抗除草剂转基因作物面临的机遇与挑战及其发展策略

2008年全球抗除草剂转基因作物已占转基因作物总种植面积的80%以上,抗除草剂成为最主要的转基因应用的性状.该技术的成功应用极大地降低了杂草防除成本、增加了安全性和减少了除草剂的残留药害.但是,抗除草剂转基因作物的大规模产业化将导致除草剂市场单一化,从而冲击除草剂产业,还会产生杂草抗药性和基因逃逸等环境安全问题,因此抗除草剂转基因作物面临着挑战.在我国转基因抗除草剂作物还没有实际的商业化,不过由于市场巨大、政策支持等面临良好发展机遇.此外,从抗除草剂基因利用、转基因抗除草剂作物的研究与开发策略等方面进行综述,以期为我国转基因抗除草剂作物的研究与开发提供可能有益的信息.     

Grain and biomass nutrient uptake of conventional corn and their genetically modified isolines

The adoption of genetically engineered crops in the United States has increased dramatically over the past decade. Differences in agronomic characteristics and protein expression between genetically engineered plants and their naturally recombinant non-genetically modified (GM) counterparts are not well-understood. Experimental field plots were established in the spring of 2005 near Brookings, SD with 18 different commonly used corn hybrids including three conventional hybrids and their corresponding transgenic modifications. Specific research objectives were to evaluate in a side by side comparison, the impact of the genetic modifications on agronomic characteristics. Results show that glyphosate or insect resistance resulting from genetic modification, in the absence of significant insect pest or weed pressure and glyphosate application, were not likely to significantly alter productivity or nutrient composition of corn residue or grain. No significant differences were observed among the hybrids in average grain yield or above-ground biomass over the three years of the experiment.     

叶绿体转化及其应用于作物改良研究的最新进展

农杆菌介导的核基因转化技术已普遍应用于作物改良和分子育种方面,而其不可避免的转基因沉默现象和生物安全问题是科研工作者和广大民众关注的焦点,进而成为基因工程技术广泛应用的主要限制因素。叶绿体遗传转化技术因具有核转化不可比拟的独特优势,成为植物基因工程发展的新方向和科研工作者研究的热点之一,通过该技术有望培育出真正生物安全的转基因作物新品种。本文围绕叶绿体转化的原理、方法、筛选标记及其应用等方面进行阐述,着重介绍其在作物改良方面研究的最新进展,并对存在问题及应用前景进行了展望。     

Composition of transgenic soybean seeds with higher γ-linolenic acid content is equivalent to that of conventional control

γ-Linolenic acid (GLA) has been used as a general nutraceutical for pharmacologic applications, particularly in the treatment of skin conditions such as eczema. Four transgenic soybean lines that produce GLA at high yields (4.21% of total fatty acids, up to 1002-fold) were generated through the stable insertion of the Delta-6-fatty acid desaturase gene isolated from Borago officinalis into the genome of a conventional soybean cultivar. As part of the safety assessment of genetically engineered crops, the transgenic soybean seeds were compared with their parental soybean seeds (nontransgenic) by applying the principle of substantial equivalence. Compositional analyses were conducted by measuring the fatty acids, proximate analysis (moisture, crude protein, crude fat, carbohydrates, TDF, and ash contents), amino acids, lectins, and trypsin inhibitor activity. The present results showed that the specific transgenic cultivar studied was similar to the conventional control.     

四价抗病虫除草剂RNAi植物表达载体构建与转化甘蔗研究

甘蔗是重要的能源作物和糖料作物,面临病虫害日益严重问题。由于甘蔗遗传背景非常复杂,且缺乏必要种质,难以通过杂交培育抗病虫、除草剂聚合品种。基因工程技术提供了新方法。本研究利用BLAST比对获得甘蔗花叶病毒ScMV-CP基因和黄叶病毒ScYLV-CP基因的保守序列作为干扰序列,通过设计内切酶位点、生物合成和连接,获得全长687 bp的两个保守序列融合片段（MYCP）。并将其正向和反向插入中间载体pHANNIBAL上,形成双价病毒干扰结构。再通过在pHANNIBAL上添加SmaⅠ内切酶位点,将双价病毒干扰结构表达框和Cry1AC基因表达框结合;利用NotⅠ酶切,回收双价病毒干扰结构和Cry1AC基因表达框片段,连接到添加Bar基因表达框的表达载体pART27-Z上,构建了抗病虫及除草剂四价植物表达载体（pART27-MYCP-Cry1AC）。采用基因枪法将构建好的四价植物表达载体转入甘蔗品种FN15的胚性愈伤组织中,通过PPT（草丁膦）筛选获得了抗性再生甘蔗植株,经过PCR、定量以及Bt蛋白试纸检测,获得同时整合四种目的基因并表达的四价转基因甘蔗植株,为培育多抗甘蔗新品种提供了材料。     

Glyphosate degradation in glyphosate-resistant and -susceptible crops and weeds

High levels of aminomethylphosphonic acid (AMPA), the main glyphosate metabolite, have been found in glyphosate-treated, glyphosate-resistant (GR) soybean, apparently due to plant glyphosate oxidoreductase (GOX)-like activity. AMPA is mildly phytotoxic, and under some conditions the AMPA accumulating in GR soybean correlates with glyphosate-caused phytotoxicity. A bacterial GOX is used in GR canola, and an altered bacterial glyphosate N-acetyltransferase is planned for a new generation of GR crops. In some weed species, glyphosate degradation could contribute to natural resistance. Neither an isolated plant GOX enzyme nor a gene for it has yet been reported in plants. Gene mutation or amplification of plant genes for GOX-like enzyme activity or horizontal transfer of microbial genes from glyphosate-degrading enzymes could produce GR weeds. Yet, there is no evidence that metabolic degradation plays a significant role in evolved resistance to glyphosate. This is unexpected, considering the extreme selection pressure for evolution of glyphosate resistance in weeds and the difficulty in plants of evolving glyphosate resistance via other mechanisms.     

ISSR variation within and among wild <Emphasis Type="Italic">Brassica juncea</Emphasis> populations: implication for herbicide resistance evolution

There is a need to understand whether weed genetic diversity is the same among different populations, especially between those exposed to herbicide selection and other without exposure history. Inter-simple sequence repeat (ISSR) were used to assess level and patterns of genetic diversity in wild Brassica juncea (L.) Czern. et Coss. populations. A total of 93 plants from 24 wild populations in China were analysed by eight primers resulting in 86 highly reproducible ISSR bands. The analysis of molecular variance (AMOVA) with distances among individuals corrected for the dominant nature of ISSRs showed that most of the variation (54.09%) occurred among populations, and the remaining 45.91% variance was attributed to differences among individuals within populations. The high differentiation was, perhaps, due to limited gene flow (Nm < 1.0) of this species. Though highest gene diversity was observed in resistant B. juncea population, the overall distribution of diversity across China was not geographic dependent. High F _ST value (0.541) corroborated AMOVA partitioning and provided significant evidence for population differentiation in wild B. juncea. UPGMA cluster analyses, based on Nei’s genetic distance, revealed grouping pattern geographically. Based on these results, the factors affect weed population genetic diversity and implication for herbicide resistance evolution were discussed in the context of transgenic crops advent and increasing herbicide usage in China.     

Protein biofortified sorghum: effect of processing into traditional african foods on their protein quality

Protein biofortification into crops is a means to combat childhood protein-energy malnutrition (PEM) in developing countries, by increasing the bioavailability of protein in staple plant foods and ensuring sustainability of the crop. Protein biofortification of sorghum has been achieved by both chemically induced mutation and genetic engineering. For this biofortification to be effective, the improved protein quality in the grain must be retained when it is processed into staple African foods. Suppression of kafirin synthesis by genetic engineering appeared to be superior to improved protein digestibility by chemical mutagenesis, because both the lysine content and protein digestibility were substantially improved and maintained in a range of African foods. For the genetically engineered sorghums, the protein digestibility corrected amino acid score was almost twice that of their null controls and considerably higher than the high protein digestibility sorghum type. Such protein biofortified sorghum has considerable potential to alleviate PEM.     

Genetic diversity in tomato (Solanum lycopersicum L.) germplasm using fruit variation implemented by tomato analyzer software based on high throughput phenotyping

The decrease of genetic variation in crops like tomato (Solanum lycopersicum L.) due to domestication, breeding, and population bottle necks for fresh market consumptions has increased the need for this cash crop’s conservation and characterizations. To study the genetic variation of this valuable genetic resource in the National Plant Gene Bank of Iran, a collection of 589 tomato accessions from different regions was characterized using semi-automatic and high-throughput techniques. Based on the fruit shape, the studied accessions were classified into ten groups and significant differences for all the fruit characteristics studied indicated an abundant diversity within the germplasm collected from different regions. Studied tomato germplasm was grouped into nine main clusters based on hierarchical cluster analysis. More than 83% of data variation was explained by seven components in which the first two components explained for 50% of the variation where fruit shape index and proximal/distal fruit end shape showed a high contribution in the variation of the first component. This unique genetically divergent germplasm could be utilized to select tomato breeding lines of interest. Also, accessions belonged to the heterotic clusters could be selected for hybridization and conservation purpose as well as enrichment of pre-breeding programs of the tomato germplasm.

     

Comparing conventional and biotechnology-based pest management

Pest management has changed dramatically during the past 15 years by the introduction of transgenes into crops for the purpose of pest management. Transgenes for herbicide resistance or for production of one or more Bt toxins are the predominant pest management traits currently available. These two traits have been rapidly adopted where available because of their superior efficacy and simplification of pest management for the farmer. Furthermore, they have substantially reduced the use of environmentally and toxicologically suspect pesticides while reducing the carbon footprint of pest management as reduced tillage became more common, along with fewer trips across the field to spray pesticides. The most successful of these products have been glyphosate-resistant crops, which cover approximately 85% of all land occupied by transgenic crops. Over-reliance on glyphosate with continual use of these crops has resulted in the evolution of highly problematic glyphosate-resistant weeds. This situation has resulted in some farmers using weed management methods similar to those used with conventional crops. Evolution of resistance has not been a significant problem with Bt crops, perhaps because of a mandated resistance management strategy. Transgenic crops with multiple genes for resistance to different herbicides and resistance to additional insects will be available in the next few years. These products will offer opportunities for the kind of pest management diversity that is more sustainable than that provided by the first generation of transgenic crops.     

Strategies for bacterial tagging and gene expression in plant-host colonization studies

Bacteria are extraordinarily diverse microorganisms with a huge potential to benefit environmental and agricultural systems. Comprehensive studies in complex habitats such as soils and plants have led to the development of genetic tools to evaluate gene expression and bacterial colonization under controlled or environmental conditions and to obtain genetically engineered organisms for environmental release. In addition, current advances in genomic and metagenomic research will add to the number of genes with potential for biotechnological applications, which will require the development of appropriate genetic systems to fulfill their potential for both industrial and agricultural applications. The aim of the present review is to assess the approaches and recent progress in vector design and genetic tools to study and manipulate plant-bacterial interactions, as well as strategies to construct genetically modified strains for environmental release.     

Evaluation of compositional equivalence for multitrait biotechnology crops

Compositional analysis is an important tool in the evaluation of the safety and nutritional status of biotechnology-derived crops. As part of the comparative assessment of a biotechnology-derived crop, its composition is evaluated by quantitative measurement of the levels of key nutrients, antinutrients, and secondary metabolites and compared to that of conventional crops. To evaluate the effect of combining multiple biotech traits through conventional breeding, the forage and grain compositions of the double combinations MON 810 × NK603, MON 863 × MON 810, and MON 863 × NK603 and the triple combination MON 863 × NK603 × MON 810 were compared to their respective near-isogenic, conventional control hybrids. Overall, a total of 241 statistical comparisons between the multitrait biotechnology crop and its corresponding conventional controls were conducted. Of these comparisons 192 (79.7%) were not statistically significantly different (p > 0.05), and all 49 of the differences were within the 99% tolerance interval for commercial hybrids grown in the same field or related field trials. These data on combined trait biotechnology-derived products demonstrated that the forage and grain were compositionally equivalent to their conventional comparators, indicating the absence of any influence of combining insect protection and herbicide tolerance traits by conventional breeding on compositional variation.     

谷子抗除草剂基因的发现及其应用

谷子田间除草一直是阻碍谷子产业化发展的主要因素之一。谷子抗除草剂品种的出现,解决了谷子田间除草难的问题,有利于谷子大面积的栽培,使谷子产业化发展成为可能。本文围绕谷子抗除草剂基因的发现、类型和机理以及抗除草剂品种在谷子生产中的应用情况等方面进行阐述,探讨谷子抗除草剂应用对环境的影响,并对谷子抗除草剂研究中存在的问题及应用前景进行了展望。     

Genetic diversity of thiamin and folate in primitive cultivated and wild potato (Solanum) species

Biofortification of staple crops like potato via breeding is an attractive strategy to reduce human micronutrient deficiencies. A prerequisite is metabolic phenotyping of genetically diverse material which can potentially be used as parents in breeding programs. Thus, the natural genetic diversity of thiamin and folate contents was investigated in indigenous cultivated potatoes (Solanum tuberosum group Andigenum) and wild potato species (Solanum section Petota). Significant differences were found among clones and species. For about 50% of the clones there were variations in thiamin and folate contents between years. Genotypes which contained over 2-fold the thiamin and 4-fold the folate content compared to the modern variety Russet Burbank were identified and should be useful material to integrate in breeding programs which aim to enhance the nutritional value of potato. Primitive cultivars and wild species with widely different amounts of thiamin and folate will also be valuable tools to explore their respective metabolic regulation.     

植物基因定点突变与定点置换技术及其在作物遗传改良中的应用

随着大量植物基因组测序工作的完成及EST数据库的充实，利用基因定点突变与定点置换技术精细的研究基因功能是功能基因组学研究的重要手段。同时，基因定点突变与定点置换技术可以克服现有转基因技术的基因沉默和位置效应等诸多缺陷，在植物遗传改良方面具有重要的意义。基因定点突变与定点置换技术在酵母和小鼠胚胎干细胞中已经比较成熟，但在高等植物中同源重组频率非常低，限制了其使用。新近研究发现，利用锌指核酶（Zinc finger nucleases，ZFNs）引入DNA分子的定点断裂（double-strand breaks, DSBs）可以高效介导基因的同源重组，使得ZFN成为遗传工程的一个研究热点。利用嵌合寡核苷酸(Chimeric oligonucleotides)可以介导基因的单碱基定点突变，在植物遗传改良上有广阔的应用前景。对同源重组相关的调控途径进行基因修饰也可以提高植物的基因打靶效率。植物基因定点突变与定点置换技术难题的攻克，必将加快植物功能基因组研究的步伐，同时给植物基因工程育种带来新的革命。     

Centres of Crop Diversity and/or Origin, Genetically Modified Crops and Implications for Plant Genetic Resources Conservation

The concept of centres of crop diversity and/or origin of agriculture is briefly reviewed. The conservation status of crop genetic resources, either ex situ or in situ, cultivated or wild, has been assessed for species of the Central American and Mexican centre, demonstrating that that region is indeed one of the important centres of crop diversity for human kind. Furthermore, biotechnological developments with regard to the creation and spread of genetically modified crops have been analyzed. The likelihood of unintentional introgression of genetically modified traits into conventional seed lots, crops as well as into germplasm collections have been assessed. Related biosafety measures as well as the possible implications of intellectual property rights on transgenic crops and/or genes are being discussed vis-a-vis the possible implications they might have for germplasm management. The Central American crop genetic resources situation has been used as a “case study” to illustrate the potential impact of the spreading of GM varieties on the genetic diversity in genebanks and farmers’ fields and the need for effective and efficient conservation efforts. Conservation management strategies and practices are being proposed of mitigate the potential negative impact of GM crops on the conservation efforts.     

分子标记在油菜育种中的应用

油菜是世界上重要的油料作物之一,为了提高油菜的产量,各国都致力于油菜的育种工作。随着分子生物学的迅速发展,分子标记技术广泛地应用油菜育种。本文从遗传和物理图谱构建、基因的定位与克隆、数量性状位点的遗传分析、遗传多样性的评估、分子标记辅助选择等方面,综述了分子标记在油菜育种中的应用。     

A survey of yield differences between transgenic and non-transgenic crops

In the current survey, there was no clear evidence that GM (genetically modified) crops are higher yielding than those conventionally bred&lt;fnoteref rid="fn1"&gt;1&lt;/fnoteref&gt;. Furthermore, there were no trials to support valid comparisons of yield per se. This article investigates GM crop yields, introducing the importance of hybrid vigour and a non-stress environment for higher percentage heritability selection and therefore more productive conventional plant breeding and improved crops. GM technology and crops are compared with proven plant breeding methods, with respect to hybrid vigour and the economic viability of both systems. These proven methods of plant breeding are (1) traditional landrace cropping, (2) conventional Mendelian breeding and (3) Isolection Mendelian breeding, and are also considered historically.     

基因工程技术在烟草研究中的应用现状与展望

介绍转基因、RAPD分子标记、基因芯片、mRNA差异显示以及蛋白质组学等基因工程技术在烟草中的应用,指出这些技术不仅在烟草遗传图谱建立、系统分析、物种及品系的鉴定、转基因烟草鉴定和检测等方面发挥着重要作用,而且在改善烟草品质和建立烟草基因组学方面也有广阔的应用前景,并从抗病性、抗虫性、抗旱性、抗除草剂等方面介绍了基因工程技术在烟草抗逆性中的研究现状,对转基因烟草的安全性问题进行了探讨。     

苏云金芽孢杆菌cry1Ia基因的克隆、表达与活性研究

根据cry1Ia类基因的全长序列设计引物，以苏云金芽孢杆菌（Bacillus thuringiensis）菌Btc008的总DNA为模板扩增出片段长为2.1kb的cry1Ia的全长基因，插入大肠杆菌（Escherichia coli）表达载体pET-21b，转化大肠杆菌BL21（DE3）菌株，诱导表达出81kD的蛋白。该蛋白由719个氨基酸组成，推导的分子量为81.2kDa。该蛋白的氨基酸序列不同于已知的12种Cry1Ia蛋白，是一种新的Cry1Ia蛋白，该基因已被国际基因命名委员会正式命名为cry1Ia8。杀虫活性测定结果表明：Cry1Ia8对亚洲玉米螟（Ostrinia furnacalis）、小菜蛾（Plutella xylostella）有很强的杀虫活性，LC50分别为0.268 µg/g、2.227 µg/ml，其杀虫效果与Cry1Ab、Cry1Ac相当。对大豆食心虫（Leguminivora glycinivorella）也有较好的活性，但对鞘翅目叶甲科害虫榆兰叶甲（Pyrrhalta aenescens）没有活性。该基因的获得将为我国抗虫转基因作物和工程菌的研制提供新的基因来源，为筛选延缓昆虫抗性产生的基因组合提供了极为重要的依据。