Bt杀虫基因与Bt转基因抗虫植物研究进展

苏云金杆菌是一类非常重要的昆虫病原体,它能产生特异性的杀虫结晶蛋白,对农业上和生物医学上的许多有害的昆虫有毒杀作用,这些害虫包括鳞翅目、双翅目、鞘翅目、膜翅目、螨类和线虫。近三十年来,以苏云金杆菌为基础的生物杀虫剂已在世界范围内商业化用于防治重要经济作物的害虫。近年来有关Ｂｔ基因的遗传、分子生物学和基因工程已取得显著进展。本文对苏云金杆菌杀虫晶体蛋白基因的分类和杀虫机理及用该类基因构建的工程转基因植物研究状况作一简要综述,同时对Ｂｔ基因工程存在的潜在问题和解决途径作了简单的探讨。     

转苏云金芽胞杆菌杀虫晶体蛋白基因抗虫植物的研究进展与商品化

本文综述了转ICPs基因植物从ICPs在植物中低表达,具杀虫效应到高表达而进行田间应用试验的研究进展及转ICPs抗虫植物的商品化现状。     

Bt转基因抗虫植物研究进展

目前Bt成为世界上植物抗虫基因工程中研究和应用最多的基因。已经有多种转单一Bt和复合Bt的转基因玉米、棉花、马铃薯等作物得到大面积种植。还有许多新的具有良好抗虫性的Bt转基因植物,如水稻、大豆、油菜、苜蓿、花椰菜、蓖麻等已经试验成功,并逐渐推广种植。Bt转基因抗虫植物的培育为提高产量,减少杀虫剂的使用和保护环境做出了举足轻重的贡献。就Bt的分类、杀虫机理、Bt转基因抗虫植物的发展状况以及种植Bt抗虫植物对环境的影响进行了概述。     

植物抗虫基因工程研究进展

植物抗虫基因工程为防治农业害虫提供了一条崭新途径。本文对植物抗虫基因工程近年来所取得的某些研究进展,包括目前已发现和利用的抗虫基因、提高抗虫基因在植物体内表达的方法以及防止或延缓害虫产生抗性的策略等方面进行了综合评述,并对植物抗虫基因工程中有待解决的问题和发展前景提出了自己的看法。     

植物抗虫基因工程研究进展

植物抗虫基因工程为防治农业害虫提供了一条崭新途径。本文对植物抗虫基因工程近年来所取得的某些研究进展,包括目前已发现和利用的抗虫基因、提高抗虫基因在植物体内表达的方法以及防止或延缓害虫产生抗性的策略等方面进行了综合评述,并对植物抗虫基因工程中有待解决的问题和发展前提提出了自己的看法。     

植物来源抗虫基因的应用

作为植物抗虫基因工程研究中的重要组成部分,植物来源的抗虫物质及其在基因工程中的应用日益受到人们的重视。本文就植物来源的蛋白酶抑制剂、凝集素、淀粉酶抑制剂等抗虫物质的定义、分类、作用机理,及其在植物抗虫基因工程中的应用进行了论述,并对面临的问题及应用前景提出了个人的看法。     

植物来源抗虫基因的研究进展

植物抗虫基因工程为农林业生产中的害虫防治提供了新的途径,随着研究的不断深入,已经获得了很多抗虫基因。本文综述了目前源于植物的抗虫基因的种类,归纳了研究较多的抗虫基因的作用机制及其在转基因植物中的应用,提出了植物抗虫基因在虫害防治中面临的问题及相应的解决策略,展望了植物抗虫基因工程研究发展的方向和前景。     

植物抗虫基因工程研究进展

植物抗虫基因工程研究进展周兆斓,朱祯（中国科学院遗传研究所,北京１００１０１）在世界范围内,虫害造成的损失约占农作物总收获量的１３％,每年大约损失数千亿美元。目前,广泛使用化学杀虫剂控制虫害,由于化学杀虫剂的作用方式是非特异的,因此在施放杀虫剂时不但杀死了害虫,同时也杀死了有益昆虫及害虫的天敌,造成生态平衡的破坏;化学杀虫剂对人、畜也有严重危害,其在自然界中的残留以及在食物链中的积累已造成了严重的环境污染;另外,由于长期使用农药,害虫已逐步对其产生抗性,目前已经出现了几百种对杀虫剂有耐受性的害虫,有些甚至到了无药可治的程度。据有关部门反应,我国棉花的主要害虫-棉铃虫对农药的耐受性有逐年提高的趋势,常规使用剂量已不能有效地控制其危害。     

抗虫植物基因工程研究进展

虫害是造成农业减产的主要原因之一。据不完全统计,全世界每年因虫害引起的作物减产达总产量的15％,损失高达数千亿美元。在我国,因虫害水稻减产在lO％以上;小麦减产近20％;棉花减产在     

苏云金杆菌4.0718质粒上杀虫晶体蛋白基因的PCR分析

苏云金杆菌（Bacillus Thuringiensis)4.0718由本实验室选育是对鳞翅目（Lepidopteran)昆虫有高毒性的菌株。采用SDS温和提取方法提取此菌株的质粒,电泳纯化后获得了其4种不同分子量的质粒P1,P2,P3,P4。利用序列分析软件对已知的cryⅠ类型基因进行阵列比较,根据其高度保过区域设计了1对通用引物,对cryⅠ类型基因的扩增产物对277bp左右的片段,用此引的对Bacillus thuringiensis 4.0718的4种不同质粒进行了PCR分析,发现质粒P1,P2,P3扩增阳性,都产生了277bp的扩增片段,而质粒P4没有扩增产物,这为进一步的基因定位打下了基础。     

苏云金芽胞杆菌杀虫晶体蛋白基因的分类

１９８９年Ｈｏｆｔｅ和Ｗｈｉｔｅｌｅｙ根据氨基酸序列的同源性和杀虫谱双重标准将苏云金芽胞杆菌杀虫晶体蛋白基因划分为５类１４亚类。根据双重标准出现的问题和冲突,给分类带来了困扰,为此１９９５年,只根据氨基酸序列的同源性进行分类。现已将基因分为２１群,４４亚群,６４类,１１６个亚类。本文介绍了纳入新系统的条件、分类原则、命名方法、定名步骤和分类中值得注意的问题。     

Genetic Engineering of Insect-Resistant Crop Plants with Bacillus thuringiensis Crystal Protein Genes

Bacillus thuringiensis, a soil bacterium, produces crystalline proteins which are toxic to the Insect larvae, The toxicity Is brought about by the protein fragments released due to the action of mid-gut proteases and their binding to the receptors on the epithelial membrane, which In turn loses Its selective permeability. The genes (cry) coding for these Insecticidal crystal proteins have been cloned, characterized and mobilized Into a number of crop plants. Such transgenic plants were shown to be resistant to insects. However, the expression of these bacterial genes in higher plants has been limited because of differential codon usage. Various strategies for maximizing the expression of these genes in transgenic plants have been described. In addition, alternative approaches have been suggested for circumventing the development of resistance in insects to the crystal proteins.     

The Biotechnology of Bacillus Thuringiensis

Abstract

Gibberellins are a classic example of the production of plant growth regulators by microorganisms. They are important biotechnological products and are increasingly used in agriculture and horticulture.

This article intends to assemble information on the history of the identification of gibberellins (GA) and producing microorganisms, especially Gibberella fujikuroi (Saw.) Wr. Furthermore, the biosynthesis of gibberelins through the isoprenoid biosynthetic pathway will be described. The main product of GA biosynthesis in Gibberella fujikuroi is gibberellic acid (GA₃), which is formed from GA₄ via GA₇. Both the amount and the type of gibberellins produced by the fungus are dependent on the genetic constitution of the strain and the fermentation conditions.

Mutation and selection for increased product formation are probably the most important factors in improving the yield of gibberellins. Some publications concerning methods of parasexual recombination will also be summarized. Beside strain improvement of wild strains, medium development and appropriate cultivation techniques (batch, fed-batch-, continuous-, and solid state-fermentation) are very important prerequisites for successful economy of gibberellin production. Furthermore, the most important ways of gibberellin recovery and purification are described. Continuing reductions in the costs make gibberellins more attractive for existing applications and open possibilities for further applications of GA₃ and some other active gibberellins like GA₄, Ga₇, and GA₉     

植物抗早基因工程研究进展

从植物抗虫基因工程的研究历史出发,论述了第一代抗虫基因、第二代抗虫基因,重点介绍了B.t．杀虫晶体蛋白基因、胆固醇氧化酶基因和营养杀虫蛋白基因,并对植物抗虫基因工程中所遇到的问题和解决办法进行了探讨。     

Epsps基因为筛选标记的多基因抗虫表达载体构建

目的:为了获得水稻的广谱抗虫性,同时也避免由潮霉素抗性基因引起的安全性问题.构建以抗除草剂基因(Epsps 基因)为筛选标记的多基因抗虫植物表达载体 pCTSK.方法:以双元载体pCAMBIA1300为基础,将潮霉素抗性基因hpt替换成携带烟草叶绿体转运肽序列TSP的草甘膦抗性突变基因aroAM12(编码细菌5-烯醇式丙酮酸莽草酸-3-磷酸合成酶Epsps)作为筛选标记基因,获得中间表达载体pCISM1300.然后再连接上3个抗虫基因(马铃薯蛋白酶抑制剂基因Pin-Ⅱ、苏云金杆菌毒蛋白基因Bt cryI(A)及雪花莲外源凝集素基因 CNA)的完整表达片段(包含各自的启动子和终止子).结果:通过PCR方法和酶切方法鉴定已成功地构建了该表达载体;通过不同实验方案的数据分析,得出在载体构建的大分子连接中优化的一次连接法连接效率高于二次连接法的结论.     

Molecular Characterization and Genetic Diversity of Insecticidal Crystal Protein Genes in Native Bacillus thuringiensis Isolates

The Western Ghats of Karnataka natural ecosystem are among the most diverse and is one of the eight hottest hotspots of biological diversity in the world, that runs along the western part of India through four states including Karnataka. Bacillus thuringiensis (Bt) strains were isolated from soils of Western Ghats of Karnataka and characterized by molecular and analytical methods as a result of which 28 new Bt-like isolates were identified. Bt strains were isolated from soil samples using sodium acetate selection method. The morphology of crystals was studied using light and phase contrast microscopy. Isolates were further characterized for insecticidal cry gene by PCR, composition of toxins in bacterial crystals by SDS-PAGE cloning, sequencing and evaluation of toxicity was done. As a result 28 new Bt-like isolates were identified. Majority of the isolates showed the presence of a 55 kDa protein bands on SDS-PAGE while the rest showed 130, 73, 34, and 25 kDa bands. PCR analysis revealed predominance of Coleopteran-active cry genes in these isolates. The variations in the nucleotide sequences, crystal morphology, and mass of crystal protein(s) purified from the Bt isolates revealed genetic and molecular diversity. Three strains containing Coleopteran-active cry genes showed higher activity against larvae Myllocerus undecimpustulatus undatus Marshall (Coleoptera: Curculionidae) than B. thuringiensis subsp. Morrisoni. Results indicated that Bt isolates could be utilized for bioinsecticide production, aiming to reduce the use of chemical insecticide which could be useful to use in integrated pest management to control agriculturally important pests for sustainable crop production.     

New Strategy for Isolating Novel Nematicidal Crystal Protein Genes from Bacillus thuringiensis Strain YBT-1518

We have developed a strategy for isolating cry genes from Bacillus thuringiensis. The key steps are the construction of a DNA library in an acrystalliferous B. thuringiensis host strain and screening for the formation of crystal through optical microscopy observation and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analyses. By this method, three cry genes—cry55Aa1, cry6Aa2, and cry5Ba2—were cloned from rice-shaped crystals, producing B. thuringiensis YBT-1518, which consists of 54- and 45-kDa crystal proteins. cry55Aa1 encoded a 45-kDa protein, cry6Aa2 encoded a 54-kDa protein, and cry5Ba2 remained cryptic in strain YBT-1518, as shown by SDS-PAGE or microscopic observation. Proteins encoded by these three genes are all toxic to the root knot nematode Meloidogyne hapla. The two genes cry55Aa1 and cry6Aa2 were found to be located on a plasmid with a rather small size of 17.7 kb, designated pBMB0228.     

苏云金芽胞杆菌杀虫晶体蛋白作用的分子机制研究进展

本文主要综述了苏云金芽胞杆菌(Bacillusthuringiensis,Bt)杀虫晶体蛋白在分子水平上作用机制的研究进展。杀虫晶体蛋白经蛋白酶活化后形成的毒性肽一般由三个结构域组成。在杀虫过程中,毒性肽首先通过结构域Ⅱ或结构域Ⅲ的特殊部位与昆虫中肠上皮细胞膜上的受体蛋白发生专一性结合。这一结合开始是可逆的,随后发生紧密的不可逆结合。继而诱发毒性肽分子发生空间构象变化,使得结构域Ⅰ中的某些α螺旋从α螺旋束中弹出并插入细胞膜,并通过寡聚合作用造成膜穿孔,导致细胞渗透平衡破坏、中肠破裂、昆虫死亡 。     

A Translation Fusion Product of Two Different Insecticidal Crystal Protein Genes of Bacillus thuringiensis Exhibits an Enlarged Insecticidal Spectrum

Two truncated Bacillus thuringiensis crystal protein genes, belonging to the classes cryIA(b) and cryIC and both coding for insecticidal N-terminal fragments of the corresponding crystal proteins, were translationally fused. Expression of the gene fusion in Escherichia coli showed a biologically active protein with a toxicity spectrum that overlapped those of both contributing crystal proteins.