Chloroplasts are a vital group of organelles of plants, yet the molecular mechanisms associated with their division remain poorly understood. Recent studies have revealed that the FtsZ protein, known as a key component in prokaryotic cell division, is involved in chloroplast division process. The NtFtsZ2-1 gene was isolated from Nicotiana tabacum by RT-PCR, and the sense and antisense expression plasmids were used to examine the function of NtFtsZ2-1 gene in transgenic tobacco. Light and confocal observations revealed that the normal chloroplast division process was severely disrupted in transgenic plants with enhanced or reduced expression of NtFtsZ2-1 gene. These chloroplasts were abnormally larger in size and fewer in number compared with that of the wild-type tobacco. But the total chloroplast plan area per mesophyll cell was conserved in sense, antisense and wild type tobaccos. Analyses of electron micrographs and chlorophyll content of different transgenic plants showed that constitutively enhancing or inhibiting the expression of NtFtsZ2-1 gene had no direct influence on the ultrastructure and photosynthetic ability of chloroplasts. Basing on these results, we suggest that NtFtsZ2-1 gene is involved in chloroplast division and expansion; the fluctuation of NtFtsZ2-1 expression level would alter normal chloroplast number and size in plant cells. In addition, the similarities of ultrastructure and photosynthetic ability of chloroplasts among sense, antisense and wild type tobaccos implies that a special mechanism regulate the relationship between chloroplast number and size to maximize photosynthetic rate.

SummaryThis study reports an Agrobacterium-mediated transformation of green-colored cotton (Gossypium hirsutum L.). A tissue culture procedure was optimized to induce callus formation from hypocotyl explants and subsequent differentiation into the embryogenic type. Callus formation could be induced by growing explants on Murashige and Skoog medium containing 2,4-dichlorophenoxyacetic acid and kinetin. Among the four genotypes studied, embryogenic calli and plant regeneration were observed only in var. G9803. Agrobacterium-mediated transformation of G9803 with the fiber-specific expansin gene GhExpl was achieved based on the establishment of these tissue culture methods. A total of 32 individual regenerants resistant to kanamycin were generated within 7 mo., with a transformation frequency of 17.8%. Transformation was confirmed by Southern blot analysis and RT-PCR. These results represent the first step towards genetic manipulation of the colors and fiber quality of green-colored cottons by biotechnology.

农杆菌介导法、花粉管通道法和基因枪法是植物遗传转化的主要方法,这些转化方法在植物基因工程育种和基因改良中起到了重要作用.这些遗传转化方法目前已经广泛应用于棉花抗虫、抗病和品质改良育种实践,并获得了抗虫、抗病等性状良好的转基因植株,有些已经进入商品化生产,取得了显著的经济效益和社会效益.本文重点综述了这3种应用较广的遗传转化方法及其在棉花抗虫、抗病和品质改良育种中的应用,并讨论了我国转基因棉花研究的进展.

Pure lines from anther culture of intergeneric hybrids between wheat and Leymus multicaulis were used to develop pure translocation lines with salt-tolerance and ideal agronomic characteristics through cytological detection, field selection,resistance assessment,comprehensive observation of agricultural characteristics,yield trials,and molecular markers (GISH).Results showed that anther culture can obtain large number of breed-true variants in a short time by avoiding excessive segregation of posterity and promoting structural variation of chromosomes in offspring from wheat distant hybridization.This greatly shortened the time in introduction of alien genes to wheat through distant hybridization.

We have determined the Rubisco components and activity, whole leaf protein and amino acid components of thermo-sensitive mutant line 1103s of rice (Oryza sativa ssp. indica) leaves during induces green and yellow banding in this study. The results are as follows: The structure and components of Rubisco in the mutant are the same as in the wild form and relatively stable, but the activity of the mutant Rubisco greatly changes as a special protein of molecular weight 56.2 kD (PI=4.5) appears and disappears. When the green-yellow bands appear, the special protein disappears, and the activity of the mutant-rice Rubisco decreases, whereas when the green-yellow bands in the same part of the leaves disappear, the special protein appears and the activity of Rubisco is increase. The above shows the changes of the activity of the mutant-rice Rubisco during photosynthesis are closely related to the special protein in the leaves and its structure and components, and possibly to the regulating protein of Rubisco. The protein particularly regulates metabolic processes of amino acids preventing regulation of the preceding amino acids, such that the formation of the structural material in chloroplast is prevented, and finally the chloroplast thylakoid structure degenerates.

1103s was a new chlorophyll national mutant,of which leaves and sheaths would tum into green-yellow band when treated with varied temperature.This paper summarized the main research achievernent of this mutant trait.

用合成的cry1Ac基因与绿色荧光蛋白基因(GFP)构成融合蛋白基因,然后和改造的GNA基因构建双价抗虫基因植物表达载体pBGbfg,经根癌农杆菌介导转化了烟草.在紫外灯照射下,观察到转基因植株叶片中有较强的绿色荧光;经抗虫试验、PCR、Southern blot和Western blot等检测,表明该重组植物表达载体能够在转基因植物中有效表达外源基因,转基因植株绿色荧光的表型与其抗虫性密切相关.从而成功地建立了以绿色荧光蛋白基因与抗虫基因组成的融合基因转化系统,简化了抗虫转基因植物筛选程序,有助于快速获得双价抗虫转基因植株.

FtsZ蛋白在叶绿体的分裂过程中担负着重要作用.高等植物中FtsZ蛋白的陆续发现和功能分析加深了人们对叶绿体分裂机制的认识.植物中的ftsZ基因可分为两个明显不同的家族,在对烟草ftsZ1家族的研究基础上,对烟草ftsZ2家族成员的NtFtsZ2-1基因做了进一步的功能分析.研究结果表明,NtFtsZ2-1在叶绿体的分裂过程中发挥着重要作用,该基因的过量与减弱表达均对叶绿体的形态和数目产生了明显的影响.

棉花抗病虫基因工程和纤维品质改良一直是国内外研究的热点之一,近年来许多实验室又开展了棉花功能基因组学研究,在这些研究中都需要提取高质量的RNA,但目前广为使用的TriZol reagent kit和异硫氰酸胍-酚-氯仿一步提取法从棉花中难以纯化出完整的RNA,尽管许多学者针对棉花也发展出了几种有效的方法,但是这些方法一般都费工、费时.本文综合比较这些技术,并对热CTAB法加以改进,提出了一种较为可行的提取高质量棉花总RNA的简便方法.

FtsZ protein plays an important role in the division of chloroplasts. With the finding and functional analysis of higher plant FtsZ proteins, people have deepened the understanding in the molecular mechanism of chloroplast division. Multiple ftsZ genes are diversified into two families in higher plants, ftsZ1 and ftsZ2. On the basis of the research on ftsZ1 family, we analyzed the function of NtFtsZ2-1 gene in Nicotiana tabacum. Microscopic analysis of the sense and antisense NtFtsZ2-1 transgenic tobacco plants revealed that the chloroplasts were abnormal in size and also in number when compared with wild-type tobacco chloroplasts. Our investigations confirmed that the NtFtsZ2-1 gene is involved in plant chloroplast division.

将具有广谱抗病作用的葡萄糖氧化酶(glucose oxidase,GO)基因插入具有潮霉素抗性选择标记的双元载体pCAMBIA1301,新构建了水稻高效表达载体pCAG1301.将此质粒导入根癌农杆菌(Agrobacterium tumefaciens)菌株LBA4404后,转化粳稻(Oryza.sativa L.ssp.japonica)品种日本晴(Nipponbare)的幼胚,并由筛选出的潮霉素抗性愈伤组织分化再生植株.对所得潮霉素抗性水稻植株的Southem杂交分析表明,GO基因已整合到受体基因组.淀粉-碘化钾显色反应检测到了转基因植株产生的过氧化氢,证实GO基因表达产生的葡萄糖氧化酶已经在水稻中发挥功能.抗病性鉴定表明,所得转基因水稻对稻瘟菌(Magnaporthegrisea)具有良好的抗性.

质体是植物细胞中一类重要的细胞器, 其正常分裂过程与植物细胞的分化和发育密切相关. 人们对于质体分裂的早期研究主要集中于质体分裂过程的形态学观察和对某些突变体材料的遗传学分析, 而对于控制质体分裂的分子基础尚无明晰的认识. 近年来, 随着原核细胞分裂基因类似物在植物中的发现以及对质体进化祖先原核生物细胞分裂机制的解析, 极大地促进了人们对质体分裂机制的认识. 通过对分裂相关基因的功能研究, 人们认为作为内共生产物的质体可能与其原核祖先具有相似的分裂机制, 特别是对在分裂过程中发挥关键作用的ftsZ基因功能的深入研究为人们从分子水平上认识质体分裂的机制奠定了坚实的基础. 本文对质体分裂的研究历史进行了简单的回顾, 并对近来质体分裂分子机制的研究进展做一简要评述.

Plastid is one of the most important cellular organelles, the normal division process of plastid is essential for the differentiation and development of plant cells. For a long time, morphological observations and genetic analyses to special mutants are the major research fields of plastid division, but the molecular mechanisms underlying plastid division are largely unknown. Because of the endosymbiotic origin, plastid division might have mechanisms in common with those involved in bacterial cell division. It has been proved that several prokaryotic cell division genes also participate in the plastid division. Recently, the mechanisms of prokaryotic cell division have been well documented, which provides a valuable paradigm for understanding the plastid division mechanisms. In plants, the functional analyses of ftsZ , a key gene involved both in bacteria and plastid division, have established the solid foundation for people to understand the plastid division in molecular level. In this paper we will make a review for the research history and progress of plastid division.

In order to elucidate the origin of the plastid division gene ftsZ in green plant lineage, and to understand the significance of this divergence for the function of FtsZ proteins in plants, two full-length cDNAs (accession numbers AF449446 and AB084236) were isolated from Chlamydomonas reinhardtii, a base species of green plant lineage. A phylogenetic analysis based on amino acid sequences of eukaryotic FtsZs reveals that an ancient duplication of the ftsZ gene occurred after the endosymbiotic event. The ancient duplication implies that two ftsZ families might play an indispensable role at the early endosymbiotic stage.

通过PCR扩增,克隆了黑曲霉编码葡萄糖氧化酶的GO基因.构建了CaMV35S启动子驱动的GO基因植物表达载体,经农杆菌介导的遗传转化,获得了转基因烟草植株.经Southern,Northern和Western杂交分析,证明了GO基因在转基因烟草中的整合、转录和翻译.转基因烟草中H2O2的含量最高可达610μmol/L,比对照提高近5倍.嫁接试验证实H2O2可由砧木远距离运输至接穗,但不能由接穗转运至砧木,说明H2O2是通过导管由下向上运输.非转基因接穗嫁接至转基因砧木上3周,接穗上不同节位叶片中H2O2的含量不存在浓度梯度,说明这是一种主动运输而不是被动扩散.

The NDRl (nonrace-specific disease resistance) gene is required in Arabidopsis resistance to both bacterial and fungal pathogen mediated by several R genes. Expression of NDRl in transgenic plants confers resistance to a broad variety of plant pathogens. The NDRl gene from Arabidopsis Wassilewskija cultivation was cloned. In contract to the gene of Columbia cultivation, this gene has 7 nucleic acids and 4 amino acids mutations, respectively. A plant high effective expression vector was constructed and transformed to tobacco by agrobacterial-mediated transformation. Analysis of PCR and southern blot showed that NDRl gene has been integrated in tobacco genome. Three of ten transgenic plants selected randomly have obviously improved its resistance to Altemaria alternata and Phytophthora infestans.

As an important group of plant cellular organelles, the molecular mechanism of plastid division is poorly understood. Recent studies have revealed that the homologs of ftsZ gene, an essential prokaryotic cell division gene, are involved in plastid division process of plant cells. Antisense and sense expression constructions were employed to investigate the functions of the two ftsZ genes, NtFtsZ1 and NtFtsZ2, in transgenic Nicotiana tabacum L. plants. Although antisense expression of NtFtsZs reduced the native protein level obviously, the size and number of chloroplasts in transgenic tobacco plants had no effect. In contrast, overexpression of NtFtsZs in transgenic plants strikingly changed the number and morphology of chloroplasts. Even only 1-2 huge chloroplasts could be seen in the mesophyll cells of some overexpression transgenic plants. Analyses of chloroplast ultrastructures and chlorophyll content of different transgenic plants suggested that NtFtsZs gene have no direct influence on the normal development and function of chloroplasts. The changes in chloroplast morphology must be a compensation for the change in chloroplast number. The different phenotypes of chloroplasts in antisense and sense transgenic plants implied that different members from the same ftsZ gene family may have similar function in controlling plastid division. Meanwhile, the changes of chloroplast morphology in sense transgenic plants represented the possible plastoskeleton function of ftsZ in higher plant.

A full-length cDNA of GlFtsZ was isolated by screening the cDNA library of Gentiana lutea. Analysis of the deduced amino acid sequence encoded by GlFtsZ indicated that GlFtsZ protein possesses the typical conservative motifs existed in all FtsZ proteins. The existence of putative plastid transit peptide in its N-terminus suggested that GlFtsZ might function inside of plastids. With the developmental process of petals of Gentiana lutea, the expression of plastid division gene GlFtsZ declined gradually, whereas the expression of carotenoids biosynthesis gene Zds increased obviously; meanwhile, in contrast to the increment of carotenoids, the content of chlorophyll in petals decreased sharply. The chloroplasts turned into chromoplasts, and the color of petals also turned from green to golden. All of these results suggested that the expression of GlFtsZ is accompanied with the development and differentiation of plastids.

FtsZ蛋白在细菌的分裂中担任着重要作用,能够在分裂位点形成一个环状结构而控制细菌的分裂过程.胞内FtsZ蛋白浓度的明显降低或异常升高均可阻断正常的细胞分裂过程进而导致丝状菌体的产生.我们为了研究烟草FtsZ蛋白与大肠杆菌FtsZ蛋白的异同,构建了烟草全长ftsZ2-1与绿色荧光蛋白EGFP的融合表达质粒并转化大肠杆菌JM109.融合表达质粒的过量表达导致宿主菌形成了丝状菌体.通过荧光显微镜观察发现NtFtsZ2-1-EGFP融合蛋白沿着宿主菌体的纵轴方向有规律地聚集成荧光点或荧光带,说明烟草FtsZ2-1蛋白能够识别宿主菌内分裂位点的定位信号并参与其细胞分裂复合物的组装.