口蹄疫疫苗非抗原蛋白对146S抗原免疫效果的影响

为探究非抗原蛋白对口蹄疫病毒A型(FMDV-A) 146S抗原免疫效果的影响,本研究以小鼠和猪为试验动物,分别制备5组小鼠注射用样品:A组(3μg 146S抗原)、B组(3μg 146S抗原+25μg非抗原蛋白)、C组(3μg 146S抗原+50μg非抗原蛋白)、D组(3μg 146S抗原+100μg非抗原蛋白)及E组(空白对照,PBS);同时制备4组猪注射用样品:A组(18μg 146S抗原)、B组(18μg 146S抗原+400μg非抗原蛋白)、C(18μg 146S抗原+4 000μg非抗原蛋白)及D组(空白对照,PBS)。免疫接种试验动物后,通过淋巴细胞增殖试验及实时荧光定量PCR评价比较不同疫苗样品组小鼠产生的细胞免疫应答水平,同时通过液相阻断ELISA方法检测、评价各试验组动物(小鼠和猪)产生的体液免疫应答水平。小鼠试验结果显示,在免疫后3个检测时间点内,4个试验组中C组的平均抗体水平最高,平均抗体效价分别为4.13、5.83和5.50,而D组的抗体水平最低,平均抗体效价分别为3.46、5.16和4.46;C组细胞增殖能力及Th-1型细胞因子IL-6、IFN-β和TNF-αmRNA表达量均高于其他组。猪试验结果显示,A组和B组间平均抗体水平差异不显著(P>0.05),但A组和B组的平均抗体效价均极显著高于C组(P<0.01)。综合上述结果表明,一定范围内少量非抗原蛋白对FMDV-A 146S抗原的免疫效果没有影响,而高浓度的非抗原蛋白则抑制FMDV-A 146S抗原的免疫效果。     

Virulence status,viral accommodation and structural protein profiles of white spot syndrome virus isolates in farmed Penaeus monodon from the southeast coast of India

The objective of this study was to investigate the reason for variation in the virulence of white spot syndrome virus (WSSV) from different shrimp farms in the Southeast coast of India. Six isolates of WSSV from farms experiencing outbreaks (virulent WSSV; vWSSV) and three isolates of WSSV from farms that had infected shrimps but no outbreaks (non‐virulent WSSV; nvWSSV) were collected from different farms in the Southeast coast of India. The sampled animals were all positive for WSSV by first‐step PCR. The viral isolates were compared using histopathology, electron microscopy, SDS‐PAGE analysis of viral structural proteins, an in vivo infectivity experiment and sequence comparison of major structural protein VP28; there were no differences between isolates in these analyses. A significant observation was that the haemolymph protein profile of nvWSSV‐infected shrimps showed three extra polypeptide bands at 41, 33 and 24 kDa that were not found in the haemolymph protein profile of vWSSV‐infected shrimps. The data obtained in this study suggest that the observed difference in the virulence of WSSV may not be due to any change in the virus, rather it could be due to the shrimp defence system producing certain factors that help it to accommodate the virus without causing any mortality.     

Disulphide Bonds in Wheat Gluten Proteins

Disulphide bonds play a key role in determining the structure and properties of wheat gluten proteins. Comparison of the sequences of monomeric gliadins and polymeric glutenin subunits allows the identification of conserved and variant cysteine residues. Direct disulphide bond determination demonstrates that the conserved cysteine residues present in S-rich prolamins (α-type gliadins, γ-type gliadins and LMW subunits) form intra-chain disulphide bonds while additional cysteines residues present only in the LMW subunits form inter-chain bonds with cysteines in HMW subunits and other LMW subunits. Conserved and variant cysteine residues are also present in the HMW subunits but their patterns of disulphide bond formation are less well understood. Further information on the abilities of individual cysteine residues to form intra- and inter-chain disulphide bonds has also been obtained by heterologous expression of wild type and mutant proteins inE. coliand, in the case of the HMW subunits, by examination of the patterns of dimers recovered on partial reduction of glutenin or resulting from the expression of subunits in transgenic tobacco plants. Wheat gluten proteins are folded and assembled within the lumen of the endoplasmic reticulum of the developing endosperm cells, where disulphide bond formation and exchange may be catalysed by the enzyme protein disulphide isomerase. Similarly, disulphide bond reduction, for example to facilitate mobilisation during germination, may be catalysed by thioredoxinh. Understanding the mechanism and specificity of disulphide bond formation in gluten is crucial for the manipulation of its functional properties by genetic engineering or chemical modification.     

Identifying proteins with insecticidal activity: use of encoding genes to produce insect-resistant transgenic crops

Crops resistant to insect attack offer an alternative strategy of pest control to a total reliance upon chemical pesticides. Transgenic plant technology can be a useful tool in producing resistant crops, by introducing novel resistance genes into a plant species. This technology is seen very much as forming an integral component of a crop management programme. Several different classes of plant proteins have been shown to be insecticidal towards a range of economically important insect pests from different orders; in some cases a role in the defence of specific plant species against phytophagous insects has been demonstrated. Genes encoding insecticidal proteins have been isolated from various plant species and transferred to crops by genetic engineering. Amongst these genes are those that encode inhibitors of proteases (serine and cysteine) and α-amylase, lectins, and enzymes such as chitinases and lipoxygenases. Examples of genetically engineered crops expressing insecticidal plant proteins from different plant species, with enhanced resistance to one or more insect pests from the orders Lepidoptera, Homoptera and Coleoptera are presented. The possibility of ‘pyramiding’ different resistance genes to improve the effectiveness of protection and durability is discussed and exemplified. The number of different crop species expressing such genes is very diverse and ever-increasing. The viability of this approach to crop protection is considered. © 1998 SCI.     

Soil application of imidacloprid and related SAR-inducing compounds produces effective and persistent control of citrus canker

Soil application of the systemic insecticide imidacloprid (Admire®, Bayer Crop Science) produced season-long control of citrus canker caused by Xanthomonas citri sbsp. citri. Imidacloprid is a neo-nicotinoid that breaks down in planta into 6-chloronicotinic acid, a compound closely related to the systemic acquired resistance (SAR) inducer isonicotinic acid. Potted Swingle citrumelo seedlings (Citrus paradisi × Poncirus trifoliata) were treated with imidacloprid and the SAR inducers, isonicotinic acid, and acibenzolar-s-methyl as soil drenches or with acibenzolar-s-methyl as a foliar spray 1week prior to inoculation of immature leaves with X. citri sbsp. citri. Seedlings were re-inoculated four times over a 24-week period. SAR induction was confirmed by expression of the PR-2 gene (β-1,3 glucanase). Soil drenches of imidacloprid, isonicotinic acid, and acibenzolar-s-methyl induced a high and persistent up-regulation of PR-2 gene expression and reduced the number of canker lesions for up to 24 weeks compared to 4 weeks for foliar acibenzolar-s-methyl. Soil applied inducers of SAR reduced canker lesions up to 70% compared with the untreated inoculated plants. Lesions on leaves were small, necrotic, and flat compared to pustular lesions on inoculated untreated plants. Populations of X. citri sbsp. citri per leaf were reduced 1–3 log units in soil-treated plants compared to inoculated untreated plants.     

维生素A对视黄醇结合蛋白基因表达效果初探

采用半定量RT-PCR技术对视黄醇结合蛋白(RBP)基因的表达进行研究.在猪的成纤维细胞内添加浓度为0.5、5、10、30μg/μL维生素A,分别作用12、24、48、72 h后,比较分析RBP的表达情况.结果表明,添加10μg/μL的维生素A作用72 h后,RBP的表达量下降了92.6%;添加5μg/μL的维生素A作用72 h后,RBP的表达量最高,为对照组的1.7倍.适量的维生素A可以诱导RBP表达,但过量的维生素A则会抑制RBP表达.     

Purification and characterisation of proteins secreted by the entomopathogenic fungus Metarhizium anisopliae with insecticidal activity against adults of the Mediterranean fruit fly,Ceratitis capitata (Diptera: Tephritidae)

BACKGROUND: The control of the Mediterranean fruit fly (medfly) Ceratitis capitata (Wied) is usually performed with protein bait sprays incorporating chemical insecticides that may have adverse effects on humans, non‐target organisms and the environment. In recent years, scientists have sought more environmentally friendly insecticides for medfly control, such as plant‐ and microorganism‐derived compounds. Among these compounds, entomopathogenic fungi are an unexplored source of natural insecticides. RESULTS: The crude soluble protein extract (CSPE) of the entomopathogenic fungus Metarhizium anisopliae (Mestch.) (strain EAMa 01/58‐Su) shows chronic insecticidal activity when administered per os. Mortality in flies exhibits a dose response. The CSPE produces an antifeedant effect in adult flies, a result probably due to a progressive deterioration of the fly midgut after ingestion of the extract. Protease and temperature treatments show that insecticidal activity against C. capitata is due to proteinaceous compounds that are highly thermostable. Four monomeric proteins from this crude extract have been purified by liquid chromatography and gel electroelution. Although all four monomers seem to be involved in the insecticidal activity of the CSPE, the 15 kDa and the 11 kDa proteins appear to be mainly responsible for the observed insecticidal effect. CONCLUSIONS: Four new fungal proteins with insecticidal activity have been purified and identified. These proteins might be combined with insect baits for C. capitata biocontrol. Copyright © 2009 Society of Chemical Industry     

不同品种马血液蛋白(酶)遗传多态性及聚类分析

采用垂直平板不连续聚丙烯酰胺凝胶电泳技术分别对蒙古马、三河马及纯血马的8种血液蛋向进行多态性检测.计算了各位点的基因型频率、基因频率、遗传杂合度,并通过聚类分析探讨了群体间的亲缘关系和遗传距离.结果表明:种血液蛋白(酶)位点在不同程度上呈现出多态.平均杂合度分别为:蒙古马0.378 2,三河马0.395 6和纯血马0.257 1.聚类分析表明,蒙古马与三河马间的亲缘关系最近,与纯血马的亲缘关系相对较远.通过分析不同马品种的遗传关系,为中国马遗传资源的保护与育种提供依据.     

极端嗜热菌Pyrococcus furiosus热休克蛋白HSP60的克隆表达和纯化

[目的]研究极端嗜热菌Pyrococcus furiosus的热休克蛋白HSP60(Pfu-HSP60)的克隆表达与纯化方法。[方法]应用加拿大HSP60数据库及其BLAST和FASTA序列比较工具对Pfu-HSP60基因序列进行同源性分析,对HSP60基因进行了体外扩增并在大肠杆菌中成功克隆与表达,还进行了Pfu-HSP60蛋白的纯化。[结果]以Taq酶为DNA聚合酶,用PCR方法扩增出预期的Pfu-HSP60基因,插入质粒pET-21a,转化入大肠杆菌受体菌BL21-Codonplus(DE)3-RIL,0.1mmol/LIPTG诱导2 h,得到可溶性高表达,经超声、离心、上清85℃加热、再离心得到初步纯化的Pfu-HSP60蛋白,再经DEAE Sepharose Fast Flow阴离子交换层析纯化,收集峰超滤浓缩后经SephacrylS-200凝胶过滤柱,收集Pfu-HSP60蛋白峰,结果获得纯度为94.5%以上的重组Pfu-HSP60蛋白,总得率为28.0%。[结论]该研究为Pfu-HSP60蛋白的结构和功能及其他热休克蛋白相互作用机制等的研究奠定基础。     

水稻种子蛋白质含量及组分在品种间的变异与分布

采用近红外光谱技术测定分析了351份不同类型水稻品种(系)糙米中的蛋白质含量,结果显示粗蛋白含量在9.3%~17.7%之间,平均为12.4%;籼稻平均蛋白质含量为13.2%,比粳稻高约1个百分点。蛋白质含量低的粳稻品种明显偏多,表现出明显的遗传不平衡现象。现有生产上主栽品种稻米蛋白质含量大多处于中等水平,而高蛋白粳稻种质极少。但仍有部分蛋白质含量极高或低的种质,如饲料稻、早籼稻和一些籼粳交后代品系蛋白质含量较高,而部分粳稻和外来籼稻品种中的蛋白质含量较低。因此可以从一些地方品种、外来品种以及籼粳交后代中筛选到极端类型的种质,为遗传育种提供研究的原材料。SDS-PAGE分析结果显示不同类型水稻间各贮藏蛋白组分具一定差异。