适合度分析对转基因逃逸潜在环境风险评价的意义北大核心CSCD

转基因作物的全球大规模种植引起了全世界的广泛关注甚至争议。经过遗传改良并具有自然选择优势的转基因作物进入商品化种植,可能将带来环境生物安全的顾虑。在这些生物安全的顾虑中,转基因通过花粉介导的基因漂移向栽培作物的野生近缘种逃逸及其导致的潜在环境风险,就是世人最为关注的环境生物安全问题之一。包括中国在内的许多国家,在转基因作物进行商品化生产之前都必须对转基因逃逸及其带来的潜在环境风险进行严格评价。按照风险评价的框架,转基因向野生近缘种逃逸及其带来潜在环境风险的评价包括3个连续的步骤:1)检测转基因漂移到作物野生近缘种的频率;2)分析转基因在野生近缘种中的表达;3)确定转基因对野生近缘种群体适合度和进化潜力的影响。大量基因漂移的研究结果已表明,转基因通过基因漂移向栽培作物邻近的野生近缘种群体逃逸难以避免,而转基因也会在作物的野生近缘种群体中正常表达。因此分析和评价转基因为野生近缘种带来的适合度效应,对于转基因逃逸及其环境风险的评价至关重要。对适合度的概念及其进化意义进行介绍,并对如何利用转基因的适合度效应分析转基因逃逸的环境风险,以及对此类环境风险进行研究和评价的具体方法予以介绍。上述知识和方法的掌握将有助于人们对转基因作物环境生物安全及其评价的全面理解。     

适合度分析对转基因逃逸潜在环境风险评价的意义

转基因作物的全球大规模种植引起了全世界的广泛关注甚至争议。经过遗传改良并具有自然选择优势的转基因作物进入商品化种植,可能将带来环境生物安全的顾虑。在这些生物安全的顾虑中,转基因通过花粉介导的基因漂移向栽培作物的野生近缘种逃逸及其导致的潜在环境风险,就是世人最为关注的环境生物安全问题之一。包括中国在内的许多国家,在转基因作物进行商品化生产之前都必须对转基因逃逸及其带来的潜在环境风险进行严格评价。按照风险评价的框架,转基因向野生近缘种逃逸及其带来潜在环境风险的评价包括3个连续的步骤:1)检测转基因漂移到作物野生近缘种的频率;2)分析转基因在野生近缘种中的表达;3)确定转基因对野生近缘种群体适合度和进化潜力的影响。大量基因漂移的研究结果已表明,转基因通过基因漂移向栽培作物邻近的野生近缘种群体逃逸难以避免,而转基因也会在作物的野生近缘种群体中正常表达。因此分析和评价转基因为野生近缘种带来的适合度效应,对于转基因逃逸及其环境风险的评价至关重要。对适合度的概念及其进化意义进行介绍,并对如何利用转基因的适合度效应分析转基因逃逸的环境风险,以及对此类环境风险进行研究和评价的具体方法予以介绍。上述知识和方法的掌握将有助于人们对转基因作物环境生物安全及其评价的全面理解。     

转基因抗除草剂油菜对近缘作物的基因漂移

以转基因抗除草剂油菜 Q3和 HCN- 19为花粉供体材料 ,油菜近缘作物为花粉受体材料 ,在自然授粉条件下研究甘蓝型油菜与芸薹属近缘作物间的基因漂移频率。结果表明 ,油菜对芸薹属 6个种甘蓝、黑芥、埃芥、芥菜型油菜、白菜型油菜和甘蓝型油菜的基因漂移率分别为 0、0 .0 2 4 %～ 0 .2 4 3%、 0 .0 2 8%～ 0 .0 92 %、 0 .10 9%～ 0 .95 1%、 0 .4 79%～ 0 .879%、 1.2 5 2 %～2 .191%。且基因漂移频率受多种因素影响 ,其中与杂交亲和性、花期同步率、种植面积等高度相关。通过花粉将抗除草剂基因漂移给近缘作物 ,油菜是需要特别关注的作物     

转基因栽培稻基因漂移是否会带来环境生物安全影响?

转基因通过基因漂移可以渐渗到作物的野生近缘种,由此而导致的环境风险是全球广泛关注的生物安全问题.有3个关键因素可以决定环境风险的程度:特定空间距离的转基因漂移频率,转基因在野生近缘种中的表达水平,以及转基因为野生近缘种群体带来的适合度效应.本文将根据现有研究结果,从上述3方面对转基因漂移到非转基因栽培稻、杂草稻和野生稻造成的潜在环境影响进行回顾.栽培稻品种之间的基因漂移频率很低,可以通过空间隔离或其他方法使其降低到可忽略的水平.在共同分布的环境中,栽培稻基因(包括转基因)向杂草稻和野生稻的漂移不可避免.尽管抗虫转基因(Bt或Bt/CpTI)在栽培稻和野生近缘种杂交后代中可以正常表达,但由于在低虫压环境中,抗虫转基因不会明显改变野生近缘种的适合度,抗虫转基因漂移所造成的环境影响十分有限.因此对基因漂移而言,抗虫转基因栽培稻的商品化种植应该比较安全.然而,抗除草剂转基因渐渗到杂草稻或野生稻会改变群体的适合度,可能会引起不可预测的环境后果.     

转基因植物的环境生物安全：转基因逃逸及其潜在生态风险的研究和评价

转基因作物的商品化生产和大规模环境释放在带来巨大利益的同时,也引起了全球对其生物安全问题的广泛关注和争议,其中转基因通过花粉介导的基因漂移逃逸到非转基因作物及其野生近缘种,进而导致的潜在环境和生态风险就是备受争议的生物安全问题之一。转基因植物的环境生物安全涉及两方面关键问题：如何科学评价转基因植物商品化种植以后带来的环境和生态影响;如何利用环境生物安全的研究成果来制定科学有效的风险监测和管理措施。对转基因逃逸及其潜在生态风险的科学评价应包括三个重要环节：（1）检测转基因的逃逸的频率;（2）检测转基因逃逸后的表达和遗传规律;（3）确定逃逸后的转基因对野生近缘种群体适合度的影响及其进化潜力,本文将围绕对转基因逃逸及其潜在环境风险的科学评价,以转基因水稻为案例来对转基因逃逸带来生态影响的研究好评价的进展进行简要介绍,并对目前依据风险评价研究成果制定的各种管理策略进行了讨论。只有提高对转基因生物环境安全研究和评价的水平,并制定有效的风险监测和管理措施,才能为我国转基因技术的发展和转基因产品的商品化应用保驾护航。     

杂交-渐渗进化理论在转基因逃逸及其环境风险评价和研究中的意义

转基因作物的商品化生产和大规模环境释放, 引起了全球对生物安全问题的广泛关注和争议, 其中转基因通过花粉介导的基因漂移逃逸到非转基因作物及其野生近缘种, 进而带来不同类型的环境风险就是备受争议的生物安全问题之一。有效的生物安全评价和研究能够为转基因作物的安全持久利用保驾护航。按照风险评价的原则, 对于转基因逃逸及其潜在环境风险的评价应包括两个重要步骤: (1)检测转基因向野生近缘种(包括杂草类型)群体逃逸的频率; (2)确定逃逸后的转基因能否通过遗传渐渗在野生近缘种群体中存留和扩散。杂交－渐渗是进化生物学中非常重要的科学命题和普遍的自然现象, 杂交－渐渗的进化理论与转基因逃逸及其潜在环境风险的研究和评价有密切的关系。杂交－渐渗过程往往导致物种形成、适应性进化和自然群体的濒危与灭绝, 这是因为在杂交－渐渗过程中, 不同的机制如遗传同化作用、群体湮没效应以及群体的选择性剔除效应等都会在很大程度上影响群体的进化过程。转基因通过杂交－渐渗进入野生群体, 使这一过程更加复杂化。如果转基因能提高群体的适合度, 则更有利于其渐渗速率, 从而在群体中迅速扩散并带来一定的生态后果。杂交－渐渗的进化理论和思想将有益于指导转基因逃逸及其潜在环境风险的研究和评价。     

转基因植物与近缘种之间基因流的研究进展

随着转基因植物的大面积种植,转基因植物的生态风险受到广泛关注,其中主要的风险是转基因植物与近缘物种之间的基因流及其影响。本文综述了目前商业化种植的转基因作物油菜、棉花、玉米和大豆,以及未商业化种植的水稻、小麦的基因流研究进展;分析了不同转基因作物与其近缘种之间发生基因流的频率和最远发生距离;介绍了降低基因流发生的方法。基因流频率受物种亲缘关系、花期重叠时间、风速风向等因素的影响,最远发生距离受气候条件、传粉媒介、地理条件等因素的影响。转基因作物与其近缘种之间的基因流频率与距花粉源的距离呈负相关关系(y=-0.59x-0.46,R²=0.25,P<0.01),亲缘关系近的基因流频率高。为了降低转基因植物与其近缘物种之间的基因流风险,建议采取“分区管理”的策略,并加强基因流发生之后的生态风险评价研究。     

防止转基因作物释放引发“超级杂草”产生的若干对策

转基因作物释放可能导致潜在的生态风险性,其中的一个重要方面是通过花粉的传播将某些转基因(主要是抗除草剂基因)漂入野生近缘种或近缘杂草而引发难以控制的“超级杂草”的产生。本文讨论了防止“超级杂草”产生的若干对策,包括物理隔离、转基因遗传调控、雄性不育性与无融合生殖机制的利用、将转基因定位于同当地杂草不亲和的基因组和叶绿体或线粒体基因组等。     

防止转基因作物释放引发“超级杂草”产生的若干对策

转基因作物释放可能导致潜在的生态风险性,其中的一个重要方面是通过花粉的传播将某些转基因（主要是抗除草剂基因）漂入野生近缘种或近缘杂草而引发难以控制的“超级杂草”的产生。本文讨论了防止“超级杂草”产生的若干对策,包括物理隔离、转基因遗传调控、雄性不育性与无融合生殖机制的利用、将转基因定位于同当地杂草不亲和的基因组和叶绿体或线粒体基因组等。     

转基因抗除草剂油菜对十字花科杂草的基因漂移

以转基因抗除草剂油菜Q3为花粉供体材料,油菜远缘杂草为花粉受体材料,在自然传粉和人工辅助授粉条件下研究甘蓝型油菜与十字花科杂草间的基因漂移频率。结果表明,以转基因油菜为父本,十字花科杂草荠菜、碎米荠、播娘蒿、诸葛菜、风花菜、遏蓝菜和菜为母本,杂交高度不亲和,基因漂移率为0 % ,无生态风险,但对野芥菜的基因漂移率高达0 .885 %。野芥菜是我国大部分地区的常见杂草,种类繁多,分布范围广,大面积种植转基因抗除草剂油菜对野生芥菜的基因污染应引起高度重视。     

Gene Flow,Risk Assessment and the Environmental Release of Transgenic Plants

The release of genetically modified plants is governed by regulations that aim to provide an assessment of potential impact on the environment. One of the most important components of this risk assessment is an evaluation of the probability of gene flow. In this review, we provide an overview of the current literature on gene flow from transgenic plants, providing a framework of issues for those considering the release of a transgenic plant into the environment. For some plants gene flow from transgenic crops is well documented, and this information is discussed in detail in this review. Mechanisms of gene flow vary from plant species to plant species and range from the possibility of asexual propagation, short- or long-distance pollen dispersal mediated by insects or wind and seed dispersal. Volunteer populations of transgenic plants may occur where seed is inadvertently spread during harvest or commercial distribution. If there are wild populations related to the transgenic crop then hybridization and eventually introgression in the wild may occur, as it has for herbicide resistant transgenic oilseed rape (Brassica napus). Tools to measure the amount of gene flow, experimental data measuring the distance of pollen dispersal, and experiments measuring hybridization and seed survivability are discussed in this review. The various methods that have been proposed to prevent gene flow from genetically modified plants are also described. The current “transgenic traits” in the major crops confer resistance to herbicides and certain insects. Such traits could confer a selective advantage (an increase in fitness) in wild plant populations in some circumstances, were gene flow to occur. However, there is ample evidence that gene flow from crops to related wild species occurred before the development of transgenic crops and this should be taken into account in the risk assessment process.     

Spontaneous hybridizations between oilseed rape and wild radish

The occurence of spontaneous hybridization between Brassica napus (oilseed rape) and Raphanus raphanistrum (wild radish) was investigated under different density conditions in cages and open-field experiments. Hybrids with wild radish as the seed parent were identified by screening for herbicide resistance belonging to rape. Small seed size and intermediate morphology were used to screen for hybrids with rape as the seed parent. Leaf isozyme patterns and flow cytometry provided confirmation of hybrids. Wild radish in an oilseed rape field produced as many as three interspecific hybrids per 100 plants. This is the first report of such a spontaneous event. The frequency of hybrids is expected to range from 0.006 to 0.2% of the total seed produced, at P = 0.05. Male-sterile oilseed rape plants surrounded by wild radish can produce up to 37 hybrids per plant. Seed production of the F₁ hybrids and their F₂ descendants was up to 0.4% and 2%, respectively, of that of wild radish. Gene escape from transgenic oilseed rape to wild related species is discussed.     

Fitness of backcross six of hybrids between transgenic oilseed rape (Brassica napus) and wild radish (Raphanus raphanistrum)

The process of introgression between a transgenic crop modified for better agronomic characters and a wild relative could lead potentially to increased weediness and adaptation to the environment of the wild species. However, the formation of hybrid and hybrid progeny could be associated with functional imbalance and low fitness, which reduces the risk of gene escape and establishment of the wild species in the field. Our work compares the fitness components of parents and different types of backcross in the sixth generation of hybrids between transgenic oilseed rape (Brassica napus, AACC, 2n = 38) resistant to the herbicide glufosinate and wild radish (Raphanus raphanistrum, RrRr, 2n = 18). The backcross with oilseed rape cytoplasm (OBC) has a fitness value 100 times lower than that of the backcross with wild radish cytoplasm (RBC). The herbicide-resistant RBC has similar growth to the susceptible RBC, but final male and female fitness values are two times lower. In turn, susceptible RBC exhibit similar fitness to the control wild radishes. The relative fitnesses of the different types are the same whether or not they grow under competitive conditions. The consequence on fitness of the chromosome location of the transgene conferring resistance and the relevance of these results to the impact of gene flow on the environment are discussed.     

抗草丁膦和抗草甘膦转基因油菜的抗性基因向野芥菜的流动

通过人工去雄授粉和田间隔行种植试验,研究了抗草丁膦和抗草甘膦转基因油菜(Brassica napus)中的bar基因和EPSPS基因向野芥菜(B. juncea var. gracilis)流动的可能性。结果表明在人工授粉的情况下,以野芥菜为母本,分别以两种转基因油菜为父本,亲和性指数都很高,达13以上,与野芥菜自交或开放授粉条件下的亲和性指数没有明显差异,说明两种转基因油菜和野芥菜的亲和性较好。经两次除草剂筛选,人工杂交获得的所有F₁对相应的除草剂都表现出了明显的抗性,且经PCR检测扩增出了各自的特异性条带,说明人工杂交获得的所有F₁都携带了相应的抗性基因。F₁的适合度研究表明,两种F₁种子萌发率和母本都没有明显差异,营养生长明显好于母本。但花粉活力和结实率明显下降,携带抗草丁膦基因F₁的花粉活力和每角果粒数分别是32.4%和0.59粒,携带抗草甘膦基因F₁的花粉活力和每角果粒数分别是35.1%和0.58粒。经两次除草剂筛选和PCR检测,表明野芥菜和抗草丁膦油菜或与抗草甘膦油菜田间隔行种植分别能产生0.02%和0.014%的携带抗性基因的F₁杂种。以上结果表明抗除草剂转基因油菜的抗性基因具有向野芥菜流动的可能性,且bar和EPSPS基因向野芥菜流动的可能性类似,但对其可能引起的环境后果需要做进一步地深入研究。     

Risk assessment of Bacillus thuringiensis in wild Brassica rapa: A field simulation of introgressed transgenes

Abstract

Brassica crops are able to hybridize with closely related wild and weedy species such as Brassica rapa and therefore these species could become recipients of transgenes from GM brassica crops, such as oilseed rape. Transgenes which protect against herbivory, such as a gene conferring the production of a Bt toxin, could increase the recipient's fitness and therefore enhance its competitiveness. We used microbial Bt and several other pesticides to exclude several guilds of herbivores and thus simulate the effect of transgene introgression on the performance of wild B. rapa. There were only minor negative effects of excluding insect herbivores on the performance of B. rapa and it appears that vertebrate herbivory has a more significant effect. The advantages and disadvantages of simulating the risks associated with GM plants are discussed. If we apply this simulation to GM risk assessment and make inferences from these findings, the implications of this for gene flow from insect-resistant transgenic plants are that it will be of little significance should it occur.     

In vivo measurements of changes in pH triggered by oxalic acid in leaf tissue of transgenic oilseed rape

Oxalic acid (OA), a non-host-specific toxin secreted by Sclerotinia sclerotiorum during pathogenesis, has been demonstrated to be a major phytotoxic and pathogenic factor. Oxalate oxidase (OXO) is an enzyme associated with the detoxification of OA, and hence the introduction of an OXO gene into oilseed rape (Brassica napus L.) to break down OA may be an alternative way of increasing the resistance of the plant to Sclerotinia sclerotiorum. In order to investigate the activation of OXO in transgenic oilseed rape, a convenient and accessible method was used to monitor changes in pH in response to stress induced by OA. The pH sensor, a platinum microcylinder electrode modified using polyaniline film, exhibited a linear response within the pH range from 3 to 7, with a Nernst response slope of 70 mV/pH at room temperature. The linear correlation coefficient was 0.9979. Changes induced by OA in the pH values of leaf tissue of different oilseed rape species from Brassica napus L. were monitored in real time in vivo using this electrode. The results clearly showed that the transgenic oilseed rape was more resistant to OA than non-transgenic oilseed rape.     

Polymorphism for interspecific hybridisation within a population of wild radish (Raphanus raphanistrum) pollinated by oilseed rape (Brassica napus)

The within-population polymorphism of wild radish (Raphanus raphanistrum) for interspecific hybridisation with two cultivars of oilseed rape (Brassica napus) was investigated by hand crossing experiments and fluorescence microscopy. Wide variability among plants was observed in the ability of oilseed rape pollen to germinate on the wild radish stigma; the frequency of pistils showing pollen tubes ranged from 0 to 1, depending on the female plant. The ratio of fertilised ovules to the total number of ovules in ovaries where pollen tubes arrived ranged from 0.02 to 0.51. Overall, the results provide evidence for the presence of different phenotypes. In 40% of the plants, pistils had no or very few pollen tubes and few fertilised ovules. In 23%, the foreign pollen tubes grew through the style towards the ovary, but had low ovule fertilisation efficiency. The remaining 37% showed a large number of pollen tubes in the style and frequent ovule fertilisation, and two plants showed no difference between foreign and conspecific pollen. With regard to post-zygotic barriers, pollen germination and ovule fertilisation represent minor barriers to interspecific hybridisation between oilseed rape and wild radish. It is suggested that the effectiveness of these barriers could be improved through plant breeding; this could reduce the risk of gene flow from transgenic oilseed rape to populations of wild relatives. Received: 15 April 2001 / Accepted: 24 May 2001     

油菜的遗传转化及抗溴苯腈转基因油菜的获得

以油菜(Brassica napus L.)的下胚轴和子叶为转化受体,建立了油菜的高效转化系统。在此基础上,将抗除草剂溴苯腈基因(bxn基因)导入油菜,获得了抗溴苯腈转基因油菜。分子检测实验证明,转基因油菜中含有bxn基因。转基因油菜可抗高达10~(-3)mol/L的溴苯腈。     

Unexpected Diversity of Feral Genetically Modified Oilseed Rape (Brassica napus L.) Despite a Cultivation and Import Ban in Switzerland

Despite cultivation and seed import bans of genetically modified (GM) oilseed rape (Brassica napus L.), feral GM plants were found growing along railway lines and in port areas at four sites in Switzerland in 2011 and 2012. All GM plants were identified as glyphosate-resistant GM event GT73 (Roundup Ready, Monsanto). The most affected sites were the Rhine port of Basel and the St. Johann freight railway station in Basel. To assess the distribution and intra- and interspecific outcrossing of GM oilseed rape in more detail, we monitored these two sites in 2013. Leaves and seed pods of feral oilseed rape plants, their possible hybridization partners and putative hybrid plants were sampled in monthly intervals and analysed for the presence of transgenes by real-time PCR. Using flow cytometry, we measured DNA contents of cell nuclei to confirm putative hybrids. In total, 2787 plants were sampled. The presence of GT73 oilseed rape could be confirmed at all previously documented sampling locations and was additionally detected at one new sampling location within the Rhine port. Furthermore, we found the glufosinate-resistant GM events MS8xRF3, MS8 and RF3 (all traded as InVigor, Bayer) at five sampling locations in the Rhine port. To our knowledge, this is the first time that feral MS8xRF3, MS8 or RF3 plants were detected in Europe. Real-time PCR analyses of seeds showed outcrossing of GT73 into two non-GM oilseed rape plants, but no outcrossing of transgenes into related wild species was observed. We found no hybrids between oilseed rape and related species. GM plants most frequently occurred at unloading sites for ships, indicating that ship cargo traffic is the main entry pathway for GM oilseed rape. In the future, it will be of major interest to determine the source of GM oilseed rape seeds.