄雪清¹ 李霞¹ 匡廷云² Ku S..B.MAURICE ³     

过量表达大叶补血草LgNHX1基因对拟南芥耐盐性的影响

利用植物表达载体pCAMBIA1301和农杆菌GV3101将LgNHX1(全长1 656 bp)基因在拟南芥中过量表达.在含30 mg/L潮霉素的培养基上筛选获得LgNHX1的纯合转化子,并对其进行了分子鉴定和耐盐性分析.结果显示,经PCR和RT-PCR鉴定,野生型植株(对照)没有出现扩增条带,而转基因株系有相应的扩增条带,表明LgNHX1的确已经整合到拟南芥的基因组中,并已正常转录.在不同盐浓度处理下,转基因株系生长情况好于野生型对照;转基因植株地上部分和根的干重、鲜重相对高于野生型对照,但差异没有达到显著水平;当盐浓度达到150-200 mmol/L时,两个特基因株系的Na+含量显著高于野生型,K+含量极显著高于野生型.以上结果表明,过量表达LgNHX1基因可能增强了拟南芥将Na+区隔化至液泡的能力,提高了转基因拟南芥的耐盐能力.     

Properties of a Protein Activator of NAD Kinase from Plants

Purification of pea (Pisum sativum) seedling NAD kinase by DEAE-cellulose column chromatography resulted in loss of activity, due to dissociation of an activator from the enzyme. The purified enzyme preparation, which was almost completely inactive, regained the activity when the activator was added back.     

转移拟南芥CBF1基因引起水稻植株脯氨酸含量提高

利用农杆菌介导的转基因技术,成功地将拟南芥抗冻转录激活因子基因CBF1转入粳稻中花11中,并获得了T1代转基因植株。CBF1基因及筛选基因HPT（潮霉素抗性基因)均在T1代中检测到,呈现单位点的孟德尔式遗传。常温和低温处理之后,T1代植株体内的脯氨酸含量均比野生型明显提高,同时,耐低温表型也在T1-1株系中出现。     

新基因水稻OsLSD1的克隆及拟南芥和水稻类 LSD1基因家族的生物信息学分析

类LSD1 (LSD1-like)基因家族是一类特殊的C2C2型锌指蛋白基因,编码植物特有的转录因子.目前已经研究的2个成员拟南芥LSD1(1esions stimulating disease resistance 1)和LOL1(LSD-One-Like 1)基因均参与植物细胞程序化死亡(programmed cell death,PCD)的调控.从水稻cDNA文库中克隆到1个类LSD1基因,命名为OsLSD1.该基因长988 bp,包含一个432bp的开放阅读框,推导的氨基酸序列(143个氨基酸)含有3个内部保守的锌指结构域.DNA印迹结果表明OsLSD1基因在水稻基因组中为单拷贝,且在根、茎和叶中表达.借助于生物信息学分析技术,从拟南芥和水稻数据库中各识别出5个和7个(包括OsLSD1)类LSD1基因.分析了这些类LSD1基因的结构,蛋白质结构域组成.系统进化分析表明,无论基于编码区的核苷酸或氨基酸序列都可以将这些类LSD1基因分为2类.虽然不存在拟南芥或水稻特有的类LSD1蛋白,但有些结构域是水稻所特有的,也有些基因是来源于复制事件.     

Positively Selected Sites in the Arabidopsis Receptor-Like Kinase Gene Family

We analyze members of the receptor-like kinase (RLK) gene family in Arabidopsis thaliana for positive selection. Likelihood analyses find evidence for positive selection in 12 of the 52 RLK family sequences groups. These 12 groups represent 97 of the 403 sequences analyzed. The majority of genes in groups subject to positive selection have not been functionally characterized, but sites under selection are predominantly located in the extracellular region. The pattern of selection in the extracellular leucine-rich repeat (LRR) motif of groups 14 and 51 is similar to previous studies where positively selected positions are located in a solvent exposed β-strand that may determine disease specificity, raising the possibility that some RLK genes function in a similar role.[Reviewing Editor: Rasmus Nielsen]     

Gene Structure Analysis of Rice ADP-ribosylation Factors (OsARFs) and Their mRNA Expression in Developing Rice Plants

ADP-ribosylation factors (ARFs) have been widely identified as being involved in regulation of cell differentiation, intracellular vesicle transport, and cytoskeleton organization in animals and plants. To systematically clarify the ARF gene family in rice, eight putative rice ARF genes (OsARFs) were detected by searching the rice genome database. The OsARFs can be classified into class I (OsARFA1a-OsARFA1e) and class II (OsARFB1a-OsARFB1c) by comparing their deduced amino acid sequences with other known ARFs. Most OsARFs are composed of six exons and five introns, though OsARFB1b has three exons and two introns. The OsARFs are usually expressed in various developing organs, whereas OsARFA1a is predominantly expressed in young roots and in caryopses at early development stages. The histochemical localization of OsARFs in various developing organs was confirmed in young leaves, the pericycle of young roots, and the aleurone layer of developing caryopses. Furthermore, the expression of OsARFA1a was confirmed specifically in the aleurone layer and immature embryos of developing caryopses. The present work provides a foundation for further clarifying the physiological functions of OsARFs in rice growth and development.     

Characterization of Transgenic Rice Plants Expressing an Arabidopsis FAD7

Fatty acid -3 desaturase (FAD) is the key enzyme catalyzing the formation of trienoic fatty acids. We utilized an Arabidopsis FAD7 gene and the seven independent transgenic rice plants harbouring 1 to 3 copies of this gene were generated. The expression of FAD7 mRNA was different among independent transgenic lines regardless of the copy number. The total linolenic acid (18:3) contents reduced by about 7 – 32 % in transgenic rice plants but the linoleic acid (18:2) content increased accordingly. With or without wounding treatments, the jasmonate content was higher in transgenic lines than in wild-type rice plant. The transgenic lines overproducing jasmonate also showed increased expression of PR1b mRNA and allene oxide synthase inresponse to wounding.     

Expression of Rice Stripe Virus Coat Protein Gene in Transgenic Rice Plants

Rice stripe virus (RSV) is a pathogen of rice stripe disease causing great damage to rice. The disease is transmitted by Laodelphax striatellus and three other planthoppers. RSV infects as much as 37 cereals including rice, wheat, maize and results in a significant reduction in yield in epidemic year. In order to develop efficient means of controlling the disease, authors have studied the amino acid composition of RSV coat protein (CP), synthesized and cloned the cDNA to CP, sequenced the full-length CP gene. Having inserted the RSV CP gene into plant expression vector pROK Ⅱ, authors transformed rice suspension culture via microprojectile bombardment and obtained transgenic plants expressing the CP gene. The suspension culture was initiated by inoculating yellowish, compact and embryogenic calli derived from seeds into suspension medium containing proline and maltose. After being cultured at 26℃ in the dark for about half a year, finely-dispersed and embryogenic suspension culture was estabolished. Before bombardment the suspension culture was evently applied onto three-layered filter-paper discs in a petri dish. CaCl2 and spermidine was employed to coat tungsten particle with plasmid DNA. 2.5 μl of coated particle was loaded onto bullet and each dish was bombarded three times. Immediately after being bombarded, the suspensions were cultured in modified N6 medium. 2 days later the suspensions were transferred to the same medium but containing G418, which were subcultured weekly. Being subject to G418 selection for two months, white and fast-growing clones were emerged from the brownish cultures. Green plants regenerated when the resistant calli were transferred to differentiation medium. The regenerated plants were firm enough to grow well in the greenhouse. 10 plants regenerated from G418 resistant calli were tested for their transformed nature by Southern blot using 32P-labelled CP gene as a probe. Among the plants tested, 2 plants showed clearly hy bridizing bands with a molecular weight corresponding to RSV CP gene. Western blot further demonstrated that RSV CP gene was expressed in transgenic rice plants. At present tests on the antiviral effects of transgenic plants by feeding plantphoppers infccted with RSV are being underway.     

Transgenic Tobacco Plants Overexpressing a Grass PpEXP1 Gene Exhibit Enhanced Tolerance to Heat Stress

Heat stress is a detrimental abiotic stress limiting the growth of many plant species and is associated with various cellular and physiological damages. Expansins are a family of proteins which are known to play roles in regulating cell wall elongation and expansion, as well as other growth and developmental processes. The in vitro roles of expansins regulating plant heat tolerance are not well understood. The objectives of this study were to isolate and clone an expansin gene in a perennial grass species (Poa pratensis) and to determine whether over-expression of expansin may improve plant heat tolerance. Tobacco (Nicotiana tabacum) was used as the model plant for gene transformation and an expansin gene PpEXP1 from Poa pratensis was cloned. Sequence analysis showed PpEXP1 belonged to α-expansins and was closely related to two expansin genes in other perennial grass species (Festuca pratensis and Agrostis stolonifera) as well as Triticum aestivum, Oryza sativa, and Brachypodium distachyon. Transgenic tobacco plants over-expressing PpEXP1 were generated through Agrobacterium-mediated transformation. Under heat stress (42°C) in growth chambers, transgenic tobacco plants over-expressing the PpEXP1 gene exhibited a less structural damage to cells, lower electrolyte leakage, lower levels of membrane lipid peroxidation, and lower content of hydrogen peroxide, as well as higher chlorophyll content, net photosynthetic rate, relative water content, activity of antioxidant enzyme, and seed germination rates, compared to the wild-type plants. These results demonstrated the positive roles of PpEXP1 in enhancing plant tolerance to heat stress and the possibility of using expansins for genetic modification of cool-season perennial grasses in the development of heat-tolerant germplasm and cultivars.