Increasing loop domain size does not diminish effects of matrix attachment regions on transgene expression in tobacco cells in culture

Mendu, N.; Massel, Mara; Spiker, Steven

doi:10.1016/s0014-5793(01)02406-1

Cited by 7 publications

(11 citation statements)

References 5 publications

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“…Early experiments in animal cells indicated that MARs increased transgene expression and decreased transformantto-transformant variability in transgene expression (Stief et al, 1989;Bonifer et al, 1990;Phi-Van et al, 1990). Subsequent work in plant systems also demonstrated an increase in transgene expression and/or a reduction in variability (Breyne et al, 1992;Allen et al, 1993Allen et al, , 1996Mlynarova et al, 1994Mlynarova et al, , 1995Han et al, 1997;Liu and Tabe, 1998;Ü lker et al, 1999;Vain et al, 1999;Cheng et al, 2001;Mendu et al, 2001;Fukuda and Nishikawa, 2003;Mankin et al, 2003;Butaye et al, 2004;van der Geest et al, 2004). The observations of increases in expression and/or reductions in variability have not, however, been consistent.…”

Section: Introductionmentioning

confidence: 99%

“…Binary vectors used for transformation carried a GFP reporter gene flanked on both sides by the tobacco Rb7 MAR, which we have previously used (Allen et al, 1996;Ü lker et al, 1999;Mendu et al, 2001;Mankin et al, 2003). GFP levels were measured in protoplasts generated from the various transgenic lines and compared with levels in protoplasts generated from lines carrying control constructs without MARs.…”

Section: Introductionmentioning

confidence: 99%

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The Rb7 Matrix Attachment Region Increases the Likelihood and Magnitude of Transgene Expression in Tobacco Cells: A Flow Cytometric Study

2005

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Many studies in both plant and animal systems have shown that matrix attachment regions (MARs) can increase expression of transgenes in whole organisms or cells in culture. Because histochemical assays often indicate variegated transgene expression, a question arises: Do MARs increase transgene expression by increasing the percentage of cells expressing the transgene (likelihood), by increasing the level of expression in expressing cells (magnitude), or both? To address this question, we used flow cytometry to measure green fluorescent protein (GFP) expression in individual tobacco (Nicotiana tabacum) cells from lines transformed by Agrobacterium tumefaciens. We conclude that MAR-mediated overall increases in transgene expression involve both likelihood and magnitude. On average, cell lines transformed with the Rb7 MAR-containing vector expressed GFP at levels 2.0- to 3.7-fold higher than controls. MAR lines had fewer nonexpressing cells than control lines (10% versus 45%), and the magnitude of GFP expression in expressing cells was greater in MAR lines by 1.9- to 2.9-fold. We also show that flow cytometry measurements on cells from isogenic lines are consistent with those from populations of independently transformed cell lines. By obviating the need to establish isogenic lines, this use of flow cytometry could greatly simplify the evaluation of MARs or other sequence elements that affect transgene expression

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Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

The Rb7 Matrix Attachment Region Increases the Likelihood and Magnitude of Transgene Expression in Tobacco Cells: A Flow Cytometric Study

2005

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“…However, we observed some differences between the effect of control constructs and the effect of 5MGUS construct where the maize adh-1 MAR-based loop should not be reconstituted. This is not inconsistent with the loop model: the DNA delivery technique we used here is known to generate complex insertion sites (Christou, 1995(Christou, , 1997Mendu et al, 2001;Svitashev et al, 2002;Makarevitch et al, 2003;Romano et al, 2003). Thus, it is possible that the rearrangements of plasmid constructs occurring at insertion sites result in a MAR-based loop structure defined only with 5 0 MAR.…”

Section: Mar-based Loop Structuresmentioning

confidence: 55%

Enhancement of maize transformation efficiency by the use of maize matrix attachment regions

Torney

Partier²,

Barret³

et al. 2005

Plant Cell Tiss Organ Cult

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Improving genetic transformation efficiency is a major concern in plant genetic engineering. While various strategies have been investigated, the enhancement of selectable marker gene expression has not been tried extensively. We used maize matrix attachment regions (MARs) to bracket an herbicide resistance transgene, bar. MARs have been reported to enhance transgene expression level and stability. We show here that MARs not only enhance transformation efficiency by 50%, but are also able to increase or decrease relative efficiencies of each step of the regeneration process depending on MAR sequence combinations. Furthermore, we assessed the trans-effect of MARs in co-bombardment experiments with two independent plasmids, one including the MAR sequences and the other one the bar gene. As for simple bombardment, MARs enhanced transformation efficiency by having a positive influence on organogenesis step in the regeneration process.Abbreviations: adh-1 -alcohol dehydrogenase; 2,4-D D -2,4-dichlorophenoxyacetic acid; gus -b-glucuronidase; MAR -matrix attachment region; SATB1 -Special AT rich binding protein 1

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“…Scientists discovered the ability of DNA sequences called scaffold/matrix attachment region (S/MAR) that could stabilize transcription and shield transgene from being repressed by chromatin structure into which they incorporated [6][7][8][9]. The site where this chromatin fibers bind to matrix protein is called S/MAR [8,10].…”

Section: Introductionmentioning

confidence: 99%

“…The site where this chromatin fibers bind to matrix protein is called S/MAR [8,10]. Interaction between S/MAR and matrix protein causes the formation of a distinct loop domain which localize gene transcription units and specific regulatory elements to regulate transcription [4,11].…”

Section: Introductionmentioning

confidence: 99%

Mapping analysis of scaffold/matrix attachment regions (s/MARs) from two different mammalian cell lines

Pilus

Johari

Ahmad

et al. 2014

AIP Conference Proceedings

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Abstract. Scaffold/matrix attachment regions (S/MARs) are potential element that can be integrated into expression vector to increase expression of recombinant protein. Many studies on S/MAR have been done but none has revealed the distribution of S/MAR in a genome. In this study, we have isolated S/MAR sequences from HEK293 and Chinese hamster ovary cell lines (CHO DG44) using two different methods utilizing 2 M NaCl and lithium-3,5-diiodosalicylate (LIS). The isolated S/MARs were sequenced using Next Generation Sequencing (NGS) platform. Based on reference mapping analysis against human genome database, a total of 8,994,856 and 8,412,672 contigs of S/MAR sequences were retrieved from 2M NaCl and LIS extraction of HEK293 respectively. On the other hand, reference mapping analysis of S/MAR derived from CHO DG44 against our own CHO DG44 database have generated a total of 7,204,348 and 4,672,913 contigs from 2 M NaCl and LIS extraction method respectively.

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Increasing loop domain size does not diminish effects of matrix attachment regions on transgene expression in tobacco cells in culture

Cited by 7 publications

References 5 publications

The Rb7 Matrix Attachment Region Increases the Likelihood and Magnitude of Transgene Expression in Tobacco Cells: A Flow Cytometric Study

The Rb7 Matrix Attachment Region Increases the Likelihood and Magnitude of Transgene Expression in Tobacco Cells: A Flow Cytometric Study

Enhancement of maize transformation efficiency by the use of maize matrix attachment regions

Mapping analysis of scaffold/matrix attachment regions (s/MARs) from two different mammalian cell lines

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