基因流——转基因生物安全性的关键问题

基因流是基因从一个生物体转移到另一个生物体的基因流动过程.本文探讨了转基因作物基因流研究的现状,综述了转基因对环境和生物体的潜在风险,并讨论了监测和管理基因流的方法.     

Bt/CpTI抗虫转基因水稻及其非转基因亲本与普通野生稻杂种F2群体的等位基因偏态分离

了解水稻转基因是否影响野生稻基因传递的频率,对于评价转基因逃逸及其生态影响有重要意义.本研究构建了不含转基因和含转基因的栽培稻与普通野生稻杂种F2群体,利用分子标记检测了F2群体各位点的基因型和基因频率.以连续卡方分析了F2群体相关位点的基因型和基因频率观察值是否符合理论分离比,同时对各位点进行了连锁不平衡分析,并对实验群体所需的最小样本量进行理论探索.结果表明两个F2群体分别有25.93%与33.33%的位点出现了显著的非随机分离,非转基因与转基因F2群体分别在偏态分离位点数与偏离亲本方向上出现一定差异,并观察到连锁不平衡位点.实验个体数应不少于280个才能保证实验结果准确.外源转基因在杂种后代群体中会因为选择而影响基因分离,进而影响杂种群体的进化潜力.     

3个茶树品种WOX基因家族的进化及密码子偏好性比较

【目的】WOX基因家族在模式植物组织培养中扮演着重要的分子调控角色。非模式植物WOX基因家族的系统发育和密码子使用偏好性研究有助于探索遗传进化规律和基因表达特征,进而为其转基因体系构建提供指导。【方法】利用生物信息学方法在3个茶树品种全基因组数据中鉴定WOX基因;通过ClustalX 2.1和MEGA X软件探索它们的进化规律。运用Perl语言、Codon W 1.4.2程序和SPSS 23.0等软件分析3个茶树品种WOX基因编码序列,探究它们的密码子使用偏好模式;基于ENc-GC_3s、PR2分析和中性分析结果解析影响密码子偏好的主要因素。【结果】共鉴定出42个WOX成员,其中‘云抗10号’(CSA)11个,‘舒茶早’(CSS)18个,‘云南野生古茶DASZ’(DASZ)13个,系统发育揭示3个茶树品种WOX基因在进化上存在较强的保守性。密码子偏好性分析揭示,3个茶树品种WOX基因密码子都偏好使用A/T和以A/T结尾;CSA和DASZ的WOX基因密码子使用模式相似,表明两者亲缘关系较近;以CSA和CSS作为转基因受体时,3个茶树品种WOX基因中个别密码子需优化;烟草是3个茶树品种WOX基因的最佳异源表达受体;ENc-GC_3s绘图、PR2分析和中性分析表明自然选择是影响3个茶树品种WOX基因密码子使用偏倚的主要因素。【结论】3个茶树品种WOX基因进化上较保守,密码子偏好使用A/T和以A/T结尾,且密码子偏好原因主要是自然选择;揭示出转茶树时密码子的优化信息和烟草为最佳异源表达受体的结果。     

Application of a nuclear localization signal gene in transgene mice

Efficient gene transfer by cytoplasm co-injection will offer a powerful means for transgenic animals. Using co-injection in cytoplasm, two independent gene constructs, including bovine α-s1-casein-hG-CSF and a mammal expression vector expressing a nuclear localization signal (mNLS), were introduced into fertilized mouse eggs. The target gene construct was docked into host nucleus probably by the nuclear localization signal. Transgene mice have been obtained at 58% (29/50) of integration ratio. Expression level of the positive transgene mice was detected by Western blotting. Maximal expression of human G-CSF was estimated about 540 mg/L of milk. The expression ratio was up to 75% (9/12). The results here have important practical implications for the generation of mammary gland bioreactors and other transgene studies. Co-injection of a target gene with an expression vector of a mammal nuclear localization signal by cytoplasm appears to be a useful, efficient and easy strategy for generating transgenic animals, which may be able to substitute the routine method of pronucleus-injection of fertilized eggs.     

Can transgenic rice cause ecological risks through transgene escape?

Alien transgene escape from genetically engineered rice to non-transgenic varieties or close wild relatives (including weedy rice) may lead to unpredictable ecological risks. However, for transgene escape to occur three conditions need to be met: (i) spatially, transgenic rice and its non-transgenic counterparts or wild relatives should have sympatric distributions; (ii) temporally, the flowering time of transgenic rice and the non-transgenic varieties or wild relatives should overlap; and (iii) biologically, transgenic rice and its wild relative species should have such a sufficiently close relationship that their interspecific hybrids can have normal generative reproduction. This paper presents research data on the geographic distribution, flowering habits, interspecific hybridization, and gene flow of cultivated rice (Oryza sativa) and its closely related wild relatives containing the AA genome. The objective is to estimate the possibility of transgene escape to non-transgenic rice varieties and wild relatives of rice, which may result in unpredictable ecological risks.     

转基因植物及其安全性问题

介绍了转基因植物在全球和我国发展概况,介绍了转基因植物的几种目的基因:抗除草剂基因、抗病虫基因、改良作物品质基因,及其在农业上的应用,从转基因植物食品的安全性和转基因生态环境的安全性讨论了正确对待转基因技术的安全性问题。     

Robust and regulatory expression of defensin A gene driven by vitellogenin promoter in transgenic Anopheles stephensi

The use of genetically modified mosquitoes to reduce or replace field populations is a new strategy to control mosquito-borne diseases. The precondition of the implementation of this strategy is the ability to manipulate the genome of mosquitoes and to induce specific expression of the effector molecules driven by a suitable promoter. The objective of this study is to evaluate the expression of defensin A gene of Anopheles sinensis under the control of a vitellogenin promoter in transgenic Anopheles ste- phensi. The regulatory region of Anopheles gambiae vitellogenin was cloned and subcloned into transfer vector pSLFa consisting of an expression cassette with defensin A coding sequence. Then, the expression cassette was transferred into transformation vector pBac[3xP3-DsRedafm] using Asc I di- gestion. The recombinant plasmid DNA of pBac[3xP3DsRed-AgVgT2-DefA] and helper plasmid DNA of phsp-pBac were micro-injected into embryos of An. stephensi. The positive transgenic mosquitoes were screened by observing specific red fluorescence in the eyes of G1 larvae. Southern blot analysis showed that a single-copy transgene integrated into the genome of An. stephensi. RT-PCR analysis showed that the defensin A gene expressed specifically in fat bodies of female mosquitoes after a blood meal. Interestingly, the mRNA of defensin A is more stable compared with that of the endogenous vitellogenin gene. After multiple blood meals, the expression of defensin A appeared as a reducible and non-cycling type, a crucial feature for its anti-pathogen effect. From data above, we concluded that the regulatory function of the Vg promoter and the expression of defensin A gene were relatively con- served in different species of anopheles mosquitoes. These molecules could be used as candidates in the development of genetically modified mosquitoes.     

神经系统细胞谱系和神经束路示踪技术

细胞谱系和神经束路示踪是指利用各种方式标记细胞，并对其及后代细胞的增殖、分化以及迁移、投射等活动进行追踪观察。自20世纪以来，示踪技术为研究神经系统发育提供了重要的手段。随着分子克隆技术的发展，荧光蛋白的发现，诱导重组酶系统Cre ／loxp的应用，极大地拓宽了示踪技术的应用范围。文章将结合该技术在神经系统发育研究中的应用，对其种类、原理、特点进行概述。     

Isolation of a pollen specific promoter in tritordeum

The promoter is a cis-acting element in regulating gene expression. A promoterless plasmid containing UidA gene was transformed into tritordeum by barmbadment. Histochemical analysis of various tissues in transgenic tritordeum was carried to examine tissue-specific expression of GUS(beta-glucuronidase) activity. The pollen-specific promoter was trapped and identified successfully in a transformant line. PCR(polymerase chain reaction) method was used to isolate this pollen-specific promoter. By sequencing and analyzing the amplified fragment from PCR, a part of UidA gene and a flanking sequence were obtained. Some essential elements of plant promoters were found in the sequence. To determine the function of it, the cloned fragment was fused with UidA gene, then cloned and transformed into Triticum durum. The transgenic plant transformed by this vector showed GUS expression only in pollen. Therefore a pollen-specific promoter was isolated successfully.