Effects of codon modification on human BMP2 gene expression in tobacco plants

Bone morphogenetic protein 2 (BMP2) has great potential in therapeutic applications. We are working on generating transgenic plants as a bioreactor to produce BMP2. We have studied the effects of codon optimization on the expression of human BMP2 (hBMP2) in tobacco plants. Three modified hBMP2 genes were transformed into tobacco under the control of either cauliflower mosaic virus 35S (CaMV35S) promoter or double-CaMV35S promoter plus alfalfa mosaic virus (AMV) enhancer. The fused β-glucuronidase (GUS) reporter gene was used to facilitate the assay of protein expression. The results indicated that codon optimization could increase the protein expression level obviously under CaMV35S promoter. However, under relatively stronger initiation condition (double-CaMV35S promoter plus AMV enhancer), only the gene with the lowest degree of codon optimization could increase the protein expression level. Our findings suggest that the action of codon optimization may be influenced by the factors of promoter strength and A+T content in tobacco plants.     

Antibody-based surface plasmon resonance detection of intact viral pathogen

Surface plasmon resonance (SPR) technique was used to directly detect an intact form of insect pathogen: the baculovirus, Autographa californica multiple nuclear polyhedrosis virus (AcMNPV). An SPR sensor chip with three bio-functional layers was used to detect the intact AcMNPV: amine-reactive crosslinker with a disulfide bond that chemisorbs to gold film, Protein A, and a mouse IgG monoclonal antibody raised against a surface protein of the target viral pathogen. A two-channel (reference & test) micro-fluidic SPR system is used for reliable measurement. Bio-specific response to the AcMNPV is compared with the response for tobacco mosaic virus (TMV) as control. Successive exposure of the sensor chip to both viruses verifies a specific response to AcMNPV. This serves as a prerequisite to the development of a new type of viral pathogen detection sensors.     

Expression of viral capsid protein antigen against Epstein-Barr virus in plastids of Nicotiana tabacum cv. SR1

Epstein-Barr virus (EBV) infects nearly 90% of adults worldwide and is the pathogenic source of a broad spectrum of malignancies originating from lymphoid and epithelial cells. Currently, no vaccine has been developed to immunologically inactivate this virus. In infected patients, anti-EBV viral capsid antigen (VCA) immunoglobins represent some of the useful diagnostic markers for carcinoma development. To demonstrate that the EBV VCA antigen can be produced in plants, the plastid genome of tobacco (Nicotiana tabacum cv. SR1) was transformed with a VCA-expressing cassette. The EBV VCA mRNA was actively transcribed in the transplastomic plants and antigen production was detected. This study indicates that plastid transformation could be a promising strategy in EBV VCA antigen production.     

Comparison of performance on different host plants between the B biotype and a non-B biotype of Bemisia tabaci from Zhejiang, China

The capacity of the B biotype of the whitefly, Bemisia tabaci (Gennadius) (Homoptera: Aleyrodidae), to invade has often been linked to its presumably wider host range than the non‐B indigenous biotypes. However, there are few experimental studies of the relative performance of the B biotype and non‐B biotypes on different host‐plant species. Here, we compared the performance of the B biotype and an indigenous non‐B biotype (China‐ZHJ‐1) of B. tabaci from Zhejiang, China on five commonly cultivated host plants, each from a different family: cotton, tobacco, cabbage, squash, and kidney bean. We also examined the effect of rearing host plants on the performance of the B biotype. Overall, the performance of the B biotype on the five species of plants was much better than that of the indigenous non‐B population. On tobacco, cabbage, and kidney bean, no individuals of ZHJ‐1 completed development to adulthood, whereas the B biotype developed successfully from egg to adult on all three plants. On squash, the B biotype survived better, developed to adulthood earlier and had a higher fecundity than ZHJ‐1. The two biotypes performed more equally on cotton, but even on this plant the B biotype female adults lived nearly twice as long as that of ZHJ‐1 and may have realized a higher life‐time fecundity. The B biotype also showed a substantial capacity to acclimatize to alternative host plants for improved survival and reproduction, on both highly suitable and marginally suitable host plants. We conclude that the host range of the B biotype of B. tabaci may be much wider than those of some indigenous biotypes, and this advantage of the B biotype over the non‐B biotypes may assist in its invasion and displacement of some indigenous biotypes in the field.     

Visualization of actin cytoskeletal dynamics during the cell cycle in tobacco (Nicotiana tabacum L. cv Bright Yellow) cells

BACKGROUND INFORMATION: The actin cytoskeleton forms distinct actin arrays which fulfil their functions during cell cycle progression. Reorganization of the actin cytoskeleton occurs during transition from one actin array to another. Although actin arrays have been well described during cell cycle progression, the dynamic organization of the actin cytoskeleton during actin array transition remains to be dissected. RESULTS: In the present study, a GFP (green fluorescent protein)-mTalin (mouse talin) fusion gene was introduced into suspension-cultured tobacco BY-2 (Nicotiana tabacum L. cv Bright Yellow) cells by a calli-cocultivation transformation method to visualize the reorganization of the actin cytoskeleton in vivo during the progression of the cell cycle. Typical actin structures were indicated by GFP-mTalin, such as the pre-prophase actin band, mitotic spindle actin filament cage and phragmoplast actin arrays. In addition, dynamic organization of actin filaments was observed during the progression of the cell from metaphase to anaphase. In late metaphase, spindle actin filaments gradually shrank to the equatorial plane along both the long and short axes. Soon after the separation of sister chromosomes, actin filaments aligned in parallel at the cell division plane, forming a cylinder-like structure. During the formation of the cell plate, one cylinder-like structure changed into two cylinder-like structures: the typical actin arrays of the phragmoplast. However, the two actin arrays remained overlapping at the margin of the centrally growing cell plate, forming an actin wreath. When the cell plate matured further, an actin filament network attached to the cell plate was formed. CONCLUSIONS: Our results clearly describe the dynamic organization of the actin cytoskeleton during mitosis and cytokinesis of a plant cell. This demonstrates that GFP-mTalin-transformed tobacco BY-2 cells are a valuable tool to study actin cytoskeleton functions in the plant cell cycle.     

烟草根际微生物研究

利用选择性培养基,对土壤肥力肥沃、中等、贫瘠的烟区根际细菌、真菌和放线菌进行了分离和测数。根据菌体形态及培养特征、生理生化指标及16S rDNA部分序列分析等,对根际自生固氮菌、磷细菌、钾细菌进行了鉴定和分类。主要结果为：土壤肥沃的烟区根际细菌的数量最多,土壤贫瘠的烟区数量最少;土壤贫瘠烟区根际真菌的数量较多,中等和肥沃烟区的较少;根际故线菌的数量随土壤肥力的降低而依次减小。从不同肥力烟区分离的根际自生固氮菌、磷细菌、钾细菌以革兰氏阴性杆菌为主,分别属于10个属。土壤的肥沃程度对根际3类细菌的种类和数量都有影响,总体上看,土壤肥沃和中等的条件下,细菌类群的多样性和丰富度较大,而贫瘠土壤细菌类群的优势度较大。     

葡萄蔗糖转运蛋白在烟草中的反义表达及其对转基因烟草的影响

将葡萄蔗糖转运蛋白VvSUC27cDNA以反义方向插入到含有CaMV35s启动子的真核表达载体pBI121载体中,然后转化到烟草(Nicotianatobacumcv.Samsun)植株中.转反义VvSUC27cDNA的烟草植株在含有20g/L蔗糖的培养基上能够正常生长发育,但是通过切片观察,发现其根部发育较弱,且叶片叶绿体含量增加.可溶性糖测定发现,转基因烟草根部蔗糖含量只有野生型烟草的51%.14C蔗糖吸收实验发现转基因烟草转运外界蔗糖的能力大大降低.     

烟草抗TMV基因工程的研究进展

烟草花叶病毒病是一种危害严重的世界性病害,可大幅度降低烟草的产量和品质.研究和探索防治烟草花叶病毒病的新技术、新途径已成为众多研究者普遍关注的焦点.分子生物学的发展,特别是基因工程的发展为防治该病毒病带来了希望.本文综述了目前普遍采用的几种烟草抗TM V基因工程策略的研究现状及评价.     

转LEA基因烟草的耐盐性分析

目的:验证柽柳LEA基因的功能,为通过基因工程手段培育耐盐植物提供基础资料。方法:对转LEA基因烟草当代(T0)和子一代(T1)分别进行不同浓度的NaCl胁迫处理,研究转基因烟草的耐盐性。结果:转基因烟草T0代组培苗耐受NaCl的临界浓度为230mmol/L,而对照耐受NaCl的临界浓度为130mmol/L以下;T1代幼苗耐受NaCl的临界浓度为150mmol/L,对照耐受NaCl的临界浓度为100mmol/L以下;在临界浓度转基因烟草的T0代、T1代根系发育良好,生长量明显高于非转基因对照烟草。结论:柽柳LEA基因的转化提高了烟草的耐盐性。     

烟碱的微生物降解研究进展

烤烟叶面微生物类群有细菌、放线菌、霉菌和酵母菌,它们在烟叶生长、调制、陈化、加工和贮存过程中对烟叶质量产生很大的影响。就烟草生长至加工过程中微生物菌群动态变化及微生物在降解烟叶烟碱上的应用作了概述,分析了微生物降烟碱的应用前景。