香蕉乙烯受体基因cDNA的克隆及其表达分析

提取香蕉果实总RNA，根据有关文献报道的乙烯受体基因序列设计引物，通过RT-PCR方法扩增其开放读框(open reading frame，ORF)，结果同时扩增出2个长短不同的特异cDNA序列。测序结果表明：其中较长的cDNA序列与该报道的香蕉乙烯受体cDNA序列的对应区段(ORF)长度一致，同源性为99．1％；另一较短cDNA序列为新序列，其与该报道序列的同源性达97．2％，但缺少该报道序列中的19和1036 bp的区段。RT-PCR扩增结果显示，报道的cDNA序列在香蕉果实的几个成熟阶段均有表达，而在根和叶片等组织中检测不到其表达，说明其表达具有果实组织特异性。Southern杂交结果显示，该报道基因序列在香蕉基因组中为单拷贝存在。因此，推测新克隆的缺失cDNA序列可能来自同一基因的转录后剪辑，其可能为1个新的香蕉乙烯受体基因。     

香蕉果实的1个14-3-3蛋白同源基因的克隆及序列分析(简报)

根据在香蕉果实抑制缩减文库中获得的1个14-3-3蛋白基因片段,采用PCR和RACE相结合的方法从香蕉(Musa acuminate L.AAA group cv.Brazilian)cDNA文库中筛选到其全长cDNA序列.序列测定和Blastx比对分析结果表明,该cDNA全长1 037 bp,含有1个完整的阅读框,其编码区编码261个氨基酸残基,具有植物14-3-3蛋白基因的保守结构域,并与其他植物来源的14-3-3蛋白具有很高的同源性,将其命名为Ma-14-3-36(Musa acuminate 14-3-3).采用遗传进化系统发育树的分析结果表明,Ma-14-3-36与来源于单子叶植物的多数14-3-3蛋白基因序列在同一个进化枝上.采用RT-PCR对其在香蕉果实发育不同时期的表达进行分析结果表明,在香蕉果实发育的不同时期差异表达,推测其可能在果实的发育中起作用.     

香蕉NPR1基因的克隆及植物表达载体构建

病程相关基因非表达子(NPR1)是植物抗病信号传导的关键基因。以香蕉抗病香蕉品种为材料,根据NPR1基因序列在保守区设计引物,从香蕉cDNA中克隆获得NPR1基因片段,以RACE的方法克隆获得香蕉NPR1基因的全长,并命名为GCT119-NPR1。香蕉GCT119-NPR1基因最大阅读框(ORF)为1 707 bp。生物信息学分析结果表明,该基因含有ANK和BTB两个结构域,其中ANK结构域是NPR1基因中非常重要的结构域,在蛋白质相互作用中起着至关重要的作用。同源性分析表明,该基因与已克隆发表的NPR1基因氨基酸序列同源性为67%~82%。将其构建植物表达载体,为下一步转化工作奠定基础。     

一个新的大豆类受体蛋白激酶基因rlpk2的克隆及其结构特征

植物类受体蛋白激酶在细胞生长及对环境胁迫反应方面起着非常重要的作用.本文报告用RACE方法(快速扩增cDNA末端)扩增一个新的大豆类受体蛋白激酶基因rlpk2的全长cDNA.根据RACE的结果,我们克隆了rlpk2的cDNA,其序列信息已经提交到GenBank(Accession No.AY687391).经初步分析,rlpk2编码产物为LRR型类受体蛋白激酶.     

香蕉MuMADS1与泛素激活酶(MuUBA)在采后果实中的相互作用

以香蕉MuMADS1为诱饵载体,采后2 d的香蕉果实的cDNA文库为猎物,采用酵母双杂交的方法得到了泛素激活酶E1的基因片段,命名为MuUBA。采用双分子荧光互补技术进一步验证MuMADS1与MuUBA在植物体内的相互作用。荧光实时定量PCR结果表明,MuMADS1和MuUBA在香蕉中子房发育第4个阶段的表达量最高,但是在茎中的表达量很低,表明其在不同的组织和发育的果实中的表达具有协同性。MuMADS1和MuUBA的表达都受外源乙烯和1-MCP的高度调控,推测MuMADS1和MuUBA在香蕉果实的采后成熟过程中具有很重要的作用。     

香蕉苯丙氨酸解氨酶基因家族的全基因组鉴定及表达分析

苯丙氨酸解氨酶（Phenylalanine ammonia-lyase, PAL）是催化苯丙烷代谢途径第1步反应的限速酶,广泛地参与植物生长发育过程中的各种生理活动。本研究通过生物信息学分析,从香蕉A基因组数据库中利用BLASTp筛选出53个可能的MaPAL编码蛋白序列。在NCBI分析蛋白保守结构域,发现仅有8个基因具有典型的PAL家族基因的特性。聚类分析结果表明香蕉MaPAL家族的8个基因可以分为Class I 和Class II两类。转录组数据表明,所有MaPAL基因在果实发育阶段高表达,MaPAL5参与果实采后成熟过程。在干旱、低温、盐处理的香蕉幼苗叶片中,MaPAL1、MaPAL2、MaPAL3、MaPAL4和MaPAL5成员均显著上调表达,MaPAL6号成员显著下调表达,这些成员参与香蕉响应上述非生物胁迫过程。8个MaPAL成员在枯萎病菌处理下均显著下调表达,表明在感病品种巴西蕉根系中,MaPAL基因的表达全部被Foc TR4抑制。本研究结果为分析MaPAL基因家族在香蕉果实品质形成和响应逆境中的作用提供了理论依据。     

大豆GmLecRlk基因耐盐功能分析

凝集素类受体蛋白激酶属于类受体蛋白激酶（RLKs）家族,在植物的抗病防御反应、生长发育、胞内信号传导以及非生物胁迫反应过程中发挥重要作用。实验室前期对过表达抗逆转录因子GmNFYB1大豆进行转录组测序,获得了差异表达基因GmLecRlk（Glyma.07G005700）,其开放阅读框长度为546 bp,编码181个氨基酸,蛋白结构域分析显示其含有两个丝氨酸/苏氨酸激酶结构域,属于一种G型凝集素类受体蛋白激酶。实时荧光定量PCR结果显示,GmLecRlk基因在大豆的根、茎、叶、荚中均有不同程度的表达,在根中表达量最高;200 mmol/LNaCl处理下,GmLecRlk 的mRNA丰度先降低后升高,在12 h时达到最高值,表明该基因参与大豆对盐胁迫的响应。利用发根农杆菌K599获得GmLecRlk过表达转基因发状根复合植株,在盐胁迫处理下,其存活率高于对照;在拟南芥中异源表达GmLecRlk基因,转基因拟南芥在盐胁迫处理下的萌发率、绿化率和根长均高于野生型拟南芥。综上所述,GmLecRlk参与大豆对盐胁迫的反应,过量表达基因能提高大豆和拟南芥的耐盐性,为培育和改良抗盐大豆新品种提供新的途径和理论指导。     

HXKs基因在香蕉果实后熟过程中功能初探

香蕉是世界主要水果之一,对乙烯非常敏感,属于呼吸跃变型果实。目前,香蕉果实后熟机理还未完全探明,后熟过程中己糖激酶(HXK)与乙烯的关系还不清楚。通过BLAST比对从香蕉基因组数据库中发现HXK基因家族的11个成员,经克隆获得这些基因的序列,并研究HXKs基因在香蕉果实自然后熟过程中的转录表达谱,重点研究乙烯利、甘露糖、NAG和1-MCP对HXKs基因表达、HXK酶活和内源乙烯生物合成的影响。结果表明:在香蕉果实后熟过程中HXKs基因呈现差异性表达,乙烯利上调大多数HXKs基因的表达和HXK酶活,1-MCP的作用正好相反,这表明外源乙烯正作用于HXK。另一方面,甘露糖加快内源乙烯生物合成,NAG却推迟内源乙烯生物合成,这说明HXK正作用于内源乙烯生物合成。所以,在香蕉果实后熟过程中乙烯和HXK之间存在相互促进的关系。这将为进一步阐明香蕉果实后熟机制提供理论依据,也将为香蕉果实保鲜新技术的挖掘提供新思路。     

香蕉MuMADS2基因表达产物的亚细胞定位

对已获得的香蕉MuMADS2基因进行生物信息学和芯片分析表明,该基因编码的蛋白可能作为转录因子定位于细胞核中,是乙烯上游的调控因子,参与调控香蕉果实的成熟过程。为进一步研究该基因功能,构建香蕉MuMADS2基因与绿色荧光蛋白(Green Fluorescent Protein,GFP)基因融合的植物表达载体pCAMBIA1302-MuMADS2,再利用基因枪转化法将其转入洋葱表皮细胞进行亚细胞定位观察。结果表明:该基因表达产物定位于细胞核中,符合转录因子特性。     

香蕉MaTET基因的克隆与表达分析

采用RACE技术克隆香蕉果实采后成熟相关基因MaTET的全长cDNA,利用生物信息学方法分析MaTET基因及推导蛋白的序列,再利用RT-PCR技术对该基因的组织器官表达特性和在果实采后成熟过程中的表达特性进行分析。结果表明,MaTET的cDNA序列全长为1 234 bp,包含一个852 bp的完整开放阅读框,编码283个氨基酸残基,5′端非编码区222 bp,3′端非编码区160 bp,起始密码子为ATG,终止密码子为TAG。MaTET蛋白具4个跨膜区域,属于四跨膜蛋白（Tetraspanins）,拥有多个与信号传导有关的修饰位点,在系统发生上更接近鹰嘴豆、野草莓和番茄等双子叶植物。MaTET可在香蕉根、茎、叶、花以及果实中表达,在根和叶中的表达量高于其它器官;MaTET在自然成熟香蕉果实中的微黄(TY)阶段开始上调表达,在黄多于绿(MY)阶段表达量急遽升高,随后降低,乙烯可增强MaTET表达且表达高峰提前至绿多于黄(MG)阶段,1-MCP抑制MaTET的表达。     

Studies on natural repellents against potato tuber moth (Phthorimaea operculella Zeller) in country stores

Summary The effects of the leaves of five plant species, one from each of the generaAmbrosia, Anemone, Eupatorium, Eucalyptus andLantana, on potato tuber moth were investigated under indigenous storage conditions at the Central Potato Research Station, Shillong (1800 m above sea level). Their action was compared with that of a biological insecticide (spores ofBacillus thuringiensis), a chemical insecticide (carbaryl), and an untreated control. The data collected after six months storage on tuber damage, sprout damage and the rotting indicated that the leaves ofLantana aculeata provided most protection to the tubers, reducing damage from over 70% in the check to below 5%, and sprout damage from over 45% to below 3%. Next best wasEucalyptus globulus followed byB. thuringiensis. They may be used on tubers stored for table use or for seed as they had no adverse effect on germination or on the yield of a subsequent crop.     

Comparative Assessment of the Efficacy of Various Nitrogen Fertilizers

Summary

The efficiency of N fertilizers is usually poor; often less than 50% of the applied N is taken up by the crop. This review focuses on various N fertilizers with respect to the significance of different N loss pathways, namely (i) ammonia volatilization, (ii) dinitrogen and nitrogen oxide emissions, and (iii) nitrate leaching. Further, the significance of biological N immobilization, ammonium fixation and, finally, the impact of nitrate vs. ammonium uptake on crop yield are also discussed. The reviewed literature shows that N fertilizers may differ markedly in their susceptibility to losses. There is, however, considerable scope to improve N efficiency of each N source by proper N management practices.     

In vitro antimicrobial activity of different accessions ofSolanum commersonii Dun. from Uruguay

Summary The in vitro antimicrobial activity of extracts from accessions ofSolanum commersonii Dun. collected in the south of Uruguay was investigated against five microorganisms including the pathogenRalstonia solanacearum. A total of 30 extracts corresponding to organic and aqueous extracts were studied. Interestingly, most of the positive results for growth inhibition were againstR. solanacearum. The extracts were also analyzed for the presence of glycoalkaloids and lectins. Six of the ten aqueous extracts showed lectin presence and a wide variation in the type and amounts of glycoalkaloids, was found. Results indicate that there is no clear relationship between the antimicrobial activity against the five microorganisms screened and the presence or amounts of lectins and glycoalkaloids, traditionally regarded as possible antimicrobial metabolites in theSolanum genus, which indicates the presence of as yet unidentified antimicrobial compounds.     

Novel food and non-food uses for sorghum and millets

Sorghum and millets have considerable potential in foods and beverages. As they are gluten-free they are suitable for coeliacs. Sorghum is also a potentially important source of nutraceuticals such antioxidant phenolics and cholesterol-lowering waxes. Cakes, cookies, pasta, a parboiled rice-like product and snack foods have been successfully produced from sorghum and, in some cases, millets. Wheat-free sorghum or millet bread remains the main challenge. Additives such as native and pre-gelatinised starches, hydrocolloids, fat, egg and rye pentosans improve bread quality. However, specific volumes are lower than those for wheat bread or gluten-free breads based on pure starches, and in many cases, breads tend to stale faster. Lager and stout beers with sorghum are brewed commercially. Sorghum's high-starch gelatinisation temperature and low beta-amylase activity remain problems with regard to complete substitution of barley malt with sorghum malt . The role of the sorghum endosperm matrix protein and cell wall components in limiting extract is a research focus. Brewing with millets is still at an experimental stage. Sorghum could be important for bioethanol and other bio-industrial products. Bioethanol research has focused on improving the economics of the process through cultivar selection, method development for low-quality grain and pre-processing to recover valuable by-products. Potential by-products such as the kafirin prolamin proteins and the pericarp wax have potential as bioplastic films and coatings for foods, primarily due to their hydrophobicity.     

Extreme resistance to infection by potato virus X in genotypes of wild tuber-bearingSolanum species

Summary Clones derived from thirty-one different accessions (nineteen of Argentine origin) belonging to eightSolanum species were screened for resistance to infection by potato virus X strain cp (PVX cp) by mechanical inoculation of plantlets that had been micropropagated in vitro. Estimates of PVX multiplication obtained by enzyme linked immunosorbent assay and slot blot nucleic acid hybridization allowed the identification of resistant clones derived from five accessions belonging toS. commersonii S. oplocense, S. sparsipilum andS. tuberosum andigena. Resistant genotypes supported PVX concentrations 5 to 15 times smaller than did the susceptible control cultivar Spunta. Graft inoculation test confirmed the presence of extreme resistance similar to that conferred by the ‘immunity’ gene X¹ (also called RX_act).     

Marine Pharmacology in 2009–2011: Marine Compounds with Antibacterial,Antidiabetic, Antifungal,Anti-Inflammatory,Antiprotozoal, Antituberculosis,and Antiviral Activities; Affecting the Immune and Nervous Systems,and other Miscellaneous Mechanisms of Action ?

The peer-reviewed marine pharmacology literature from 2009 to 2011 is presented in this review, following the format used in the 1998–2008 reviews of this series. The pharmacology of structurally-characterized compounds isolated from marine animals, algae, fungi and bacteria is discussed in a comprehensive manner. Antibacterial, antifungal, antiprotozoal, antituberculosis, and antiviral pharmacological activities were reported for 102 marine natural products. Additionally, 60 marine compounds were observed to affect the immune and nervous system as well as possess antidiabetic and anti-inflammatory effects. Finally, 68 marine metabolites were shown to interact with a variety of receptors and molecular targets, and thus will probably contribute to multiple pharmacological classes upon further mechanism of action studies. Marine pharmacology during 2009–2011 remained a global enterprise, with researchers from 35 countries, and the United States, contributing to the preclinical pharmacology of 262 marine compounds which are part of the preclinical pharmaceutical pipeline. Continued pharmacological research with marine natural products will contribute to enhance the marine pharmaceutical clinical pipeline, which in 2013 consisted of 17 marine natural products, analogs or derivatives targeting a limited number of disease categories.     

Recent Modifications of Chitosan for Adsorption Applications: A Critical and Systematic Review

Chitosan is considered to be one of the most promising and applicable materials in adsorption applications. The existence of amino and hydroxyl groups in its molecules contributes to many possible adsorption interactions between chitosan and pollutants (dyes, metals, ions, phenols, pharmaceuticals/drugs, pesticides, herbicides, etc.). These functional groups can help in establishing positions for modification. Based on the learning from previously published works in literature, researchers have achieved a modification of chitosan with a number of different functional groups. This work summarizes the published works of the last three years (2012–2014) regarding the modification reactions of chitosans (grafting, cross-linking, etc.) and their application to adsorption of different environmental pollutants (in liquid-phase).     

Pharmacokinetics of Marine-Derived Drugs

Marine organisms represent an excellent source of innovative compounds that have the potential for the development of new drugs. The pharmacokinetics of marine drugs has attracted increasing interest in recent decades due to its effective and potential contribution to the selection of rational dosage recommendations and the optimal use of the therapeutic arsenal. In general, pharmacokinetics studies how drugs change after administration via the processes of absorption, distribution, metabolism, and excretion (ADME). This review provides a summary of the pharmacokinetics studies of marine-derived active compounds, with a particular focus on their ADME. The pharmacokinetics of compounds derived from algae, crustaceans, sea cucumber, fungus, sea urchins, sponges, mollusks, tunicate, and bryozoan is discussed, and the pharmacokinetics data in human experiments are analyzed. In-depth characterization using pharmacokinetics is useful for obtaining information for understanding the molecular basis of pharmacological activity, for correct doses and treatment schemes selection, and for more effective drug application. Thus, an increase in pharmacokinetic research on marine-derived compounds is expected in the near future.