植物安全转基因技术研究现状与展望

转基因技术是进行基因功能研究和作物遗传改良的重要工具。根癌农杆菌介导转化法和基因枪轰击法是两种主要的遗传转化方法。从1996年首例转基因作物商业化种植以来,2012年全球有28个国家和地区种植转基因作物,种植面积已达1.7亿公顷。随着全球转基因作物的大面积商业化种植,转基因植物的安全性越来越多地受到公众关注。安全转基因技术的研发对于转基因植物的商业化生产具有重要意义。文章详细介绍了目前已报道的安全转基因技术原理及其应用现状。按其作用原理和目的将其分为4类：安全选择标记、标记基因删除及基因叠加、基因漂移防控法和基因定点编辑整合技术。其中,安全选择标记按其筛选原理分为糖代谢相关标记、氨基酸代谢相关标记、激素相关标记和抗逆相关标记。与抗生素和除草剂抗性标记相比,这类标记基因及其表达产物对人和其他生物更安全。标记基因删除及基因叠加技术包括共转化法、位点特异性重组法、转座子法、同源重组法以及基于位点特异性重组的基因叠加技术,其中共转化法又包括农杆菌介导的共转化法和基因枪介导的表达盒共转化法。基因叠加技术为复合性状转基因植物研发奠定了基础,有望成为未来多性状转基因植物研发的重要技术之一。基因漂流防控法包括叶绿体转化法和基因拆分法。基因定点编辑和整合技术包括锌指核酸酶、TALEN和CRISPR/Cas9系统介导的基因定点编辑和整合技术。其中,TALEN和CRISPR/Cas9技术,设计简单,成本低,易操作,靶点分布广泛,有望成为安全转基因研发和应用的重要技术。文章对这些技术原理及其应用现状进行了综述,讨论了其优缺点,并展望了安全转基因技术研发重点和发展趋势。     

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。     

Effects of Active Modified Atmosphere Packaging on Postharvest Quality of Shiitake Mushrooms (Lentinula edodes) Stored at Cold Storage

The effects of active modified atmosphere packaging (MAP) on the postharvest quality of shiitake mushrooms stored at cold temperature (4°C) were investigated. The gas components were 2% O2+7% CO2 (MAP1), 2% O2+10% CO2 (MAP2) and 2% O2+13% CO2 (MAP3), respectively. The results showed that active MAP could extend the shelf-life of shiitake mushrooms to 17 d and the concentration of carbon dioxide could influence the postharvest quality of shiitake mushrooms. MAP2 treatment inhibited the increase in respiration rate and malondinaldehyde (MDA) contents, delayed the decrease in firmness, soluble sugar and vitamin C, and obviously reduced the activity of polyphenol oxidase (PPO) and the degree of browning, therefore maintaining better quality.     

氯化钠改性沸石对氨氮的吸附作用

采用30℃和90℃的NaCl溶液改性浙江缙云产天然沸石,通过静态吸附实验考察天然沸石及改性沸石对溶液中氨氮的吸附能力及机制,结果表明,NaCl改性可以提高沸石对氨氮的吸附能力。天然沸石及NaCl改性沸石对氨氮的吸附动力学过程符合“初期快速吸附,后期缓慢稳定”的特点。假二级动力学模型适合描述天然沸石及NaCl改性沸石对氨氮的吸附过程,颗粒内扩散模型仅适合于描述吸附反应初期天然沸石及NaCl改性沸石对氨氮的吸附过程。天然沸石和NaCl改性沸石对溶液中氨氮的吸附过程满足Langmuir和Freundlich等温吸附模型。90℃ NaCl改性沸石、30℃ NaCl改性沸石及天然沸石的氨氮饱和吸附量分别为19.5 mg/g、17.8 mg/g和17.2 mg/g。离子交换作用决定了溶液中氨氮向天然沸石及NaCl改性沸石的全部转移量。     

转基因大豆MON89788检测质粒标准分子构建与应用

[目的]构建适用于转基因大豆MON89788检测的质粒标准分子.[方法]利用定性PCR和连接、转化等分子克隆技术,将大豆内标准基因lectin、MON89788的3'端特异性序列和5'端特异性序列依次克隆到pMD18-T载体上,获得质粒标准分子pMD-LM3M5,并进行适用性验证.[结果]获得了3 700 bp的质粒标准分子,其中重组DNA片段1 029 bp.该质粒标准分子的定性PCR检测灵敏度达到10 copy.[结论]该研究构建的质粒标准分子pMD-LM3M5能替代NON89788基体标准品,用于MON89788大豆及其产品的定性PCR检测. Abstract： [Objective] The aim was to construct a plasmid reference molecule (PRM) for detection of transgenic soybean MON89788.[Method]the lectin gene sequence,3'-junction and 5'-junction sequence between host plant DNA integrated DNA of MON89788 soybean were amplified independently,and the three fragments were cloned into the cloning vector pMD18-T in order through molecular manipulation method to construct pMD-LM3M5,the applicability of the constructed novel PRM was tested. [Result] Sequencing confirmation result showed that the PRM was 3 700 bp in length,containing 1 029 bp of recombined DNA fragment.The limits of qualitative detection of the PRM were 10 copies,[Conclusion]The PRM constructed in this study was suitable for the identification of MON89788 event.     

Risk Assessment and Biosafety Studies of Transgenic Bt Rice （Oryza sativa L.）

Transgenic crops having alien genes from different sources are getting popularity. A Rice （Oryza sativa L.） containing gene from Bacillus thuringiensis, a soil bacterium, was assessed to study the potential risks of transgenic plants on environment. The crop was found resistant to target insect pests. Rice variety Basmati-370 transformed with two insecticidal genes, crylAc and cry2A, was grown under field conditions for several years. Data were collected at different stages of the plant growth. Fate of Cry protein in soil, effect of Bt protein on non-target insects, risks of vertical and horizontal gene flow were evaluated. No potential hazard was found at all levels. Bt protein was unstable and degraded significantly in soil within 30 days after harvesting the crop. No harmful effects were found on non-target insects （insects other than order lepidoptera）. Maximum gene flow of 0.02% was observed at close spacing and no evidence of horizontal gene transfer to Rhizobium spp. was found. In conclusion, the transgenic rice plants transformed with Bt genes have no harmful effects on the environment.     

Persistence of an oilseed rape transgene in the environment

Transgenic bromoxynil (BX)-resistant spring oilseed rape (Brassica napus L.) was grown commercially in Canada only for two crop seasons – 2000 and 2001. We investigated the persistence of suspected BX-resistant oilseed rape volunteers in a 64-ha wheat field in Saskatchewan, Canada in 2007, 7 years after the BX-resistant cultivar BX Armor was grown. A small oilseed rape volunteer population, estimated at less than 100 plants, was observed in three main areas or patches in the field. These patches were located in low-lying areas that were too wet to plant or spray with herbicides in 2007. Viable seed was collected from 35 mature volunteers and F₁ progeny screened with BX at 280 g ai/ha in the greenhouse. Progeny of all of the volunteers were visually rated as BX-resistant; the presence of the Oxy 235 transgene in leaf tissue of progeny of all volunteers was confirmed by PCR analysis. This study has documented the longest persistence of oilseed rape volunteers in Canada. Volunteers were not observed in 2008 or 2009, because of drought conditions in spring of both years. Results support the findings from previous studies that persistence of volunteer oilseed rape populations in western Canada is generally ephemeral or transitory in the absence of seed bank immigration. However, this study shows that oilseed rape transgenes can persist in the environment for a number of years even after all cultivars with the conferred trait are removed from the market.     

感官剖面分析法在调制乳感官分析技术中的应用研究

以麦香调制乳的感官剖面分析为例,通过描述词的收集和删减、使用描述词培训感官评价小组、建立感官剖面图来分析说明感官剖面分析法在调制乳的感官分析技术中的应用。通过分析比较说明,感官剖面分析法可以量化感官特征,建立行之有效的感官分析控制体系,为调制乳产品的研发和质量控制提供感官分析技术。     

冷却猪肉中假单胞菌生长预测模型的建立与验证

以从腐败猪肉中分离得到的假单胞菌P1,P2,P3,P4,P5,P6的混合培养物为受试菌液,应用Origin7．5软件拟合假单胞菌0-10℃下的生长情况。经比较发现,修正的Gompertz模型对假单胞菌生长曲线的拟合效果优于线性模型,R2均在0．96以上。应用平方根模型和Arrhenius模型对由修正的Gompertz模型得出的最大比生长速率进行拟合,所得Arrhenius模型的R2为0．9716,拟合效果优于平方根模型（R2为0．9326）。结果表明,所建二级模型,能真实快速有效地预测0-10℃冷藏条件下假单胞菌的生长。     

变温压差膨化干燥对冬枣总黄酮和水溶性糖含量的影响

研究了变温压差膨化干燥对冬枣中总黄酮和水溶性糖含量的影响。比较了超声波-微波辅助乙醇法和乙醇法提取冬枣总黄酮的效果,采用水提法提取冬枣中水溶性糖,参照Fenton法比较了膨化前后冬枣黄酮的清除羟基自由基能力,采用Sehpadex G-150凝胶层析法测定冬枣水溶性糖分子量分布。结果表明:微波-超声波辅助乙醇法提取总黄酮优于乙醇法。冬枣在膨化前后总黄酮和水溶性糖含量发生了显著的变化。冬枣鲜果、预干燥冬枣和膨化冬枣中黄酮含量分别为1.59%、0.71%、0.38%,膨化冬枣清除羟基自由基能力较冬枣鲜果下降。冬枣鲜果、预干燥冬枣和膨化冬枣中水溶性总糖分别为45.23%、55.84%、44.34%,还原糖分别为38.91%、47.29%、38.56%。Sehpadex G-150凝胶层析结果表明,冬枣鲜果中含有较多的大分子多糖,而膨化枣中含有较多的小分子糖。