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A 577-bp promoter segment of Agrobacterium rhizogenes rolC, previously known as the phloem-specific gene expression promoter, was fused to the 5′ end of a reporter gene, β-glucuronidase
(GUS), uidA. This rolC-promoter-driven expression of the GUS gene was found to be significantly strong in glandular cells in transgenic tobacco plants. Analysis of this segment of the
promoter sequence revealed a myb response element. 相似文献
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Characterization of the regulatory elements of the maize P-rr gene by transient expression assays 总被引:2,自引:0,他引:2
The maize P-rr gene conditions floral-specific flavonoid pigmentation, especially in the kernel pericarp and cob. We analyzed the P-rr promoter by transient expression assays, in which segments of the P-rr promoter were fused to the GUS reporter gene and introduced into maize cells by particle bombardment. A basal P-rr promoter fragment (–235 to +326) gave low, but significant, levels of GUS reporter gene expression. Interestingly, two widely spaced segments containing enhancer-like activity were found. When tested individually, both the proximal (–1252 to –236) and distal (–6110 to –4842) segments boosted expression of the basal P-rr promoter::GUS construct about five-fold. A 1.6 kb segment of the P-rr promoter (–1252 to +326) containing the proximal enhancer and the 5-untranslated leader driving the GUS reporter gene showed preferential expression in BMS and embryogenic suspension cell cultures vs. endosperm-derived suspension cell cultures. These results demonstrate the application of transient assay techniques for the identification of regulatory elements responsible for floral-specific regulation of the complex P-rr gene promoter in maize. 相似文献
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The presence of expansins was investigated in various developmental and ripening stages of cherry fruits by SDS-PAGE and immunoblotting.
An expansin gene and three fragments (242, 607 and 929 bp) of its promoter region were cloned. The genomic clone of the expansin
gene contained three introns, two exons spanning a 1.6 and a 1.0 kb upstream region. Semi-quantitative PCR analysis showed
that this gene was ripening specific. Chimeric promoter—GUS constructs were made and truncated forms of the expansin promoter
were introduced into tomatoes by agroinjection and fruits were analyzed for GUS expression by histochemical GUS staining and
enzyme activity assays. The 0.60 kb expansin promoter efficiently induced GUS expression in transgenic tomatoes, whereas constructs
with the 0.25 kb promoter did not display significant GUS staining. The highest GUS activity was detected in tomatoes containing
the 1.0 kb promoter construct. Both large base pair promoter constructs drove the expression of the GUS gene at an equal or
higher rate than the tomato E8 promoter. 相似文献
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缺铁是世界范围内农业生产面临的严重问题,玉米通过分泌脱氧麦根酸(2’-deoxymugineic acid, DMA)吸收利用土壤中的难溶性铁。为探明玉米DMA分泌通道蛋白基因YS3的表达和调控机制,本文通过克隆获得长为2813 bp的YS3基因启动子,该序列含有大量TATA-box、CAAT-box等启动子基本元件,以及光响应、激素调控等多个顺式调控元件;构建YS3启动子驱动GUS基因的植物表达重组载体pCAMBIA-YS3GUS,利用农杆菌介导转化拟南芥,获得pYS3::GUS转基因植株,对转基因植株进行GUS组织化学染色,并通过石蜡切片技术对转基因植株进行组织观察,分析pYS3::GUS转基因植株中YS3基因启动子的活性。结果表明,YS3启动子主要驱动GUS基因在拟南芥根部表达,且主要集中在根部表皮细胞,机械损伤可激发YS3启动子活性,驱动GUS基因在损伤临近部位表达。本研究对于理解玉米DMA分泌的分子调控机理方法od3 gmaigensuan有重要意义。 相似文献
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A 330 bp region of the spinach nitrite reductase gene promoter directs nitrate-inducible tissue-specific expression in transgenic tobacco 总被引:4,自引:0,他引:4
Rajeev Rastogi Eduard Back Alois Schneiderbauer Caroline G. Bowsher Barbara Moffatt Steven J. Rothstein 《The Plant journal : for cell and molecular biology》1993,4(2):317-326
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利用PCR技术从哥伦比亚型拟南芥基因组DNA中分离了AtSTP3绿色组织特异表达的启动子,序列分析表明,扩增片段(1774bp)与已报道序列的相应区域同源性达99.9%。将其与GUS报告基因融合在一起,构建了植物表达载体,并由农杆菌介导法导入水稻品种‘中花11’中。对转基因水稻植株中的GUS活性进行定性与定量测定结果表明,AtSTP3启动子可驱动GUS报告基因在转基因水稻植株叶片中特异性表达,而在根和种子等器官中不表达或表达活性极弱,AtSTP3启动子表现出明显的组织特异性。 相似文献
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利用PCR技术从毛白杨基因组DNA中扩增获得花器官发育相关的SEPALLATA2类似基因PtSEP25′侧翼约2.3kb的一段序列,经PlantCARE序列分析表明,该序列中含有启动子特征的保守序列及多种光应答元件,初步推测其为PtSEP2基因启动子.进一步以GUS为报告基因,构建了pPtSEP2 promoter::... 相似文献
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Isolation and functional analysis of a strong specific promoter in photosynthetic tissues 总被引:2,自引:0,他引:2
Constitutive promoters such as CaMV (cauliflower mosaic virus) 35S and nos (nopaline syn-thase) have been used extremely as useful tools in many plant transgenic researches. Because of lacking temporal and spatial regulation, constitutive promoters have a number of potential drawbacks in genetically improved crops[1]. For example, constitutive expression of viral capsid proteins in plants may increase the risk of transvapsidation or viral recombination to generate new strains of phytopathogen… 相似文献
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为了探明拟南芥内膜反向转运体AtNHX6基因的组织表达模式,从基因组中克隆了AtNHX6基因开放阅读框(ORF)上游侧翼调控区1 922bp序列,并成功构建AtNHX6基因启动子与GUS融合表达载体pCAM-BIA1381-proNHX6-GUS,通过农杆菌花序浸染法转化野生型拟南芥获得T3代纯合转基因拟南芥株系,经PCR检测扩增得到2 187bp目的条带。利用组织染色法鉴定转基因拟南芥的GUS表达模式发现,在子叶、下胚轴和花中GUS活性显著。在这些广泛表达的部位中,微管系统中的表达最为显著,真叶中只有局部检测到GUS表达;在根中GUS在根毛和侧根生长部位表达;在未成熟果荚中只有在果荚顶端和基部存在GUS活性,成熟果荚中只在果柄检测到GUS表达;在花中,雄蕊的花丝和花粉粒及雌蕊的柱头中检测到GUS表达。GUS染色分析结果表明,AtNHX6基因启动子与GUS的融合表达载体成功构建并正常启动GUS基因表达,且AtNHX6基因主要在拟南芥的子叶、下胚轴、根、花、果荚中的微管系统、根毛和侧根生长部位以及花丝、花粉、柱头中表达。 相似文献
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竹节花黄斑驳病毒启动子的缺失分析及功能 总被引:4,自引:0,他引:4
竹节花黄斑驳病毒(CoYMV)是侵染单子叶植物竹节花的一
种双链环状DNA病毒,它的启动子可介导外源基因在烟草韧皮部特异表达。为了研究其组织
特异性表达的最佳启动子区域,对CoYMV启动子进行了5′端五种不同长度的缺失分析,用不同长度的启动子片段与GUS基因及NOS3′端转录中止序列构建了全长启动子及5
个缺失启动子序列的六个嵌合GUS基因植物表达载体。利用农杆菌将上述嵌合基因转化烟草
外植体后,每种表达载体都获得了一批转基因烟草植株。转化再生烟草植株的PCR分析、GUS
酶活测定及GUS组织染色的结果表明六种类型的嵌合基因已整合到烟草染色体中,并有五种
表达出GUS活性。缺失到870bp的启动子比全长启动子(1040bp)的活性约高78%,870bp比585bp启动子介导的GUS活性略高但差别不明显,缺失到447和232时GUS活性有明显下
降,但仍具有韧皮部特异表达的特性。当缺失到TATA box附近的44bp时启动子丧失组织特
异性,GUS活性也降低到测不出来的水平。以上结果表明CoYMV启动子从转录起始位点上游
870bp~230bp及232bp下游区分别与启动子的活性和韧皮部组织特异性密切相关,870bp上游可能存在一个负调控序列,所以该启动子的活性和组织特异性的最佳调控区应在87
0bp或585bp的下游区。CoYMV启动子与35S启动子驱动GUS基因在烟草中表达的活性相比,
前者为后者的70%左右,考虑到前者仅在韧皮部细胞表达而后者为组成型表达,所以CoYMV启
动子在韧皮部的活性可能与35S启动子相当或更高。CoYMV启动子在其它转基因植物中驱动外
源基因表达的特点正在研究中。 相似文献
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To develop a strong constitutive gene expression system, the activities of ibAGP1 promoter and its transit peptide were investigated using transgenic Arabidopsis and a GUS reporter gene. The ibAGP1 promoter directed GUS expression in almost entire tissues including rosette leaf, inflorescence stem, inflorescence, cauline
leaf and root, suggesting that the ibAGP1 promoter is a constitutive promoter. GUS expression mediated by ibAGP1 promoter was weaker than that by CaMV35S promoter in all tissue types, but when GUS protein was targeted to plastids with
the aid of the ibAGP1 transit peptide, GUS levels increased to higher levels in lamina, petiole and cauline leaf compared to those produced by
CaMV35S promoter. The enhancing effect of ibAGP1 transit peptide on the accumulation of foreign protein was tissue-specific; accumulation was high in lamina and inflorescence,
but low in root and primary inflorescence stem. The transit peptide effect in the leaves was maintained highly regardless
of developmental stages of plants. The ibAGP1 promoter and its transit peptide also directed strong GUS gene expression in transiently expressed tobacco leaves. These
results suggest that the ibAGP1 promoter and its transit peptide are a strong constitutive foreign gene expression system for transgenesis of dicot plants. 相似文献
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Pollen-specific gene expression in transgenic plants: coordinate regulation of two different tomato gene promoters during microsporogenesis 总被引:29,自引:0,他引:29
To investigate the regulation of gene expression during male gametophyte development, we analyzed the promoter activity of two different genes (LAT52 and LAT59) from tomato, isolated on the basis of their anther-specific expression. In transgenic tomato, tobacco and Arabidopsis plants containing the LAT52 promoter region fused to the beta-glucuronidase (GUS) gene, GUS activity was restricted to pollen. Transgenic tomato, tobacco and Arabidopsis plants containing the LAT59 promoter region fused to GUS also showed very high levels of GUS activity in pollen. However, low levels of expression of the LAT59 promoter construct were also detected in seeds and roots. With both constructs, the appearance of GUS activity in developing anthers was correlated with the onset of microspore mitosis and increased progressively until anthesis (pollen shed). Our results demonstrate co-ordinate regulation of the LAT52 and LAT59 promoters in developing microspores and suggest that the mechanisms that regulate pollen-specific gene expression are evolutionarily conserved. 相似文献