香蕉束顶病毒复制酶基因克隆及转基因表达

以广州市郊获得的香蕉束项病毒(BBTV)的DNA为模板,进行PCR扩增得到香蕉束项病毒复制酶基因的1.1 kb DNA.所获得的DNA序列与澳大利亚的BBTV序列的同源性达90%,这部分序列编码香蕉束顶病毒复制酶基因的羧基端.将改造的BBTV复制酶基因克隆到pBll21的CaMV 35S和NOS终止序列之间,构建表达载体,并采用基因枪轰击香蕉试管苗生长点组织的方法,经PCR检测和Westem blot分析,获得4株具有BBTV复制酶基因整合表达的To代转基因香蕉.转基因植株的抗病性正在检测之中.     

香蕉转基因研究进展

香蕉转基因研究逐渐成为国内外植物分子生物学研究的热点。人们从 2 0世纪 60年代开始就进行了香蕉的组织培养 ,70年代茎尖培养成功 ,近 1 0年来相继在微繁殖、单克隆抗体、分子生物学和遗传转化等方面开展了工作。香蕉转化受体系统已从茎尖发展到胚状体和愈伤组织受体系统 ,遗传转化方法也在不断改进 ,从单一的方法发展为根癌农杆菌感染法与基因枪法的结合。从香蕉组织培养、遗传转化受体系统、基因转化方法、筛选方法的研究等方面进行综述 ,并提出了一些看法。     

ISAAA信息

用转基因拯救香蕉产业昆士兰理工大学(QUT)JamesDale等科学家正在种植能在热带4号真菌下存活的转基因香蕉。热带4号是一种存在于土壤中的真菌Fusarium,它由于生存在土壤中数十年并且难以被化学药品消灭而给种植者带来麻烦。     

ISAAA信息

抗黑叶条斑病的转基因香蕉转基因香蕉(MUSA ACUMINATA)GROS MICHEL携带一或二个水稻几丁质酶基因,是由比利时鲁汶大学GABRIELLA KOVCS团队培育的。通过叶盘法(LEAF DISK ASSAY),研究小组检测了转基因香蕉对黑叶条斑病(MYCOSPHAERELLA FIJIENSIS)的抗     

香蕉乙二醛酶基因增强酿酒酵母对非生物胁迫抵抗能力的研究

从香蕉cDNA文库中克隆到了一个香蕉(Musa acuminata AAA subgroup)乙二醛酶(glyoxalase,GLO)基因(MaGLO14)。构建了带有MaGLO14的酵母表达载体PYES2-MaGLO14,转化酿酒酵母(Saccharomyces cerevisiae)尿嘧啶营养缺陷型菌株INVSC1,挑取转化子进行PCR和酶切鉴定,证实获得了转基因菌株。通过比较转基因菌株和非转基因菌株在NaCl、高温、低温、干旱、UV胁迫下的生长状况,证明转基因菌株在以上非生物胁迫条件下的存活菌落数均高于非转基因菌株。利用酿酒酵母初步证明MaGLO14具有增强酵母菌对非生物胁迫抵抗力的功能。     

香蕉抗病基因工程

长期以来,香蕉生产遭受病害的严重威胁,制约了其发展,目前,随着转基因抗病香蕉基因工程技术的日趋成熟,为培育抗病香蕉品种开辟了新途径。     

根癌农杆菌介导的香蕉遗传转化研究进展

香蕉的栽培品种绝大多数是三倍体, 常规育种难度大, 工作周期长, 难以采用传统的育种方法进行遗传改良, 因此转基因技术成为改良香蕉种质的有效方法。根癌农杆菌介导的香蕉的遗传转化从20世纪90年代起取得的了较大的进步, 但仍然存在着很多问题。文章分析了影响农杆菌介导香蕉转化的几个重要环节, 并对其研究现状、存在的问题及应用前景作简要概述。     

香蕉生物技术研究现状与展望

本文对香蕉生物技术研究现状进行了全面的概括,包括微繁殖、脱病毒、人工种子、诱变育种、单克隆抗体用于病毒检测、转基因受体及基因转化等,并对今后香蕉生物技术的发展重点进行了讨论。     

烤烟栽培品种G28和K326抗黄瓜花叶病毒的遗传转化

采用根癌农杆菌介导的叶盘转化法,将ＣＭＶＢＳ（香蕉株系）的ＣＰ基因转入广东烤烟栽培品种Ｇ２８和Ｋ３２６,获得了再生转基因烟株。在卡那霉素（Ｋｍ）１００μｇ／ｍＬ浓度下转化体叶片分化形成愈伤组织;用ＰＣＲ和ＤＡＳＥＬＩＳＡ分析,结果证明ＣＰ基因已整合到转化烟株的染色体中,并得到不同程度的表达。利用广东地区香蕉、烟草和番茄上的ＣＭＶＢＳ、２７和３７株系,采用机械接种和蚜虫传毒法分别进行攻毒试验,结果证明转基因烟株对这三个株系的攻击都表现出明显的抗病能力,其抗性程度与烟株中的ＣＰ表达量呈正相关。采用五种攻毒浓度处理,发现随着攻毒浓度的增加,转基因烟株的抗病能力逐渐减弱。     

富含维生素A的转基因香蕉将在非洲推广

<正>据国外媒体报道,澳大利亚科学家培育了一种富含维生素A的转基因香蕉,它或许能够应对导致全世界数十万人死亡的营养不良问题,而且能够减少全世界发生的婴儿失明问题。这项转基因技术能够增加东非地区某种煮食香蕉中的β-胡萝卜素(进入人体后可转变成维生素A,因此     

Construction and characterization of a plant transformation-competent BIBAC library of the black Sigatoka-resistant banana Musa acuminata cv. Tuu Gia (AA)

A plant transformation-competent binary bacterial artificial chromosome (BIBAC) library was constructed from Musa acuminata cv. Tuu Gia (AA), a black Sigatoka-resistant diploid banana. After digestion of high-molecular-weight banana DNA by HindIII, several methods of DNA size selection were tested, followed by ligation, using a vector/insert molar ratio of 4:1. The library consists of 30,700 clones stored in 80 384-well microtiter plates. The mean insert size was estimated to be 100 kb, and the frequency of inserts with internal NotI sites was 61%. The majority of insert sizes fell into the range of 100±20 kb, making them suitable for Agrobacterium-mediated transformation. Only 1% and 0.9% of the clones contain chloroplast and mitochondrial DNA, respectively. This is the first BIBAC library for banana, estimated to represent five times its haploid genome (600 Mbp). It was demonstrated by hybridization that the library contains typical members of resistance gene and defense gene families that can be used for transformation of disease susceptible banana cultivars for banana genetic improvement.     

巴西蕉高效间接器官发生再生体系研究

香蕉间接器官发生再生体系是一种较好的突变育种和基因转化受体再生系统。本研究以巴西蕉低代试管苗茎段薄层切片、中代试管苗茎段薄层切片、吸芽薄层切片、雄花薄层切片为外植体,对巴西蕉间接器官发生再生体系进行了系统研究。研究结果表明,以MS为基本培养基,巴西蕉低代试管苗茎段薄层切片在0.5～1.0mg/L2,4-D+4～6mg/L6-BA激素条件下;雄花薄层切片在0.5～1.0mg/L2,4-D+0.2～0.5mg/LZT激素条件下能通过花菜体间接器官发生途径实现高效再生;同时,花菜体薄层切片在1.0mg/L2,4-D+5mg/L6-BA激素条件下,再次通过花菜体途径实现循环高效再生。本研究为建立香蕉高效间接器官发生再生体系和相关育种技术奠定了基础。     

香蕉生物技术研究进展

近50年来香蕉的生产得到了稳步的发展,同时正遭受越来越严重的病虫、霜冻和台风的侵害。鲜食蕉雌雄性高度不育的特性,使得传统的育种方法难以进行香蕉的遗传改良。这些现状迫切要求香蕉生物技术的不断发展和深入。近10年来,在香蕉体细胞胚胎发生、原生质体培养、基因克隆和序列分析以及基因转化等方面都取得了可喜的成果,并预计于2006年完成香蕉基因组的测序 。     

Construction of banana bunchy top nanovirus-DNA-3 encoding the coat protein gene and its introducing into banana plants cv. Williams

Banana (Musa sp.) is considered as one of the most important fruit crops worldwide as well as in Egypt. The main goal of this study was to construct the open reading frame (ORF) of banana bunchy top nanovirus (BBTV)-DNA-3 that encodes the viral coat protein (cp) gene for banana transformation. The previously sequenced BBTV-G-DNA-3-ORF that cloned into plasmid pH1 was used as a template for PCR amplification using two specific primers containing Bam H1 site. A new plasmid called pRHA1 containing the amplified ORF under the control of maize polyubiquitin (ubi) promoter was created. The bar gene (herbicide-resistance gene as a selectable marker) cassette (bar gene, Cauliflower mosaic caulimovirus (CaMV) 35S promoter and nos terminator) was released from plasmid pAB6 using Hind III-digestion and subcloned into the Hind III-digested plasmid pRHA1 to create the plasmid pRHA2 via the microprojectile bombardment transformation system. The plasmid pRHA2 was successfully introduced in the applied banana cultivar. Leaf painting test was conducted to confirm the expression of the bar gene in the putative transformed banana lines. The presence and expression of BBTV-G-cp gene were also detected using some molecular (polymerase chain reaction and dot blot using a cold DNA probe) and serological (ELISA and western blot) techniques, respectively, in the obtained transgenic banana lines.     

香蕉基因工程研究进展

香蕉是世界上一种重要的作物.基因工程技术的发展为培育具有优良性状的香蕉品种提供了有效的手段.本文综述了近年来香蕉基因工程研究的最新进展,包括香蕉遗传转化受体系统的建立,香蕉作物中一些有益功能基因的克隆以及香蕉遗传转化的最新科研成果.并对各种香蕉遗传转化受体系统和目前主要的香蕉遗传转化进行了客观评价,提出了香蕉遗传转化研究中存在的问题,最后对通过基因工程手段改良香蕉品种进行了展望.     

Establishment of embryogenic cell suspension cultures and Agrobacterium-mediated transformation in an important Cavendish banana cv. Robusta (AAA)

Establishment of an efficient protocol for regeneration and genetic transformation is required in banana for the incorporation of useful traits. Therefore an efficient method has been developed for somatic embryogenesis, plant regeneration and transformation of Cavendish banana cultivar Robusta (AAA). Embryogenic cell suspension culture (ECS) was established using immature male flowers. Percentage appearance of embryogenic callus and distinct globular embryos was 10.3 and 11.1, respectively. ECS obtained was cocultivated under different cocultivation conditions with Agrobacterium tumefaciens strain EHA105 harboring pCAMBIA 1301 plant expression vector. Up to 30 transgenic plants/50 mg settled cell volume (SCV) was obtained with cocultivation in semisolid medium whereas no transgenics could be obtained with parallel experiments carried out in liquid medium. Histochemical GUS assay in different tissues of putatively transformed plants demonstrated expression of uidA gene. Among the putatively transformed plants obtained, a set of 4 were confirmed by PCR analysis and stable integration of the transgene by Southern analysis. GUS specific activity measured by a MUG (4-methylumbelliferyl-β-d-glucuronide) based flourometric assay revealed increase in transient GUS expression in semisolid as well as liquid cocultivation with centrifugation. This is the first report showing somatic embryogenesis and Agrobacterium tumefaciens mediated transformation using embryogenic cell suspension cultures in an important Cavendish banana cultivar Robusta. The present protocol will make possible agronomic improvement of this important commercially grown cultivar by introduction of disease resistance characteristics and antisense-mediated delayed fruit ripening strategies. Further, it will also assist in functional characterization of new gene or promoter elements isolated from this or other cultivars of banana.     

Biotechnology of the Banana: A Review of Recent Progress

Abstract: A number of biotechnological tools have been developed which could help breeders to evolve new plant types to meet the demand of the food industry in the next century. Available techniques for the transfer of genes could significantly shorten the breeding procedures and overcome some of the agronomic and environmental problems which would otherwise not be possible through conventional methods. In vitro protocols have been standardized to allow commercially viable propagation of desired clones of Musa. An overview of the regeneration of banana by direct and indirect organogenesis, and somatic embryogenesis is presented in this article. In addition, the use of several other biotechnological techniques to enrich the genome of banana, such as selection of somaclonal variants, screening for various useful characteristics, cryopreservation, genetic transformation and molecular genetics are reviewed. In conclusion, the improvement of banana through modern biotechnology should help ensure food security by stabilizing production levels in sustainable cropping systems geared towards meeting domestic and export market demands.