共查询到20条相似文献,搜索用时 890 毫秒
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Characterization of a plant scaffold attachment region in a DNA fragment that normalizes transgene expression in tobacco. 总被引:22,自引:3,他引:19 下载免费PDF全文
Using a low-salt extraction procedure, we isolated nuclear scaffolds from tobacco that bind specific plant DNA fragments in vitro. One of these fragments was characterized in more detail; this characterization showed that it contains sequences with structural properties analogous to animal scaffold attachment regions (SARs). We showed that scaffold attachment is evolutionarily conserved between plants and animals, although different SARs have different binding affinities. Furthermore, we demonstrated that flanking a chimeric transgene with the characterized SAR-containing fragment reduces significantly the variation in expression in series of transformants with an active insertion, whereas a SAR fragment from the human beta-globin locus does not. Moreover, the frequency distribution patterns of transgene activities showed that most of the transformants containing the plant SAR fragment had expression levels clustered around the mean. These data suggest that the particular plant DNA fragment can insulate the reporter gene from expression-influencing effects exerted from the host chromatin. 相似文献
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In an analysis of a 90-kb region around the human beta-globin gene complex we have identified at least eight sites of attachment to the nuclear scaffold (SARs). While these have many potential functions, there appears to be a particular association with sequences important in the regulation of the complex. Two SARs are close to the known enhancer-like elements of the beta-globin gene. SARs flanking the complex co-habit with the boundaries of the putative beta-like globin gene regulatory domain. In contrast, we have detected no SARs within a 140-kb region of the human alpha-globin gene complex. If SARs play a role in the regulation of gene expression then this structural difference would imply a difference in the regulation of the two complexes. 相似文献
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High-level transgene expression in plant cells: effects of a strong scaffold attachment region from tobacco. 总被引:10,自引:0,他引:10 下载免费PDF全文
G C Allen G Hall Jr S Michalowski W Newman S Spiker A K Weissinger W F Thompson 《The Plant cell》1996,8(5):899-913
We have previously shown that yeast scaffold attachment regions (SARs) flanking a chimeric beta-glucuronidase (GUS) reporter gene increased per-copy expression levels by 24-fold in tobacco suspension cell lines stably transformed by microprojectile bombardment. In this study, we examined the effect of a DNA fragment originally identified in a tobacco genomic clone by its activity in an in vitro binding assay. The tobacco SAR has much greater scaffold binding affinity than does the yeast SAR, and tobacco cell lines stably transformed with constructs containing the tobacco SAR accumulated greater than fivefold more GUS enzyme activity than did lines transformed with the yeast SAR construct. Relative to the control construct, flanking the GUS gene with plant SARs increased overall expression per transgene copy by almost 140-fold. In transient expression assays, the same construct increased expression only approximately threefold relative to a control without SARs, indicating that the full SAR effect requires integration into chromosomal DNA. GUS activity in individual stable transformants was not simply proportional to transgene copy number, and the SAR effect was maximal in cell lines with fewer than approximately 10 transgene copies per tobacco genome. Lines with significantly higher copy numbers showed greatly greatly reduced expression relative to the low-copy-number lines. Our results indicate that strong SARs flanking a transgene greatly increases expression without eliminating variation between transformants. We propose that SARs dramatically reduce the severity or likelihood of homology-dependent gene silencing in cells with small numbers of transgenes but do not prevent silencing of transgenes present in many copies. 相似文献
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Scaffold attachment regions increase reporter gene expression in stably transformed plant cells. 总被引:19,自引:2,他引:17 下载免费PDF全文
G C Allen G E Hall Jr L C Childs A K Weissinger S Spiker W F Thompson 《The Plant cell》1993,5(6):603-613
The yeast ARS-1 element contains a scaffold attachment region (SAR) that we have previously shown can bind to plant nuclear scaffolds in vitro. To test effects on expression, constructs in which a chimeric beta-glucuronidase (GUS) gene was flanked by this element were delivered into tobacco suspension cells by microprojectile bombardment. In stably transformed cell lines, GUS activity averaged 12-fold higher (24-fold on a gene copy basis) for a construct containing two flanking SARs than for a control construct lacking SARs. Expression levels were not proportional to gene copy number, as would have been predicted if the element simply reduced position effect variation. Instead, the element appeared to reduce an inhibitory effect on expression in certain transformants containing multiple gene copies. The effect on expression appears to require chromosomal integration, because SAR constructs were only twofold more active than the controls in transient assays. 相似文献
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Chromatin integrity is maintained throughout the cell cycle through repair mechanisms and intrinsically by the ordered packaging of DNA in association with histone proteins; however, aberrant rearrangements within and between chromosomes do occur. The role of the nuclear matrix protein topoisomerase II (TopoII) in generating chromosome breakpoints has been a focus of recent investigations. TopoII preferentially binds in vitro to scaffold-associated regions (SARs) and is involved in many DNA processing activities that require chromosome untangling. SARs, biochemically defined DNA elements rich in A + T, have been proposed to serve as structural boundaries for chromatin loops and to delineate functional domains. In our investigation of gene compartmentalization in a eukaryotic genome, SAR-associated nucleotide motifs from Drosophila were mapped in the regions of three histone gene clusters in an in silico analysis of the genome of Caenorhabditis elegans. Sites with similarity to the 15 bp consensus for TopoII cleavage were found predominantly in A + T enriched intergenic regions. Reiteration of sites matching the TopoII core consensus led to the identification of a novel core histone gene on chromosome IV and provided evidence for duplication and inversion in each of the three histone gene clusters. Breakpoint analysis of DNA flanking reiterated regions revealed potential sites for TopoII cleavage and a base composition phenomenon suggestive of a trigger for inversion events. 相似文献
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N. Maira T. M. Torres A. L. de Oliveira S. R. B. de Medeiros L. F. Agnez‐Lima J. P. M. S. Lima K. C. Scortecci 《Plant biology (Stuttgart, Germany)》2014,16(3):622-631
Unlike bacteria and mammals, plant DNA repair pathways are not well characterised, especially in monocots. The understanding of these processes in the plant cell is of major importance, since they may be directly involved in plant acclimation and adaptation to stressful environments. Hence, two sugarcane ESTs were identified as homologues of AP endonuclease from the base‐excision repair pathway: ScARP1 and ScARP3. In order to understand their probable function and evolutionary origin, structural and phylogenetic studies were performed using bioinformatics approaches. The two predicted proteins present a considerable amino acid sequence similarity, and molecular modelling procedures indicate that both are functional, since the main structural motifs remain conserved. However, inspection of the sort signal regions on the full‐length cDNAs indicated that these proteins have a distinct organelle target. Furthermore, variances in their promoter cis‐element motifs were also found. Although the mRNA expression pattern was similar, there were significant differences in their expression levels. Taken together, these data raise the hypothesis that the ScARP is an example of a probable gene duplication event that occurred before monocotyledon/dicotyledon segregation, followed by a sub‐functionalisation event in the Poaceae, leading to new intracellular targeting and different expression levels. 相似文献
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A cDNA and genomic DNA encoding an abscisic acid responsive gene (ASR) homologue (Asr1) was isolated from an inodorus melon, Cucumis melo var. kuwata, cDNA and genomic library. The Asr1 gene showed the strongest fruit-specific expression and differential expression profiles during fruit development, which were expressed from a low copy gene. The promoter region of the Asr1 gene contained several putative functional cis-elements, which may be involved in the response to plant hormones and environmental stresses. These results suggest that Asr1 may play an important role in the regulation of melon fruit ripening. 相似文献
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The R2R3-MYB transcription factor gene family in maize 总被引:2,自引:0,他引:2
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Large volumes of genomic data have been generated for several plant species over the past decade, including structural sequence data and functional annotation at the genome level. Various technologies such as expressed sequence tags (ESTs), massively parallel signature sequencing (MPSS) and microarrays have been used to study gene expression and to provide functional data for many genes simultaneously. This review focuses on recent advances in the application of microarrays in plant genomic research and in gene expression databases available for plants. Large sets of Arabidopsis microarray data are publicly available. Recently developed array platforms are currently being used to generate genome-wide expression profiles for several crop species. Coupled to these platforms are public databases that provide access to these large-scale expression data, which can be used to aid the functional discovery of gene function. 相似文献
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Functional network analysis of genes differentially expressed during xylogenesis in soc1ful woody Arabidopsis plants 下载免费PDF全文
Nicolas Davin Patrick P. Edger Charles A. Hefer Eshchar Mizrachi Mathias Schuetz Erik Smets Alexander A. Myburg Carl J. Douglas Michael E. Schranz Frederic Lens 《The Plant journal : for cell and molecular biology》2016,86(5):376-390
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M A Sobol' 《T?Sitologii?a i genetika》2001,35(3):72-84
Contemporary data on ultrastructural reconstruction and changes in functional nucleolus activity in plant cells affected by physical environmental factors are presented. The main attention is paid to modifications of ribosomal gene expression under these conditions and induced changes in r-chromatin structure and ribosomal DNA localization. Recent data on fine nucleolus structure and molecular aspects of its organization as well as influence of microgravitation and clinostating on structural and functional organization of plant nucleoli are reported. 相似文献
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The recent discovery of DNA sequences responsible for the specific attachment of chromosomal DNA to the nuclear skeleton (MARs/SARs) was an important step towards our understanding of the functional and structural organization of eukaryotic chromatin [Mirkovitch et al.: Cell 44:273-282, 1984; Cockerill and Garrard: Cell 44:273-282, 1986]. A most important question, however, remains the nature of the matrix proteins involved in the specific binding of the MARs. It has been shown that topoisomerase II and histone H1 were capable of a specific interaction with SARs by the formation of precipitable complexes [Adachi et al.: EMBO J8:3997-4006, 1989; Izaurralde et al.: J Mol Biol 210:573-585, 1989]. Here, applying a different approach, we were able to "visualize" some of the skeletal proteins recognizing and specifically binding MAR-sequences. It is shown that the major matrix proteins are practically the same in both salt- and LIS-extracted matrices. However, the relative MAR-binding activity of the individual protein components may be different, depending on the method of matrix preparation. The immunological approach applied here allowed us to identify some of the individual MAR-binding matrix proteins. Histone H1 and nuclear actin are shown to be not only important components of the matrix, but to be involved in a highly efficient interaction with MAR-sequences as well. Evidence is presented that proteins recognized by the anti-HMG antibodies also participate in MAR-interactions. 相似文献
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