无选择标记基因植物转化系统研究进展

在转基因植物中,将选择标记基因去掉,将提高转基因植物的食用安全性和对环境的安全性,更易为广大消费者所接受,也有利于对同一个植物品种进行多次转基因操作。科学工作者已经在建立无选择标记基因转化系统方面作了大量尝试,获得了无标记基因的转基因植物（MFTPs：Marker-Free Transgenic Plants)。本文将这方面的研究进展介绍给大家,以推动植物生物技术产业化进程。     

代谢关键酶基因作为转基因植物选择标记的研究进展

在植物基因工程中,需要用到选择标记基因以便筛选转化子。随着转基因植物的商品化,人们越来越关注其安全性问题,其中主要涉及选择标记的安全性。为了提高转基因植物的安全性,生物学家们开始寻找生物安全的选择标记。本文分别从糖代谢、氨基酸代谢、激素代谢、核苷酸代谢、蛋白质代谢等方面综述了当前以代谢关键酶基因作为转基因植物选择标记的研究进展,此类选择标记为解决转基因植物的安全性问题提供了一条有效途径。     

无选择标记转基因植物的培育

植物转基因研究中通常都要使用选择标记基因来筛选转化细胞并获得转基因植株。但是当转基因植株育成后,选择标记基因就失去了存在的意义。为了消除由选择标记基因引起的安全性隐患,人们发展了一些培育无选择标记转基因植物的策略。这些策略主要包括共转化、位点特异性重组和转座子转座等。去除选择标记基因将促进公众对转基因作物的接受。评述了无选择标记转基因植物的研究进展。     

转基因植物中标记基因的安全性新策略

转基因植物中的除草剂和抗生素抗性标记基因潜在的生态环境和食用安全性令人担忧。解决转基因植物中抗性标记基因安全性问题有两种途径：一是转化时仍使用抗性标记基因,转基因植物再生成功后,在释放大田前将标记基因剔除;二是发展安全性标记基因用于植物遗传转化。本文综述了三种标记基因剔除系统和几种安全性标记基因在植物转化中的应用进展。     

转基因植物中标记基因去除方法的研究进展

随着越来越多的转基因植物品种的出现,转基因植物的环境安全及食用安全越来越受到人们的广泛关注。目前存在的问题之一是转基因植物中抗生素或除草剂抗性的选择标记基因的存留。为了提高转基因植物的安全性,将转基因植物中选择性标记基因去除,不仅利于同一转基因植物的多次操作,而且更容易被人们所接受。就目前转基因植物中选择性标记基因去除的方法及其优缺点进行了横纵向的比较,希望对相关研究领域方法的选择方向具有指导意义。     

转基因植物中的卡那霉素抗性

在转基因植物中一个经常使用的标记基因就是卡那霉素抗性基因。对转基因植物的安全性估价将对于现代植物育种的普及和接受十分有利。对于这个标记基因的生物安全性进行估测认为,没有必要对含这个基因的转基因植物进行注册的限制。     

安全标记基因在转基因植物中的应用

转基因植物的抗性标记一直是转基因生物安全性争论的焦点,是限制转基因植物应用的瓶颈之一。筛选安全标记基因替代抗生素标记基因已成为解决转基因植物安全性和促进转基因植物应用的重要策略。综述了生物安全标记基因的产生背景、系统分类、筛选原理及不同起源的标记基因在植物基因工程中的应用和存在问题。选用植物内源标记基因已成为转基因植物安全标记基因研究的重要方向。     

无筛选标记转基因作物的研究进展

目前,大部分植物遗传转化都要使用选择标记基因,诸如抗生素、除草剂等来筛选转化子,由此而引起的转基因生物安全性问题受到了人们的关注。为了消除转基因作物中的筛选标记,一种全新的策略即无选择标记的转基因技术迅速发展起来。该技术既能够减轻大众对转基因植物中含有选择标记基因而引起的恐惧,又能将多个基因逐个整合到同一作物以实现基因聚合。因此这种新策略有着巨大的应用价值。对获得无筛选标记转基因作物的一些方法及其最新研究进展进行综述。     

无标记（Marker—Free）：转基因植物研究的新趋势

目前 ,几乎所有的植物遗传转化中都要使用选择性标记基因诸如抗生素或除草剂抗性基因等来筛选转化子。为了消除由此而引起的公众的安全性顾虑 ,一种全新的发展策略即获取无选择标记的转基因植物应运而生。无选择标记的转基因植物具有许多独特的优势 ,如消除大众对转基因植物中含有选择标记基因而引起的恐惧及可以反复地向已转化的植物中叠加外源基因等 ,因此 ,这种新策略 (无标记 )有着巨大的应用潜力。本文对获得无标记转基因植物的一些途径做一综述。     

培育无选择标记的转基因植物

目前,几乎所有的植物遗传转化都要通过使用选择标记基因,如抗生素或除草剂抗性基因等来筛选转化子,虽然没有研究结果表明选择标记基因影响人类健康或环境安全,但近年来也引发了人们对转基因产品安全性的担心。为了消除公众对转基因食品的安全性顾虑,无选择标记的转基因植物应运而生。本文综述了共转化系统、位点特异性重组系统(包括FLP/FRT、Cre/lox、R/RS及Gin/gix系统)和转座子系统(Ac/Ds转座子系统)在培育无选择标记转基因植物中的应用。     

可视安全标记花青素合成途径相关基因在植物转化研究中的应用进展

近年来有关转基因产品可能带来的环境和食品安全问题的争论多集中在作为标记基因的抗生素抗性基因以及抗除草剂基因的广泛使用。因此寻找安全、有效的标记基因替代上述有争议标记基因就显得十分必要和紧迫。花青素合成酶类及其合成调控因子可以控制植物体内的色素合成,一些转化研究已证明其转化体表型发生了颜色改变。加之花青素类物质是一些天然色素,对人类有利而无害,可以利用花青素的这些特点,将花青素合成酶类及其调控因子基因作为一类可视化、安全和有效的标记基因来转化植物。本文就可视安全标记花青素合成酶类基因及其调控基因在植物转化研究中的应用进展进行了简要介绍,以期为提高标记基因的安全性提供参考。     

Marker Gene Controversy in Transgenic Plants

Biosafety implications of selectable marker genes that are integrated into the transgenic plants are discussed. In the laboratory, selectable marker genes are used at two stages to distinguish transformed cells out of a large population of nontransformed cells: 1) initial assembly of gene cassettes is generally done in E. coli on easily manipulatable plasmid vectors that contain the selectable marker genes which often code for antibiotic inactivating enzymes, and 2) Then the gene cassettes are inserted into the plant genome by various transformation methods. For selection of transformed plant cells, antibiotic and herbicide resistance genes are widely used. Consequently, transgenic plants can end up with DNA sequences of selectable markers that are functional in E. coli and plants. The potential for horizontal gene transfer of selectable markers from transgenic plants to other organisms both in the environment and in the intestine of humans and animals is evaluated. Mechanisms and consequences of the transfer of marker genes from plants to other organisms is examined. Strategies to avoid marker genes in plants are discussed. It is possible to avoid the use of controversial selectable markers in the construction of transgenic plants.     

Selectable marker genes in transgenic plants: applications, alternatives and biosafety

Approximately fifty marker genes used for transgenic and transplastomic plant research or crop development have been assessed for efficiency, biosafety, scientific applications and commercialization. Selectable marker genes can be divided into several categories depending on whether they confer positive or negative selection and whether selection is conditional or non-conditional on the presence of external substrates. Positive selectable marker genes are defined as those that promote the growth of transformed tissue whereas negative selectable marker genes result in the death of the transformed tissue. The positive selectable marker genes that are conditional on the use of toxic agents, such as antibiotics, herbicides or drugs were the first to be developed and exploited. More recent developments include positive selectable marker genes that are conditional on non-toxic agents that may be substrates for growth or that induce growth and differentiation of the transformed tissues. Newer strategies include positive selectable marker genes which are not conditional on external substrates but which alter the physiological processes that govern plant development. A valuable companion to the selectable marker genes are the reporter genes, which do not provide a cell with a selective advantage, but which can be used to monitor transgenic events and manually separate transgenic material from non-transformed material. They fall into two categories depending on whether they are conditional or non-conditional on the presence of external substrates. Some reporter genes can be adapted to function as selectable marker genes through the development of novel substrates. Despite the large number of marker genes that exist for plants, only a few marker genes are used for most plant research and crop development. As the production of transgenic plants is labor intensive, expensive and difficult for most species, practical issues govern the choice of selectable marker genes that are used. Many of the genes have specific limitations or have not been sufficiently tested to merit their widespread use. For research, a variety of selection systems are essential as no single selectable marker gene was found to be sufficient for all circumstances. Although, no adverse biosafety effects have been reported for the marker genes that have been adopted for widespread use, biosafety concerns should help direct which markers will be chosen for future crop development. Common sense dictates that marker genes conferring resistance to significant therapeutic antibiotics should not be used. An area of research that is growing rapidly but is still in its infancy is the development of strategies for eliminating selectable marker genes to generate marker-free plants. Among the several technologies described, two have emerged with significant potential. The simplest is the co-transformation of genes of interest with selectable marker genes followed by the segregation of the separate genes through conventional genetics. The more complicated strategy is the use of site-specific recombinases, under the control of inducible promoters, to excise the marker genes and excision machinery from the transgenic plant after selection has been achieved. In this review each of the genes and processes will be examined to assess the alternatives that exist for producing transgenic plants.     

转基因植物中的标记基因研究新进展

文章综述了转基因植物中标记基因研究的新进展,主要包括以下3个方面：第一是采用共转化、位点特异性重组和转座子等技术对传统抗性标记基因进行消除,以利于对同一作物进行多次转基因操作;第二是完善各种已应用的以糖类代谢酶基因、耐胁迫酶类基因和绿色荧光蛋白基因等为安全标记基因的转化体系,并大力研究、开发潜在的汞离子还原酶基因、叶绿体合成关键酶基因等作为安全标记基因;第三是着力发展无标记基因、无载体骨架的简单高效转化体系。此外,还展望了安全标记的应用前景。     

无标记(Marker-Free):转基因植物研究的新趋势

目前,几乎所有的植物遗传转化中都要使用选择性标记基因诸如抗生素或除草剂抗性基因等来筛选转化子.为了消除由此而引起的公众的安全性顾虑,一种全新的发展策略即获取无选择标记的转基因植物应运而生.无选择标记的转基因植物具有许多独特的优势,如消除大众对转基因植物中含有选择标记基因而引起的恐惧及可以反复地向已转化的植物中叠加外源基因等,因此,这种新策略(无标记)有着巨大的应用潜力.本文对获得无标记转基因植物的一些途径做一综述.     

Biosafety and risk assessment framework for selectable marker genes in transgenic crop plants: a case of the science not supporting the politics

Selectable marker gene systems are vital for the development of transgenic crops. Since the creation of the first transgenic plants in the early 1980s and their subsequent commercialization worldwide over almost an entire decade, antibiotic and herbicide resistance selectable marker gene systems have been an integral feature of plant genetic modification. Without them, creating transgenic crops is not feasible on purely economic and practical terms. These systems allow the relatively straightforward identification and selection of plants that have stably incorporated not only the marker genes but also genes of interest, for example herbicide tolerance and pest resistance. Bacterial antibiotic resistance genes are also crucial in molecular biology manipulations in the laboratory. An unprecedented debate has accompanied the development and commercialization of transgenic crops. Divergent policies and their implementation in the European Union on one hand and the rest of the world on the other (industrialized and developing countries alike), have resulted in disputes with serious consequences on agricultural policy, world trade and food security. A lot of research effort has been directed towards the development of marker-free transformation or systems to remove selectable markers. Such research has been in a large part motivated by perceived problems with antibiotic resistance selectable markers; however, it is not justified from a safety point of view. The aim of this review is to discuss in some detail the currently available scientific evidence that overwhelmingly argues for the safety of these marker gene systems. Our conclusion, supported by numerous studies, most of which are commissioned by some of the very parties that have taken a position against the use of antibiotic selectable marker gene systems, is that there is no scientific basis to argue against the use and presence of selectable marker genes as a class in transgenic plants.     

转基因植物中标记基因的剔除

在目前的植物转化系统中,要求在关注基因或目的基因转入细胞时,同时有标记基因存在.标记基因主要是抗生素或除草剂的抗性基因.借标记基因的表达可以将转化细胞从大量的未转化细胞中筛选出来,但标记基因的继续存在,特别是在转基因食品中,是人们广泛关注的问题.培育无标记基因的转基因植株已成为植物生物工程研究中的新课题.该文介绍了剔除标记基因的两种方法:分离剔除和重组剔除,并对近年来这两种方法在培育无标记基因的转基因植物中的应用和进展作了介绍.