The Experimental and Commercial Release of Transgenic Crop Plants

With advances in recombinant DNA methods and transformation procedures, it is possible to transfer genes into crop plants from unrelated plants, microbes and animals. Many of the modifications being carried out, or envisaged, are for disease and pest resistance, product quality and tolerance to environmental stress, but there are additional opportunities to modify crops to give specialized products for industrial or pharmaceutical use. Some of the characteristics of transgenic plants are considered, including: transgene copy number, position, expression, stability, pleiotropy, selectable marker genes and somaclonal variation. There have been several hundreds of field trials with transgenic plants, and the first transgenic varieties are likely to be approved for commercial production in 1993. Before releasing transgenic plants, it is necessary to carry out a risk assessment to determine whether the transgenic variety will behave differently from a conventionally bred variety. Assessment procedures are being harmonized internationally by various organizations. There is a growing commitment to apply these genetic modification methods to crops in developing countries, as genes relevant to their crops and environments become available.     

Herbicide-tolerant crops in agriculture: oilseed rape as a case study

Oilseed rape has been modified extensively by conventional breeding for the production of varieties useful for human consumption (blended vegetable oil and margarine) and industrial processes (rubber additives and high‐temperature lubricants). Because much is now known about its genetic and biochemical composition, it has been an obvious choice for genetic modification and is now at the forefront of the commercial development of genetically modified (GM) or transgenic crops. Around the world, the increase in commercial plantings of all transgenic crops has been rapid. In 1996, 1.7 million hectares were planted, but by 1998 this figure had jumped to 27.8 million ha. The area in the year 2001 is likely to be about 50 million ha. With the possible introduction of transgenic varieties into European agriculture, it is essential that the associated farming practices employed are appropriate for their growth, both from a commercial and an environmental viewpoint. Some of the first transgenic crops are those carrying agronomic traits, e.g. herbicide tolerance transgenes. However, before full commercialization occurs, important agronomic and environmental questions need to be answered. How are these new crops to be incorporated into existing cropping practices? How will this change the current herbicide use profile for a given crop? Do herbicide‐tolerant varieties enhance or impede integrated pest management schemes? What is the likely uptake of such crops in agriculture? What are the ecological implications of their introduction? Are there effective measures to control the spread of transgenes to wild relatives? This paper addresses these questions, with special emphasis on oilseed rape production in the UK, but includes examples from other crops and countries where appropriate.     

Assessing the threat to resistant cultivars from public risk diseases and pests

A framework is developed on epidemiological criteria to classify the degree of public risk and the threat to resistant cultivars from a range of diseases within a region. A public risk disease or pest is one that is an external threat to the productivity of a field, posed by a plant pathogen or pest that originates elsewhere. Yield losses can occur in neighbouring crops when susceptible cultivars are grown and become diseased. There is also an increased risk of the emergence of new pathotypes. Costs arise directly from yield losses and from maintenance breeding that is necessary to counter the risk of new pathotypes or biotypes. The framework is based on the four components of the disease tetrahedron (host, pathogen, environment, management) and their interactions with the principal criteria and modifying factors that are identified. The principal criteria that influence public risk are inoculum dispersal and pathogen variability. These can be modified by the degree of pathogen variability, reproductive capacity, seasonal favourability, host non compensation, production system, and overseasoning location. The major threat arising from public risk diseases is their potential to reduce the effective life of resistant cultivars. As an illustration, the framework is applied to the wheat industry in southern New South Wales, Australia, to classify diseases into those with public risk that can result in high, medium, low and negligible threats to resistant cultivars. Measures for reducing the threats posed by public risk diseases are discussed.     

转基因作物环境安全性研究进展

随着转基因作物在全球范围内的广泛应用,转基因作物的环境安全性问题日益受到重视,科研工作者们对此开展了大量的研究工作。本文对近年来这方面的研究成果进行了综述,主要内容包括:1)转基因作物的杂草化问题,这里面又包含两层含义,一是转基因作物自身的杂草化问题,多数研究表明转基因并没有提高作物的生存竞争能力,在没有选择压力的自然条件下,即使转入了抗病抗逆基因,转基因植株的生存竞争能力也没有增加,因此杂草化的可能性很小;二是转基因作物通过基因漂移使得同种或近缘野生种或得某种抗性而成为更加难以防除的“超级杂草”,由于不同植物种间杂交能力不同,外源基因转移并稳定遗传的几率受到多种因素的影响,不同作物的风险性也不同,因此必须经过长期的监测才能得出科学的结论;2)转基因作物对作物遗传多样性、物种多样性及生态系统多样性的影响:多数观点认为转基因作物会通过基因漂移,外来基因在农家品种或野生种中固定及其竞争优势导致遗传多样性减少乃至丧失,也有观点认为,从长远看,转基因作物将会增加作物的生产力,从而少用农田,少用农药,有助于保护生物多样性;转基因作物对物种多样性的影响正反两方面的报道均有,有待进一步的研究,尤其是研究分析方法亟待规范;转基因作物对生态系统多样性的影响仍在研究争论之中,尚无定论。     

Occasional loss of expression of phosphinothricin tolerance in sexual offspring of transgenic oilseed rape (Brassica napus L.)

The commercial and economic value of genetically modified crops is determined by a predictable, consistent and stable transmission and expression of the transgenes in successive generations. No gene inactivation is expected after selfings or crosses with non-transformed plants of homozygous transgenic oilseed rape plants if the expression of the transgene in homozygous or hemizygous nature in such plants is stable. The segregation ratios of phosphinothricin (PPT) tolerance in successive generations of selfings and mutual crosses of a few independent transgenic PPT-tolerant oilseed rape plants indicated a dominant, monogenic inheritance. In within-variety and between-variety crosses no transgene inactivation was observed. However, after selfings and backcrosses with non-transgenic oilseed rape infrequent loss of the expression of the PPT tolerance transgene was observed independent from its homozygous or hemizygous nature. Molecular analysis of PPT-susceptible plants showed that the loss of expression was due to gene inactivation and not to the absence of the transgene. Methylation and co-suppression are mechanisms that might cause reduced or even loss of expression of the transgene in later generations. The implications of this observation for seed multiplication of varieties and breeding activities with transgenic oilseed rape are discussed. This revised version was published online in July 2006 with corrections to the Cover Date.     

The release of transgenic plants from containment, and the move towards their widespread use in agriculture

Summary The use of transgenic plants in breeding makes it possible to utilise a wide variety of novel genes from unrelated plants, microbes and animals. Because of the diversity of genes that have now become available for modifying crop plants, it is agreed internationally that there should be a risk assessment before transgenic plants are grown outside the laboratory or glasshouse. Various aspects are considered in a risk assessment including any non-target effects of the transgene, changes in plant persistence and invasiveness, and the possibility of movement of the transgenes to wild populations by cross pollination. It is generally argued that the need for risk assessment and regulation should be determined by an analysis of certain products of transformation, rather than a risk assessment being required for all plants modified by the process of transformation. A possible consequence of considering the product only, however, could be that some of the products of conventional breeding may need to be assessed by the risk assessment procedures developed for transgenic plants. There are discussions with interest groups on the use of transgenic plants in the environment and in food products. It is likely that some form of labelling will be required for certain foods containing ethically-sensitive genes. There is little doubt that transgenic plants will make a significant contribution to agriculture in the coming decades. Developments in the patenting of genes, release regulations, food labelling, consumer reaction etc., will influence the rate of progress and should be considered in the strategic planning of plant breeding programmes.     

甘蔗转基因的研究进展

甘蔗作为重要的糖料作物和遗传转化高安全级别的经济作物,其遗传转化的研究一直倍受重视。目前应用于甘蔗中的转基因技术主要是基因枪介导法和农杆菌介导法。近年来甘蔗的转基因研究主要以新型抗病虫、抗寒、抗旱和高糖以及作为生物反应器生产高附加值产品为主,同时在转基因生物的检测研究中也取得了较大的进展。但是甘蔗的转基因还是受到一些因素的限制,转化效率低、基因表达效率不高、转基因安全性研究少以及可能存在的法律和社会问题等。建立多个甘蔗主栽品种高效、快速、稳定的遗传转化体系,研究安全型系统,开展大片段基因、多基因转化,实现基因的时空控制表达等将是甘蔗转基因技术的新的发展趋势。     

Identifying indicator species for post-release monitoring of genetically modified,herbicide resistant crops

In Europe, regulations for release and placing-on-the-market of genetically modified (GM) crops require post-release monitoring of their impact on the environment. Monitoring potential adverse effects of GM crops includes direct effects as well as indirect effects, e.g. GM crop specific changes in land and pest management. Currently, there is a gap in the pre-release risk assessments conducted for regulatory approval of GM herbicide resistant (HR) crops. Since the relevant non-selective herbicides have been registered many years ago, in current dossiers requesting regulatory approval of GM HR crops, the environmental impacts of the corresponding non-selective herbicides are either entirely omitted or the applicant simply refers to the eco-toxicological safety assessments conducted for its original pesticide approval that do not address environmental issues arising in conjunction with the cultivation of GM HR crops. Since the ‘Farm-scale Evaluations’, it is clear that consequences for farmland biodiversity can be expected. The objective of this project was to identify relevant indicator species for the long-term impact of GM HR maize cultivation and the application of their corresponding non-selective herbicides, glyphosate and glufosinate. In this article, we describe the outcome of a modified Event Tree Analysis, essentially a funnel-like procedure allowing to reduce the large number of potentially affected non-target species to those with greatest ecological relevance and highest risk to be adversely affected based on a number of ecological criteria. This procedure allowed us to identify a total of 21 weed-Lepidoptera associations that we proposed for post release monitoring of GM HR maize in Germany.     

GAFP基因导入对彩色棉抗病性和农艺及经济性状的影响

通过对转GAFP基因品系ZB-1-31、ZB-1-49、LB-7-8与原受体品种新彩棉1号、绿9903进行了抗枯、黄萎病性的对比和农艺、经济性状方面的对比研究,结果表明,3个转GAFP基因品系对枯、黄萎病的抗性显著高于受体品种,生育期有所提前,产量均明显高于受体品种,纤维品质也有不同程度的提高。     

转基因作物安全研究进展及南繁转基因生物安全管理对策

本文从生物多样性、生态系统和人畜安全三方面概述了转基因作物生物安全研究进展,并介绍了全世界转基因生物安全评价原则与方法和我国转基因作物安全管理法规.表明转基因作物存在潜在的生物安全风险,全世界正在积极构建转基因生物安全管理体系.通过对我国南繁转基因环境安全问题和情况进行分析,认为南繁地区有成为染缸的潜在威胁.为保护南繁...     

Environmental risks of genetic engineering

Summary Before release into commerce, genetically engineered organisms are first assessed for possible risks, including risks to the environment. The present paper first identifies the environmental risks recognized by regulators, and reviews the parameters considered predictive of risk. Recent field-scale studies suggest opportunities for improvement of the environmental risk assessment process. Risks unique to genetically engineered crops – if any – could pertain to the specific traits chosen for commercialization and to unintended trait expression caused by the process of transgene insertion itself. Both the standard against which to compare genetically engineered traits and the scale of exposure need to be considered when assessing environmental impact. Evidence of environmental risk in the recognized areas of weediness on agricultural land, invasiveness of unmanaged systems, and non-target impacts from Bacillus thuringiensis (Bt) maize is presented. Targeted, statistically sound, rigorously conducted, multi-trophic studies analogous to the Field Scale Evaluation trials recently completed in the UK are needed to clarify the many questions which remain unanswered.     

Overwintering of genetically modified sugar beet, Beta vulgaris L. subsp. vulgaris, as a source for dispersal of transgenic pollen

The potential impact of transgenic crops on community ecology will depend on the distribution and establishment of the new transgenic traits, on the sexual transfer of their new genes to the environment (Bartsch &; Pohl-Orf, 1996) and on the potential ecological impact of the transgenic trait. Flowering and pollen dispersal is important for outcrossing of the genetically engineered trait. For a biennial plant, like the cultivars of Beta vulgaris L., overwintering is normally necessary to become generative and to produce pollen and seeds (Abe et al., 1997), which usually does not happen with sugar beet as a field crop harvested in autumn (Longden 1989). The starting point for the project was a transgenic sugar beet, Beta vulgaris L. subsp. vulgaris (Lange et al., 1998), with rhizomania and herbicide ( Basta®, Liberty®) resistance. Cold tolerance is one of the most important factors for survival of sugar beet in Central- and North-Europe. Among other ways, spreading of transgenic traits into weed beet (Boudry et al., 1993) or wild beet can occur if genetically engineered – biennial – plants survive the winter, flower in spring and spread their pollen. Field experiments were performed with transgenic breeding lines and their hybrids, transgenic and non-transgenic hybrids with Swiss chard and three conventional beet cultivars to evaluate winter survival rates at seven different field sites. We could show that survival of sugar beet – transgenic as well as conventional ones – in Germany and at the Dutch border is possible. Survival rates were well correlated with temperature data and were unexpectedly high. Differences between sugar beet hybrids and breeding lines could be detected but not within different breeding lines or hybrids. There were no differences detectable between transgenic and non-transgenic plants. The data are crucial for the risk assessment of the release of transgenic sugar beet and are the basis for further experiments towards outcrossing and establishment.     

植物抗病毒侵染的分子机制

植物病毒病是一类严重危害农作物生产的重要病害。已报道的植物抗病毒基因主要在抑制病毒增殖和阻止病毒扩散中起作用。病毒的复制涉及自身的编码蛋白及其与寄主蛋白间的互作, 参与病毒复制的寄主蛋白很多, 如真核翻译起始因子eIF4E和eIF4G, 植物的内膜系统等, 相关蛋白的功能丧失或构型改变可阻滞病毒的复制;此外, 植物细胞内的硫氧还蛋白可调节细胞的氧化还原状态, 进而阻断病毒的增殖。病毒在植物体内的扩散包括胞间移动和长距离迁移, 植物抗病蛋白(R蛋白)通过识别病毒的无毒因子(Avr)促发防御反应, 诱导过敏性坏死, 限制病毒在细胞间的扩散, 编码这类抗病蛋白的基因主要为TIR-NBS-LRR和CC-NBS-LRR。病毒的长距离迁移涉及的因素很多, 目前仅发现韧皮部的RTM蛋白可能以多聚蛋白的形式抵制病毒的长距离移动。另外, RNA沉默也是植物抵制病毒侵染的免疫反应机制。本文旨在综述植物的各种抗病毒机制和相关的抗病基因, 并探讨分子标记辅助选择(marker-assisted selection, MAS), 定向诱导基因组局部突变(targeting induced local lesions in genomes, TILLING)和转基因等生物技术在抗病改良中的应用前景。     

转SCK基因水稻田间抗虫性评价

用转sck基因的M81、R527、C162、秋B、D62B系列水稻及对照,进行了田间抗三化螟性调查和评价。结果表明：与对照相比,用枯心指数和白穗指数作评价指标,所有转sck水稻在早造和晚造大田种植中对三化螟抗性均明显提高,达中抗以上,而对照均为中感或感级。而且评价指标枯心指数和白穗指数之间,早造和晚造之间所得结果较一致,在以后抗螟虫鉴定中可考虑选用枯心指数或白穗指数作指标,在早造或晚造进行。     

利用叶蝉评价转基因玉米对非靶标生物的风险

转基因抗虫作物应用是替代杀虫剂的重要途径。但转基因作物可能存负面效应,特别是对非靶标生物的影响及害虫产生抗性。采用直接和陷阱调查结合方法,连续多年对转Cry1Ac 抗虫基因玉米实验田叶蝉和主要节肢动物数量动态跟踪调查,结果发现叶蝉是玉米田优势非靶标生物种群之一,中期发生比例占刺吸式害虫的28.9%~43.1%,且种群动态与主要非靶标节肢动物群落动态基本一致。又鉴于叶蝉直接刺吸接触到Bt 杀虫蛋白,可直接反映转基因作物对非靶标生物的安全性,因此可利用叶蝉作为转基因抗虫玉米安全性评价指示生物来评价转基因玉米对非靶标生物群落风险,田间测试结果证明此方法是可行的。     

Direct comparison of pollen-mediated movement of native and engineered genes

Despite the commercial approval of twenty-five transgenic crops in the U.S. as of mid-1996, concern is still being expressed regarding the potential risks associated with genetically engineered crops. One recurring issue is the possibility of pollen-mediated escape of engineered genes into populations of crop wild relatives. To address this concern, the scientific community has depended on literature on pollen dispersal generated from non-transgenic organisms. Utilization of this information requires the assumption that the pollen mediated movement of native and transgenes is the same. To test the validity of this assumption, we directly compared the pollen-mediated gene movement of native and engineered marker genes using melon plants (Cucumis melo L.) expressing dominant morphological and transgenic traits. Movement into both contiguous border plots and non-contiguous satellite plots were monitored. Dispersal of the native gene and transgene into the satellite plots was identical. Dispersal of the two traits into the plot borders was nearly identical. Of the nearly 4600 seedlings screened for both morphological (presence of green vs. virescent cotyledons) and transgene movement (presence of NPT II protein by ELISA), in no case was the NPT II gene observed in the absence of green cotyledons. However, 39 seedlings were green but did not express NPT II as measured by ELISA. PCR analysis revealed transgene inactivation as a cause of the NPT II ELISA-seedlings. This revised version was published online in July 2006 with corrections to the Cover Date.     

Counteracting virulence mechanisms of grain legume pathogens

Summary Grain legumes, in common with all other plants, are subject to biotic constraints of which pathogens form an important group. They are variable in type, number, space and time and, most insidiously, in genetic constitution. Consequently, resistance in the plant to a given pathogen may be quickly nullified by genetic alteration of the pathogen, particularly where this is conferred by a single resistance gene. The products of such resistance genes usually recognise, directly or indirectly, a component of the pathogen, which is encoded by a corresponding avirulence gene. Thus resistance and avirulence genes are specific and complementary and the arrangement is referred to as a gene-for-gene relationship. It follows that alteration of the avirulence gene of the pathogen to give a product that is no longer recognised by the product of the resistance gene of the plant gives rise to a susceptible reaction. A possible solution to this problem is to pyramid several resistance genes, a procedure now facilitated by the techniques of genetic modification. In other interactions genes that reduce susceptibility rather than confer complete resistance have been found and in some cases the loci (quantitative trait loci) responsible have been mapped to specific regions of particular chromosomes. The mechanisms by which these genes limit the virulence of the pathogen are generally unknown. However, by gaining an understanding of the fundamental properties of a pathogen that are necessary for pathogenicity or virulence it may be possible to counteract them. Candidates for such properties are toxins, enzymes and mechanisms that interfere with constitutive or active defence of the plant. Reciprocally, understanding the properties of the plant that confer susceptibility may allow selection of germplasm that lacks such properties. Among the candidates here are germination stimulants of pathogen propagules and signals that promote the formation of infection structures.     

转基因玉米转化体特异性寡核苷酸芯片测试方法的研制

根据7种转基因玉米Bt11、Bt176、Mon810、Mon863、TC1507、GA21和NK603的重组DNA结构,利用Primer premier 5.0引物设计软件分别设计转化体特异性引物,优化PCR反应条件,对引物特异性及灵敏度进行验证。结果表明,所设计引物特异性强,灵敏度达0.1%。在植物基因组与外源基因的连接区域,利用Primer premier 5.0和Oligo 6.0软件分别设计和筛选7种转基因玉米40mer oligo转化体特异性探针,制备转基因玉米的寡核苷酸芯片,并对芯片检测的特异性、重复性及检测灵敏度进行检验。证明所设计探针特异性强,灵敏度达0.01%。与PCR检测方法相比,基因芯片检测法灵敏度高、特异性强,可有效检测及鉴定多种转基因玉米,大大提高检测的准确率和效率。     

利用微滴式数字PCR技术分析转基因玉米抗除草剂标记基因EPSP拷贝数

基于微滴式数字PCR(Droplet digital PCR,ddPCR)平台,以转基因玉米为例,建立了转基因作物(Genetically modified crops,GM crops)外源基因拷贝数分析方法,对待测样品进行了快速鉴定,并从T_0转基因玉米株系中鉴定出多个单拷贝单株。对该方法与实时荧光定量PCR(Quantitative real-time PCR,qRT-PCR)方法在分析结果的准确性方面进行了比较,从试验数据可以看出,2种检测方法的结果比较一致,单拷贝检测结果高度一致;但是ddPCR试验操作更加简便,试验结果可重复性强,试验数据更加准确可靠。研究表明,ddPCR方法是一种更加便捷、快速和准确的外源基因拷贝数分析新方法,基于其在准确性和灵敏度方面的显著优势,将会在转基因作物的外源基因拷贝数分析中得到广泛的应用。