A Simple and Reliable Assay for Detecting Specific Nucleotide Sequences in Plants Using Optical Thin-film Biosensor Chips

Here we report the adaptation and optimization of an efficient, accurate and inexpensive assay that employs custom-designed silicon-based optical thin-film biosensor chips to detect unique transgenes in genetically modified （GM） crops and SN-P markers in model plant genomes. Briefly, aldehyde-attached sequence-specific singlestranded oligonucleotide probes are arrayed and covalently attached to a hydrazine-derivatized biosensor chip surface. Unique DNA sequences （or genes） are detected by hybridizing biotinylated PCR amplicons of the DNA sequences to probes on the chip surface. In the SN-P assay, target sequences （PCR amplicons） are hybridized in the presence of a mixture of biotinylated detector probes and a thermostable DNA ligase. Only perfect matches between the probe and target sequences, but not those with even a single nucleotide mismatch, can be covalently fixed on the chip surface. In both cases, the presence of specific target sequences is siL, nified by a color change on the chip surface （gold to blue/purple） after brief incubation with an anti-biotin IgG horseradish peroxidase （HRP） to generate a precipitable product from an HRP substrate.     

植酸酶基因定性PCR检测方法及阳性质粒分子的构建

植酸酶基因在农业生产中具有很高的应用价值,正被广泛用于农作物的基因工程研究。为了满足转植酸酶基因作物安全监管的需要,通过优化PCR检测条件,建立了植酸酶基因特异性定性PCR检测方法。该方法具有良好的扩增特异性和检测灵敏度,可以满足我国植酸酶转基因作物监管需要。此外,我们将6大作物(小麦、水稻、棉花、大豆、玉米和油菜)的内标准基因和植酸酶基因克隆到同一载体上,构建成了阳性质粒分子pBS Endogenous-phytase。该质粒分子适用于这6大作物中植酸酶基因的筛查检测。本研究为转植酸酶基因作物的安全监管提供了阳性材料和检测方法。     

加快发展现代农业|大力推进转基因生物育种产业化

生物技术同常规农业技术结合，推进转基因生物育种研究开发和产业化，将对我国现代农业的发展发挥巨大作用。综述了国内外转基因农作物研究和产业化的最新进展，并建议：①实行积极的产业化政策；②加强科普宣传，理性认识转基因安全；③加强基础研究，提高自主创新能力。     

转基因生物技术的生态风险及对策研究

转基因生物技术是现代分子生物技术的一个分支学科,它的开发和应用是人类智慧的产物,人们在利用生物技术造福的同时,也可能带来安全问题,特别是转基因生物技术的滥用对生物多样性、生态环境及人体健康都可能构成危险或潜在的风险,即生态风险问题。各个国家和地区通过制定一系列的法律、法规对转基因生物技术及应用进行监管。我国当前没有关于转基因生物技术生态风险防范的全面而系统的法规体系,在分析和比较国内外转基因生物技术相关立法的基础上,提出相应的完善对策。     

基于多重串联式PCR的基因碟片技术检测转基因作物

【目的】建立基于多重串联式PCR（multiplexed tandem PCR,MT-PCR）的基因碟片技术,并使之应用于转基因作物的大量、快速和稳定地检测。【方法】以转基因作物研究中常用的调控序列NOS终止子、FMV35S启动子及外源基因NPTⅡ、Cry1Ab、Cry1Ab/Ac、CP4-EPSPS、PAT和玉米内源基因IVR、棉花内源基因sad1、菜籽粕内源基因PEP为检测对象,针对每个基因设计内外2对引物,先进行一次循环数较少（10-20 cycles）的高通量多重PCR,以便在均匀地扩增各基因和调控序列的同时避免引物之间的竞争,然后利用巢式荧光定量PCR检测各个基因和调控序列,最后根据扩增曲线和熔融曲线分析结果。【结果】该方法能够快速（＜2 h）、高通量、准确地（＞0.000292 ng）检测出棉花、玉米、大米、菜籽粕中的多种转基因成分,可以分辨出3种转基因物种,适合大批量检测。【结论】该方法适合转基因作物的高通量、定量检测,具有较好的应用前景。     

甘蔗转基因改良及转基因检测机构的角色

我国甘蔗转基因研究取得了长足进展，现对我国甘蔗转基因研究实践中使用的改良方法进行回顾总结，并结合国际甘蔗转基因研究的现状指出未来甘蔗转基因研究的发展方向。针对转基因研究的高度社会敏感性，阐述法定转基因检测机构对保护和促进我国甘蔗转基因研究健康发展将起到的重要作用。     

Tipping Points in Seaweed Genetic Engineering: Scaling Up Opportunities in the Next Decade

Seaweed genetic engineering is a transgenic expression system with unique features compared with those of heterotrophic prokaryotes and higher plants. This study discusses several newly sequenced seaweed nuclear genomes and the necessity that research on vector design should consider endogenous promoters, codon optimization, and gene copy number. Seaweed viruses and artificial transposons can be applied as transformation methods after acquiring a comprehensive understanding of the mechanism of viral infections in seaweeds and transposon patterns in seaweed genomes. After cultivating transgenic algal cells and tissues in a photobioreactor, a biosafety assessment of genetically modified (GM) seaweeds must be conducted before open-sea application. We propose a set of programs for the evaluation of gene flow from GM seaweeds to local/geographical environments. The effective implementation of such programs requires fundamentally systematic and interdisciplinary studies on algal physiology and genetics, marine hydrology, reproductive biology, and ecology.     

Evaluation of the feed value of a transgenic strain of the narrow‐leaf lupin (Lupinus angustifolius) in the diet of the marine fish,Pagrus auratus

This study assessed the nutritional and biological value of a noncommercial, transgenic line of the Australian sweet lupin, Lupinus angustifolius, (cv. Warrah), produced to increase the methionine content of the seed. An initial experiment demonstrated that differences in the methionine content of the transgenic and nontransgenic control lupins had no apparent influence on the growth of juvenile red seabream fed practical diets. Re‐evaluation of the nutritional characteristics of the lupin meals with subsequent digestibility studies allowed the determination of the digestible value of the protein and energy content of each of the varieties. The digestible protein content of either variety was similar, however significant differences in the digestible energy value of each variety existed (56.3% cf. 64.0%). This re‐evaluation of the nutritional value of the genetically manipulated (GM) and non‐GM lupin varieties enabled the reformulation of diets on a digestible protein and energy basis. A second growth trial was undertaken using sub‐satietal pair‐feeding regimes, with the experiment also involving protein‐restrictive diets to allow expression of the differences in the methionine content of the transgenic and nontransgenic lupin meals. A significant benefit of the enhanced methionine level in the transgenic lupin was observed. It is argued that in the high‐protein fish diets used, the importance of amino acid composition is relatively limited. Economic modelling of the potential value of the increased methionine in the transgenic lupin suggests that this will have limited benefit for aquaculture industries, and that greater value would be attributable to higher protein and energy levels in ingredients.     

草胺膦抗性基因bar的原核表达与蛋白质纯化

通过PCR扩增,从pCAMBIA1300-bar质粒中获得bar基因编码区全长,经酶切鉴定和测序后证实,bar基因分离成功。用BamH和Hind酶切,将bar基因与原核表达载体pET28a(+)连接,构建成重组质粒pET28bar。将重组质粒pET28 bar转化大肠杆菌菌株BL21(DE3)后,获得高效表达。SDS-PAGE电泳分析显示,目的蛋白主要以包涵体形式存在,蛋白质分子量约为27 ku。将包涵体离心、洗涤和纯化后,得到高纯度的目的蛋白。该蛋白可以替代植物外源蛋白进行转bar基因产品的食用安全性检测,也可用于转bar基因产品ELISA检测的抗体制备。     

转基因农产品标识管理和检测技术研究进展

对国内外转基因植物和转基因农产品及其加工食品的释放情况进行综述,并介绍了世界主要国家和地区对转基因食品的安全管理法规,重点概述了转基因农产品及其加工食品检测技术的进展情况,对各种检测方法及其应用范围做了比较,提出了今后我国在转基因检测技术方面的研究领域和发展趋势。