Improved Carica papaya Tolerance to Carmine Spider Mite by the Expression of Manduca sexta chitinase Transgene

Papaya plants producing the tobacco hornworm (Manduca sexta) chitinase protein were obtained following microprojectile bombardment of embryogenic calli derived from the hypocotyls of the cultivar Kapoho. Polymerase chain reaction (PCR) was carried out to confirm the presence of the transgene. RT-PCR and a quantitative chitinase assay showed increased levels of chitinase activity in every selected transgenic line. Insect bioassays in the laboratory showed that plants expressing the Manduca sexta chitinase gene significantly inhibited multiplication of carmine spider mites (Tetranychus cinnabarinus Boisd.). Experiments conducted to evaluate reaction of the transgenic plants to natural infection by carmine spider mites showed that the Manduca sexta chitinase gene provided increased tolerance under field conditions.     

基因枪法转化基因在小麦条锈菌中的瞬时表达

以小麦条锈菌(Puccinia striiformis f.sp.tritici)野生毒性菌株为转化受体,以含有gus报告基因的质粒(pGUS6L20)和潮霉素抗性基因的质粒(pKLHyg14)为载体,应用基因枪法研究了小麦条锈菌夏孢子遗传转化的瞬时表达特征。结果表明,在金粉直径为0.6μm、射程6cm、载体DNA5μL、可裂膜压力为900Psi或1100Psi时,gus基因和潮霉素抗性基因的瞬时表达率相对较高。     

百合组织培养和遗传转化的研究进展

百合是单子叶球茎类观赏花卉,具有食用和药用的作用。本文综述了国内外百合组织培养和遗传转化研究进展。包括营养器官、生殖器官和原生质体再生系统的建立。详细介绍了基因转化的方法,例如农杆菌、基因枪、电激穿孔等。浅析百合外源基因的表达,例如GUS、半夏凝集素基因、几丁质酶基因等。同时,讨论了百合遗传转化中存在的问题,以期为百合基因工程方面的研究提供依据。     

Structural insights into the mechanisms of drug resistance in HIV-1 protease NL4-3

The development of resistance to anti-retroviral drugs targeted against HIV is an increasing clinical problem in the treatment of HIV-1-infected individuals. Many patients develop drug-resistant strains of the virus after treatment with inhibitor cocktails (HAART therapy), which include multiple protease inhibitors. Therefore, it is imperative that we understand the mechanisms by which the viral proteins, in particular HIV-1 protease, develop resistance. We have determined the three-dimensional structure of HIV-1 protease NL4-3 in complex with the potent protease inhibitor TL-3 at 2.0 A resolution. We have also obtained the crystal structures of three mutant forms of NL4-3 protease containing one (V82A), three (V82A, M46I, F53L) and six (V82A, M46I, F53L, V77I, L24I, L63P) point mutations in complex with TL-3. The three protease mutants arose sequentially under ex vivo selective pressure in the presence of TL-3, and exhibit fourfold, 11-fold, and 30-fold resistance to TL-3, respectively. This series of protease crystal structures offers insights into the biochemical and structural mechanisms by which the enzyme can overcome inhibition by TL-3 while recovering some of its native catalytic activity.     

基因枪法介导GNA基因遗传转化甘蔗的研究

目的:将含有雪花莲外源凝集素(GNA)基因的植物表达载体用基因枪法分别导入一个果蔗和一个糖蔗品种中,以期获得转基因植株。方法:将GNA基因插入到植物表达载体上,构建出不同选择标记、不同启动子的表达载体,并用基因枪法将之导入甘蔗胚性愈伤组织,分别在G418、PPT和Hyg的选择压力下,筛选抗性植株,并进行分子杂交鉴定。结果:通过斑点杂交和PCR-Southern杂交证明GNA基因已整合到甘蔗基因组中。结论:用基因枪法成功获得了含有GNA基因的甘蔗转化株,为培育抗甘蔗绵蚜(Ceratovacuna lanigeraZehnther)的新品种提供了基础。     

RNAi Screening for Host Factors Involved in Vaccinia Virus Infection using Drosophila Cells

Viral pathogens represent a significant public health threat; not only can viruses cause natural epidemics of human disease, but their potential use in bioterrorism is also a concern. A better understanding of the cellular factors that impact infection would facilitate the development of much-needed therapeutics. Recent advances in RNA interference (RNAi) technology coupled with complete genome sequencing of several organisms has led to the optimization of genome-wide, cell-based loss-of-function screens. Drosophila cells are particularly amenable to genome-scale screens because of the ease and efficiency of RNAi in this system ¹. Importantly, a wide variety of viruses can infect Drosophila cells, including a number of mammalian viruses of medical and agricultural importance ^2,3,4. Previous RNAi screens in Drosophila have identified host factors that are required for various steps in virus infection including entry, translation and RNA replication ⁵. Moreover, many of the cellular factors required for viral replication in Drosophila cell culture are also limiting in human cells infected with these viruses ^{4,6,7,8, 9}. Therefore, the identification of host factors co-opted during viral infection presents novel targets for antiviral therapeutics. Here we present a generalized protocol for a high-throughput RNAi screen to identify cellular factors involved in viral infection, using vaccinia virus as an example.     

麦类作物小孢子培养与遗传转化研究进展

小孢子作为单个单倍体细胞,通过诱导培养能再生成纯合的二倍体植株,不仅为分子生物学、遗传学、形态发生学研究提供稳定的材料,也对小麦等作物的转基因研究提供新方向和新思路。本文主要介绍麦类作物小孢子培养的基本步骤及关键因素,简要介绍了以小孢子为材料进行小麦基因遗传转化的方法。     

玉米花粉体外萌发方法改进及其对花粉介导转基因的作用

超声波处理花粉介导植物基因转化方法由山西省农业科学院生物技术研究中心发明, 已被国家知识产权局授予发明专利(专利号ZL 99121152.9)。在该专利的基础上, 针对玉米(Zea mays)花粉取样、保存和处理条件等因素对其体外萌发的影响进行深入研究, 提出了改进玉米花粉体外萌发实验的方法。研究结果表明, 在不同时期对开花的玉米进行花粉培养时所需蔗糖溶液的浓度不同; 确定了玉米花粉的保存时间、条件及其对超声波处理后花粉萌发率的影响, 以提高该转化方法中花粉的活力, 并进一步验证了该转基因方法的可靠性; 讨论了玉米花粉体外萌发的操作技巧和各因子的参数, 对提高花粉介导植物基因转化效率有一定的参考价值。