转Bt基因抗虫欧洲黑杨安全性评价研究
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摘要
植物基因工程在过去短短二十年内取得了飞速的发展。转基因植物大规模商业化种植中的生物安全性问题,由于关系到生物及自然环境的保护和人体健康,一直受到广泛的关注。本文的工作包括二个部分,第一部分是转基因欧洲黑杨杂交后代Bt基因表达特性的研究。利用PCR检测了转基因欧洲黑杨当代和杂交后代群体,外源Bt基因在7年生的转基因欧洲黑杨14个无性系中稳定表达,Bt基因在杂交后代中分离比例为1:1。转基因欧洲黑杨作母本进行杂交时有落花、落果现象,可能是基因改造影响了转基因的某些生理功能。第二部分是B.t.转基因杨树对杨树昆虫群落结构的影响。通过研究表明小面积的新疆转基因杨树和非转基因杨树之间群落结构和多样性差别不大;而对100亩以上的转基因杨树纯林和1:1的转基因和非转基因杨树混交林中昆虫和蜘蛛群落结构的比较研究说明,Bt转基因杨树会改变其昆虫群落结构,在转基因纯林中杨叶蜂(Pristiphora benjingensis Zhou and Zhang)为优势种,而在混交林中杨扇舟蛾(Clostera anachoreta (Fabricus))为优势种,这说明Bt转基因杨树在抑制鳞翅目昆虫的同时,有利于其他食叶昆虫。为了避免其它非鳞翅目食叶害虫的大量危害,在转基因时应采用对非鳞翅目食叶害虫有抗性的种类或品系。转基因纯林和混交林的叶片被害率分别为11.26%和18.48%,转基因纯林抑制目标食叶害虫的作用较好,但与混交林之间的差异不显著,两种林分的被害率都在不成灾的水平。转基因纯林的多样性和均匀性都较混交林高,有利于系统的稳定性,但转基因混交林中瓢虫的数量明显比转基因纯林高,分别为0.21头/枝和0.021头/枝,相差10倍。蜘蛛的数量是纯林中较多,分别为0.125头/枝和0.0625头/枝,相差1倍。转基因杨树对非目标昆虫和天敌的影响还需进行进一步详细的研究。安全性评价工作起始的时间较短,资料积累不足,在研究的理论和技术方面都有待于发展。
Plant engineering developed rapidly during recent 20 years.Transgenictechnology has been effectively used for the improvement of plant cuntivars.A greatamount of transgenic plant have been released for commercial use in recentyears.Transgenic plant is different from the traditional one in that the formercontains foreign genes which leads to public concern.The environmental safety offoreign genes in transgenic plants has been disscussed in this paper.The safety oftransgenic plant commercialized in a large scale received much concern since it hasa close relationship with creatures and environment protection and human health.Thegoal of this paper is to provide a scientific Framework to assess potentialenvironment risks associated with transgenic woody biomass crops. Trees differentfrom most agricultural crops in that they persist in the landscape for long periodsof time.This different timeframe not only increase the probability that any one treecrop will be subjected to a much wider range of stress conditions,but also that anystress-induced side-effect of GM technology will be much harder to detect andaddress.Meanwhile,remote location and less intense management regimes mean limitedopportunities for monitoring ,control and enforcement of regulations while makingearly detection of unanticipated problems highly unlikely.The work in this paperincludes three aspects.
The first is about the literature.The literature pertaining to environmenteffects of genetically engineered insect resistance crops is briefly reviewed andthe relevance of this information to transgenic woody biomass crops isdiscussed ,particularly in regard to how ecological and genetic factors influencerisk.
The second is to detect the stability of foreign gene in Bt poplar,thetransmission of the transgene to the tree progenies and develop a protocol ofdetection Bt gene in tree sample by means of PCR technique.PCR is adopted to analyzethe expression of insectidal protein gene Bt in the transgenic poplar plants.The
results show that the expression of the protein gene was stable at the seven yearof the plantation.Several flowers were collected from a male transgenic tree andpollinated in controlled conditions.Progeny seeds were germinated and grown invitro.Analyses revealed that segregation appreas to follow Mendal's rule as 67 outof 120 individual tested exhibit transgene expression in accordance with the awaited1:1 segregation.A stable expression of the Bt gene was detected in both the parentand F1 progenies.These physiological differences,maybe caused by gene transfer andtissues culture,could induce changes in secondary metabolism of Bt poplar and thenbring ecological effects to environment.This kind of effect should also be animportant aspects of ecological risk assessment of transgenic plants release intoenvironment.The application of PCR detection in safety assessment is discussed.The thrid level of this work is to reveal that there was no much insect and spidercommunity difference between transgenic and non-transgenic poplar trees of the smallplanted area while in large planted area obvious difference of that was observed.Through the comparison of insect and spider community structures between puretransgenic poplar stand and 1:1 mixed stand of transgenic and non-transgenic poplars,it is clear that Bt transgenic poplar can influence the insect community structure.In pure transgenic poplar stand, dominant species is poplar sawfly (Pristiphorabenjingensis Zhou et Zhang) while in mixed poplar stand is poplar prominent (Closteraanachoreta (Fabricus)). It demonstrates that transgenic poplar can suppress theLepidopteran species and at same time may favorable to other defoliators. To reducethe risky of being damaged by other non-lepidoptera defoliators resistant speciesto this kind of pest species should be used when B.t. gene is transferred. The damagerate of pure transgenic poplar stand and mixed transgenic poplar stand were 11.26%and18.48% respectively, the former was better but no significant difference. Thediversity and uniformity of pure transgenic poplar stand were higher than that ofMixed transgenic poplars stand which leads to higher stability. However the densityof lady beetles was 10 times higher in mixed transgenic poplar stand than puretransgenic poplar stand, averagely 0.21 per twig and 0.021per twig respectivelywhile the density of spiders in pure transgenic poplar stand was higher than that
in mixed transgenic poplar stand, averagely 0.125 per twig and 0.0625 per twigrespectively. Further study is needed in the effect of transgenic poplars to thenon-target insects and natural enemies.Because the subject arise only less than 30yaer and the data are not rich enough,the theory and technique of safety assessmentneed much improvement.This work applies technique of molecular biology and community ecology ,combinesthe phenomena in field and completes a comprehensive safety assessment of transgenicBt poplar.
Plant engineering developed rapidly during recent 20 years.Transgenictechnology has been effectively used for the improvement of plant cuntivars.A greatamount of transgenic plant have been released for commercial use in recentyears.Transgenic plant is different from the traditional one in that the formercontains foreign genes which leads to public concern.The environmental safety offoreign genes in transgenic plants has been disscussed in this paper.The safety oftransgenic plant commercialized in a large scale received much concern since it hasa close relationship with creatures and environment protection and human health.Thegoal of this paper is to provide a scientific Framework to assess potentialenvironment risks associated with transgenic woody biomass crops. Trees differentfrom most agricultural crops in that they persist in the landscape for long periodsof time.This different timeframe not only increase the probability that any one treecrop will be subjected to a much wider range of stress conditions,but also that anystress-induced side-effect of GM technology will be much harder to detect andaddress.Meanwhile,remote location and less intense management regimes mean limitedopportunities for monitoring ,control and enforcement of regulations while makingearly detection of unanticipated problems highly unlikely.The work in this paperincludes three aspects.
The first is about the literature.The literature pertaining to environmenteffects of genetically engineered insect resistance crops is briefly reviewed andthe relevance of this information to transgenic woody biomass crops isdiscussed ,particularly in regard to how ecological and genetic factors influencerisk.
The second is to detect the stability of foreign gene in Bt poplar,thetransmission of the transgene to the tree progenies and develop a protocol ofdetection Bt gene in tree sample by means of PCR technique.PCR is adopted to analyzethe expression of insectidal protein gene Bt in the transgenic poplar plants.The
results show that the expression of the protein gene was stable at the seven yearof the plantation.Several flowers were collected from a male transgenic tree andpollinated in controlled conditions.Progeny seeds were germinated and grown invitro.Analyses revealed that segregation appreas to follow Mendal's rule as 67 outof 120 individual tested exhibit transgene expression in accordance with the awaited1:1 segregation.A stable expression of the Bt gene was detected in both the parentand F1 progenies.These physiological differences,maybe caused by gene transfer andtissues culture,could induce changes in secondary metabolism of Bt poplar and thenbring ecological effects to environment.This kind of effect should also be animportant aspects of ecological risk assessment of transgenic plants release intoenvironment.The application of PCR detection in safety assessment is discussed.The thrid level of this work is to reveal that there was no much insect and spidercommunity difference between transgenic and non-transgenic poplar trees of the smallplanted area while in large planted area obvious difference of that was observed.Through the comparison of insect and spider community structures between puretransgenic poplar stand and 1:1 mixed stand of transgenic and non-transgenic poplars,it is clear that Bt transgenic poplar can influence the insect community structure.In pure transgenic poplar stand, dominant species is poplar sawfly (Pristiphorabenjingensis Zhou et Zhang) while in mixed poplar stand is poplar prominent (Closteraanachoreta (Fabricus)). It demonstrates that transgenic poplar can suppress theLepidopteran species and at same time may favorable to other defoliators. To reducethe risky of being damaged by other non-lepidoptera defoliators resistant speciesto this kind of pest species should be used when B.t. gene is transferred. The damagerate of pure transgenic poplar stand and mixed transgenic poplar stand were 11.26%and18.48% respectively, the former was better but no significant difference. Thediversity and uniformity of pure transgenic poplar stand were higher than that ofMixed transgenic poplars stand which leads to higher stability. However the densityof lady beetles was 10 times higher in mixed transgenic poplar stand than puretransgenic poplar stand, averagely 0.21 per twig and 0.021per twig respectivelywhile the density of spiders in pure transgenic poplar stand was higher than that
in mixed transgenic poplar stand, averagely 0.125 per twig and 0.0625 per twigrespectively. Further study is needed in the effect of transgenic poplars to thenon-target insects and natural enemies.Because the subject arise only less than 30yaer and the data are not rich enough,the theory and technique of safety assessmentneed much improvement.This work applies technique of molecular biology and community ecology ,combinesthe phenomena in field and completes a comprehensive safety assessment of transgenicBt poplar.
引文
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