@article{wear_song_zynda_mickelson-young_leblanc_lee_deppong_allen_martienssen_vaughn_et al._2020, title={Comparing DNA replication programs reveals large timing shifts at centromeres of endocycling cells in maize roots}, volume={16}, ISSN={["1553-7404"]}, DOI={10.1371/journal.pgen.1008623}, abstractNote={Plant cells undergo two types of cell cycles–the mitotic cycle in which DNA replication is coupled to mitosis, and the endocycle in which DNA replication occurs in the absence of cell division. To investigate DNA replication programs in these two types of cell cycles, we pulse labeled intact root tips of maize (Zea mays) with 5-ethynyl-2’-deoxyuridine (EdU) and used flow sorting of nuclei to examine DNA replication timing (RT) during the transition from a mitotic cycle to an endocycle. Comparison of the sequence-based RT profiles showed that most regions of the maize genome replicate at the same time during S phase in mitotic and endocycling cells, despite the need to replicate twice as much DNA in the endocycle and the fact that endocycling is typically associated with cell differentiation. However, regions collectively corresponding to 2% of the genome displayed significant changes in timing between the two types of cell cycles. The majority of these regions are small with a median size of 135 kb, shift to a later RT in the endocycle, and are enriched for genes expressed in the root tip. We found larger regions that shifted RT in centromeres of seven of the ten maize chromosomes. These regions covered the majority of the previously defined functional centromere, which ranged between 1 and 2 Mb in size in the reference genome. They replicate mainly during mid S phase in mitotic cells but primarily in late S phase of the endocycle. In contrast, the immediately adjacent pericentromere sequences are primarily late replicating in both cell cycles. Analysis of CENH3 enrichment levels in 8C vs 2C nuclei suggested that there is only a partial replacement of CENH3 nucleosomes after endocycle replication is complete. The shift to later replication of centromeres and possible reduction in CENH3 enrichment after endocycle replication is consistent with a hypothesis that centromeres are inactivated when their function is no longer needed.}, number={10}, journal={PLOS GENETICS}, author={Wear, Emily E. and Song, Jawon and Zynda, Gregory J. and Mickelson-Young, Leigh and LeBlanc, Chantal and Lee, Tae-Jin and Deppong, David O. and Allen, George C. and Martienssen, Robert A. and Vaughn, Matthew W. and et al.}, year={2020}, month={Oct} } @article{wear_song_zynda_leblanc_lee_mickelson-young_concia_mulvaney_szymanski_allen_et al._2017, title={Genomic Analysis of the DNA Replication Timing Program during Mitotic S Phase in Maize (Zea mays) Root Tips}, volume={29}, ISSN={["1532-298X"]}, url={http://europepmc.org/abstract/med/28842533}, DOI={10.1105/tpc.17.00037}, abstractNote={The time during S phase at which different maize DNA sequences replicate reveals a complex temporal program influenced by genomic features, transcriptional activity, and chromatin structure. All plants and animals must replicate their DNA, using a regulated process to ensure that their genomes are completely and accurately replicated. DNA replication timing programs have been extensively studied in yeast and animal systems, but much less is known about the replication programs of plants. We report a novel adaptation of the “Repli-seq” assay for use in intact root tips of maize (Zea mays) that includes several different cell lineages and present whole-genome replication timing profiles from cells in early, mid, and late S phase of the mitotic cell cycle. Maize root tips have a complex replication timing program, including regions of distinct early, mid, and late S replication that each constitute between 20 and 24% of the genome, as well as other loci corresponding to ∼32% of the genome that exhibit replication activity in two different time windows. Analyses of genomic, transcriptional, and chromatin features of the euchromatic portion of the maize genome provide evidence for a gradient of early replicating, open chromatin that transitions gradually to less open and less transcriptionally active chromatin replicating in mid S phase. Our genomic level analysis also demonstrated that the centromere core replicates in mid S, before heavily compacted classical heterochromatin, including pericentromeres and knobs, which replicate during late S phase.}, number={9}, journal={PLANT CELL}, author={Wear, Emily E. and Song, Jawon and Zynda, Gregory J. and LeBlanc, Chantal and Lee, Tae-Jin and Mickelson-Young, Leigh and Concia, Lorenzo and Mulvaney, Patrick and Szymanski, Eric S. and Allen, George C. and et al.}, year={2017}, month={Sep}, pages={2126–2149} } @article{mickelson-young_wear_mulvaney_lee_szymanski_allen_hanley-bowdoin_thompson_2016, title={A flow cytometric method for estimating S-phase duration in plants}, volume={67}, ISSN={["1460-2431"]}, url={http://europepmc.org/abstract/med/27697785}, DOI={10.1093/jxb/erw367}, abstractNote={Highlight We estimated S-phase duration for several plant species by following EdU-labeled nuclei from G1 to G2 using bivariate flow cytometry. S-phase duration is relatively consistent over a range of genome sizes.}, number={21}, journal={JOURNAL OF EXPERIMENTAL BOTANY}, author={Mickelson-Young, Leigh and Wear, Emily and Mulvaney, Patrick and Lee, Tae-Jin and Szymanski, Eric S. and Allen, George and Hanley-Bowdoin, Linda and Thompson, William}, year={2016}, month={Nov}, pages={6077–6087} } @article{bass_hoffman_lee_wear_joseph_allen_hanley-bowdoin_thompson_2015, title={Defining multiple, distinct, and shared spatiotemporal patterns of DNA replication and endoreduplication from 3D image analysis of developing maize (Zea mays L.) root tip nuclei}, volume={89}, ISSN={["1573-5028"]}, DOI={10.1007/s11103-015-0364-4}, abstractNote={Spatiotemporal patterns of DNA replication have been described for yeast and many types of cultured animal cells, frequently after cell cycle arrest to aid in synchronization. However, patterns of DNA replication in nuclei from plants or naturally developing organs remain largely uncharacterized. Here we report findings from 3D quantitative analysis of DNA replication and endoreduplication in nuclei from pulse-labeled developing maize root tips. In both early and middle S phase nuclei, flow-sorted on the basis of DNA content, replicative labeling was widely distributed across euchromatic regions of the nucleoplasm. We did not observe the perinuclear or perinucleolar replicative labeling patterns characteristic of middle S phase in mammals. Instead, the early versus middle S phase patterns in maize could be distinguished cytologically by correlating two quantitative, continuous variables, replicative labeling and DAPI staining. Early S nuclei exhibited widely distributed euchromatic labeling preferentially localized to regions with weak DAPI signals. Middle S nuclei also exhibited widely distributed euchromatic labeling, but the label was preferentially localized to regions with strong DAPI signals. Highly condensed heterochromatin, including knobs, replicated during late S phase as previously reported. Similar spatiotemporal replication patterns were observed for both mitotic and endocycling maize nuclei. These results revealed that maize euchromatin exists as an intermingled mixture of two components distinguished by their condensation state and replication timing. These different patterns might reflect a previously described genome organization pattern, with "gene islands" mostly replicating during early S phase followed by most of the intergenic repetitive regions replicating during middle S phase.}, number={4-5}, journal={PLANT MOLECULAR BIOLOGY}, author={Bass, Hank W. and Hoffman, Gregg G. and Lee, Tae-Jin and Wear, Emily E. and Joseph, Stacey R. and Allen, George C. and Hanley-Bowdoin, Linda and Thompson, William F.}, year={2015}, month={Nov}, pages={339–351} } @misc{bass_wear_lee_hoffman_gumber_allen_thompson_hanley-bowdoin_2014, title={A maize root tip system to study DNA replication programmes in somatic and endocycling nuclei during plant development}, volume={65}, ISSN={["1460-2431"]}, DOI={10.1093/jxb/ert470}, abstractNote={The progress of nuclear DNA replication is complex in both time and space, and may reflect several levels of chromatin structure and 3-dimensional organization within the nucleus. To understand the relationship between DNA replication and developmental programmes, it is important to examine replication and nuclear substructure in different developmental contexts including natural cell-cycle progressions in situ. Plant meristems offer an ideal opportunity to analyse such processes in the context of normal growth of an organism. Our current understanding of large-scale chromosomal DNA replication has been limited by the lack of appropriate tools to visualize DNA replication with high resolution at defined points within S phase. In this perspective, we discuss a promising new system that can be used to visualize DNA replication in isolated maize (Zea mays L.) root tip nuclei after in planta pulse labelling with the thymidine analogue, 5-ethynyl-2'-deoxyuridine (EdU). Mixed populations of EdU-labelled nuclei are then separated by flow cytometry into sequential stages of S phase and examined directly using 3-dimensional deconvolution microscopy to characterize spatial patterns of plant DNA replication. Combining spatiotemporal analyses with studies of replication and epigenetic inheritance at the molecular level enables an integrated experimental approach to problems of mitotic inheritance and cellular differentiation.}, number={10}, journal={JOURNAL OF EXPERIMENTAL BOTANY}, author={Bass, Hank W. and Wear, Emily E. and Lee, Tae-Jin and Hoffman, Gregg G. and Gumber, Hardeep K. and Allen, George C. and Thompson, William F. and Hanley-Bowdoin, Linda}, year={2014}, month={Jun}, pages={2747–2756} } @article{barampuram_allen_krasnyanski_2014, title={Effect of various sterilization procedures on the in vitro germination of cotton seeds}, volume={118}, ISSN={["1573-5044"]}, DOI={10.1007/s11240-014-0472-x}, number={1}, journal={PLANT CELL TISSUE AND ORGAN CULTURE}, author={Barampuram, Shyam and Allen, George and Krasnyanski, Sergei}, year={2014}, month={Jul}, pages={179–185} } @article{pascuzzi_flores-vergara_lee_sosinski_vaughn_hanley-bowdoin_thompson_allen_2014, title={In Vivo Mapping of Arabidopsis Scaffold/Matrix Attachment Regions Reveals Link to Nucleosome-Disfavoring Poly(dA:dT) Tracts}, volume={26}, ISSN={["1532-298X"]}, DOI={10.1105/tpc.113.121194}, abstractNote={This work uses tiling microarrays to map S/MARs on Arabidopsis chromosome 4. S/MARs were found to be spaced more closely than in the large plant and animal genomes studied to date and preferentially enriched in poly(dA:dT) tracts, sequences that resist nucleosome formation. Most S/MARs occur near gene transcription start sites, and these genes show an increased probability of expression. Scaffold or matrix attachment regions (S/MARs) are found in all eukaryotes. The pattern of distribution and genomic context of S/MARs is thought to be important for processes such as chromatin organization and modulation of gene expression. Despite the importance of such processes, much is unknown about the large-scale distribution and sequence content of S/MARs in vivo. Here, we report the use of tiling microarrays to map 1358 S/MARs on Arabidopsis thaliana chromosome 4 (chr4). S/MARs occur throughout chr4, spaced much more closely than in the large plant and animal genomes that have been studied to date. Arabidopsis S/MARs can be divided into five clusters based on their association with other genomic features, suggesting a diversity of functions. While some Arabidopsis S/MARs may define structural domains, most occur near the transcription start sites of genes. Genes associated with these S/MARs have an increased probability of expression, which is particularly pronounced in the case of transcription factor genes. Analysis of sequence motifs and 6-mer enrichment patterns show that S/MARs are preferentially enriched in poly(dA:dT) tracts, sequences that resist nucleosome formation, and the majority of S/MARs contain at least one nucleosome-depleted region. This global view of S/MARs provides a framework to begin evaluating genome-scale models for S/MAR function.}, number={1}, journal={PLANT CELL}, author={Pascuzzi, Pete E. and Flores-Vergara, Miguel A. and Lee, Tae-Jin and Sosinski, Bryon and Vaughn, Matthew W. and Hanley-Bowdoin, Linda and Thompson, William F. and Allen, George C.}, year={2014}, month={Jan}, pages={102–120} } @article{bajwa_wang_blackburn_goshe_mitra_williams_bishop_krasnyanski_allen_huber_et al._2013, title={Identification and Functional Analysis of Tomato BRI1 and BAK1 Receptor Kinase Phosphorylation Sites}, volume={163}, ISSN={["1532-2548"]}, DOI={10.1104/pp.113.221465}, abstractNote={Abstract Brassinosteroids (BRs) are plant hormones that are perceived at the cell surface by a membrane-bound receptor kinase, BRASSINOSTEROID INSENSITIVE1 (BRI1). BRI1 interacts with BRI1-ASSOCIATED RECEPTOR KINASE1 (BAK1) to initiate a signal transduction pathway in which autophosphorylation and transphosphorylation of BRI1 and BAK1, as well as phosphorylation of multiple downstream substrates, play critical roles. Detailed mechanisms of BR signaling have been examined in Arabidopsis (Arabidopsis thaliana), but the role of BRI1 and BAK1 phosphorylation in crop plants is unknown. As a foundation for understanding the mechanism of BR signaling in tomato (Solanum lycopersicum), we used liquid chromatography-tandem mass spectrometry to identify multiple in vitro phosphorylation sites of the tomato BRI1 and BAK1 cytoplasmic domains. Kinase assays showed that both tomato BRI1 and BAK1 are active in autophosphorylation as well as transphosphorylation of each other and specific peptide substrates with a defined sequence motif. Site-directed mutagenesis revealed that the highly conserved kinase domain activation loop residue threonine-1054 was essential for tomato BRI1 autophosphorylation and peptide substrate phosphorylation in vitro. Furthermore, analysis of transgenic lines expressing full-length tomato BRI1-Flag constructs in the weak tomato bri1 allele, curl3-abs1, demonstrated that threonine-1054 is also essential for normal BRI1 signaling and tomato growth in planta. Finally, we cloned the tomato ortholog of TGF-β Receptor Interacting Protein (TRIP1), which was previously shown to be a BRI1-interacting protein and kinase domain substrate in Arabidopsis, and found that tomato TRIP1 is a substrate of both tomato BRI1 and BAK1 kinases in vitro.}, number={1}, journal={PLANT PHYSIOLOGY}, author={Bajwa, Vikramjit S. and Wang, Xiaofeng and Blackburn, R. Kevin and Goshe, Michael B. and Mitra, Srijeet K. and Williams, Elisabeth L. and Bishop, Gerard J. and Krasnyanski, Sergei and Allen, George and Huber, Steven C. and et al.}, year={2013}, month={Sep}, pages={30–42} } @article{williamson_desai_krasnyanski_ding_guo_nguyen_olson_dole_allen_2013, title={Overexpression of mannitol dehydrogenase in zonal geranium confers increased resistance to the mannitol secreting fungal pathogen Botrytis cinerea}, volume={115}, DOI={10.1007/s11240-013-0368-1}, number={3}, journal={Plant Cell, Tissue and Organ Culture}, author={Williamson, J. D. and Desai, A. and Krasnyanski, S. F. and Ding, F. and Guo, W. W. and Nguyen, T. T. and Olson, H. A. and Dole, J. M. and Allen, G. C.}, year={2013}, pages={367–375} } @article{jia_li_allen_feng_xue_2012, title={A Novel Glyceraldehyde-3-Phosphate Dehydrogenase (GAPDH) Promoter for Expressing Transgenes in the Halotolerant Alga Dunaliella salina}, volume={64}, ISSN={["0343-8651"]}, DOI={10.1007/s00284-012-0102-y}, abstractNote={A major challenge for efficient transgene expression in Dunaliella salina is to find strong endogenous promoters to drive the transgene expression. In the present study, a novel glyceraldehyde-3-phosphate dehydrogenase (GAPDH) promoter was cloned and used to drive expressions of the bialaphos resistance (bar) gene and of the N-terminal fragment of human canstatin (Can-N). The results showed that the bar gene was transcribed by the GAPDH promoter and integrated into the genome of the transformants of D. salina. Furthermore, the PCR identification, Southern and western blots indicated that Can-N was expressed in transgenic D. salina, demonstrating that the promoter of the D. salina GAPDH gene is suitable for driving expression of heterologous genes in transgenic D. salina.}, number={5}, journal={CURRENT MICROBIOLOGY}, author={Jia, Yanlong and Li, Shenke and Allen, George and Feng, Shuying and Xue, Lexun}, year={2012}, month={May}, pages={506–513} } @article{lee_pascuzzi_settlage_shultz_tanurdzic_rabinowicz_menges_zheng_main_murray_et al._2010, title={Arabidopsis thaliana Chromosome 4 Replicates in Two Phases That Correlate with Chromatin State}, volume={6}, ISSN={1553-7404}, url={http://dx.doi.org/10.1371/journal.pgen.1000982}, DOI={10.1371/journal.pgen.1000982}, abstractNote={DNA replication programs have been studied extensively in yeast and animal systems, where they have been shown to correlate with gene expression and certain epigenetic modifications. Despite the conservation of core DNA replication proteins, little is known about replication programs in plants. We used flow cytometry and tiling microarrays to profile DNA replication of Arabidopsis thaliana chromosome 4 (chr4) during early, mid, and late S phase. Replication profiles for early and mid S phase were similar and encompassed the majority of the euchromatin. Late S phase exhibited a distinctly different profile that includes the remaining euchromatin and essentially all of the heterochromatin. Termination zones were consistent between experiments, allowing us to define 163 putative replicons on chr4 that clustered into larger domains of predominately early or late replication. Early-replicating sequences, especially the initiation zones of early replicons, displayed a pattern of epigenetic modifications specifying an open chromatin conformation. Late replicons, and the termination zones of early replicons, showed an opposite pattern. Histone H3 acetylated on lysine 56 (H3K56ac) was enriched in early replicons, as well as the initiation zones of both early and late replicons. H3K56ac was also associated with expressed genes, but this effect was local whereas replication time correlated with H3K56ac over broad regions. The similarity of the replication profiles for early and mid S phase cells indicates that replication origin activation in euchromatin is stochastic. Replicon organization in Arabidopsis is strongly influenced by epigenetic modifications to histones and DNA. The domain organization of Arabidopsis is more similar to that in Drosophila than that in mammals, which may reflect genome size and complexity. The distinct patterns of association of H3K56ac with gene expression and early replication provide evidence that H3K56ac may be associated with initiation zones and replication origins.}, number={6}, journal={PLoS Genetics}, publisher={Public Library of Science (PLoS)}, author={Lee, Tae-Jin and Pascuzzi, Pete E. and Settlage, Sharon B. and Shultz, Randall W. and Tanurdzic, Milos and Rabinowicz, Pablo D. and Menges, Margit and Zheng, Ping and Main, Dorrie and Murray, James A. H. and et al.}, editor={Copenhaver, Gregory P.Editor}, year={2010}, month={Jun}, pages={e1000982} } @article{shultz_lee_allen_thompson_hanley-bowdoin_2009, title={Dynamic Localization of the DNA Replication Proteins MCM5 and MCM7 in Plants}, volume={150}, ISSN={["1532-2548"]}, DOI={10.1104/pp.109.136614}, abstractNote={AbstractGenome integrity in eukaryotes depends on licensing mechanisms that prevent loading of the minichromosome maintenance complex (MCM2-7) onto replicated DNA during S phase. Although the principle of licensing appears to be conserved across all eukaryotes, the mechanisms that control it vary, and it is not clear how licensing is regulated in plants. In this work, we demonstrate that subunits of the MCM2-7 complex are coordinately expressed during Arabidopsis (Arabidopsis thaliana) development and are abundant in proliferating and endocycling tissues, indicative of a role in DNA replication. We show that endogenous MCM5 and MCM7 proteins are localized in the nucleus during G1, S, and G2 phases of the cell cycle and are released into the cytoplasmic compartment during mitosis. We also show that MCM5 and MCM7 are topologically constrained on DNA and that the MCM complex is stable under high-salt conditions. Our results are consistent with a conserved replicative helicase function for the MCM complex in plants but not with the idea that plants resemble budding yeast by actively exporting the MCM complex from the nucleus to prevent unauthorized origin licensing and rereplication during S phase. Instead, our data show that, like other higher eukaryotes, the MCM complex in plants remains in the nucleus throughout most of the cell cycle and is only dispersed in mitotic cells.}, number={2}, journal={PLANT PHYSIOLOGY}, author={Shultz, Randall W. and Lee, Tae-Jin and Allen, George C. and Thompson, William F. and Hanley-Bowdoin, Linda}, year={2009}, month={Jun}, pages={658–669} } @article{santa-maria_pecota_yencho_allen_sosinski_2009, title={Rapid shoot regeneration in industrial 'high starch' sweetpotato (Ipomoea batatas L.) genotypes}, volume={97}, ISSN={["1573-5044"]}, DOI={10.1007/s11240-009-9504-3}, number={1}, journal={PLANT CELL TISSUE AND ORGAN CULTURE}, author={Santa-Maria, Monica and Pecota, Kenneth V. and Yencho, Craig G. and Allen, George and Sosinski, Bryon}, year={2009}, month={Apr}, pages={109–117} } @article{allen_flores-vergara_krasnyanski_kumar_thompson_2006, title={A modified protocol for rapid DNA isolation from plant tissues using cetyltrimethylammonium bromide}, volume={1}, ISSN={["1750-2799"]}, DOI={10.1038/nprot.2006.384}, abstractNote={We describe a modification of the DNA extraction method, in which cetyltrimethylammonium bromide (CTAB) is used to extract nucleic acids from plant tissues. In contrast to the original method, the modified CTAB procedure is faster, omits the selective precipitation and CsCl gradient steps, uses less expensive and toxic reagents, requires only inexpensive laboratory equipment and is more readily adapted to high-throughput DNA extraction. This protocol yields approximately 5-30 microg of total DNA from 200 mg of tissue fresh weight, depending on plant species and tissue source. It can be completed in as little as 5-6 h.}, number={5}, journal={NATURE PROTOCOLS}, author={Allen, G. C. and Flores-Vergara, M. A. and Krasnyanski, S. and Kumar, S. and Thompson, W. F.}, year={2006}, pages={2320–2325} } @misc{kumar_allen_thompson_2006, title={Gene targeting in plants: fingers on the move}, volume={11}, ISSN={["1360-1385"]}, DOI={10.1016/j.tplants.2006.02.002}, abstractNote={Zinc-finger endonucleases (ZFNs) make targeted double-stranded breaks in genomic DNA and, thus, stimulate recombination and repair processes at specific sites. ZFNs can now be harnessed to stimulate homologous recombination and gene targeting in plants, which represents a major step towards modifying the plant genome more precisely. ZFN-mediated gene targeting is likely to become a powerful tool for genome research and genetic engineering. Zinc-finger endonucleases (ZFNs) make targeted double-stranded breaks in genomic DNA and, thus, stimulate recombination and repair processes at specific sites. ZFNs can now be harnessed to stimulate homologous recombination and gene targeting in plants, which represents a major step towards modifying the plant genome more precisely. ZFN-mediated gene targeting is likely to become a powerful tool for genome research and genetic engineering.}, number={4}, journal={TRENDS IN PLANT SCIENCE}, author={Kumar, S and Allen, GC and Thompson, WF}, year={2006}, month={Apr}, pages={159–161} } @misc{helmer_allen_thompson_2006, title={High efficiency gene targeting in plants}, volume={7,126,041}, number={2006 Oct. 24}, publisher={Washington, DC: U.S. Patent and Trademark Office}, author={Helmer, G. L. and Allen, G. C. and Thompson, W. F.}, year={2006} } @article{abranches_shultz_thompson_allen_2005, title={Matrix attachment regions and regulated transcription increase and stabilize transgene expression}, volume={3}, ISSN={["1467-7644"]}, DOI={10.1111/j.1467-7652.2005.00144.x}, abstractNote={SummaryTransgene silencing has been shown to be associated with strong promoters, but it is not known whether the propensity for silencing is caused by the level of transcription, or some other property of the promoter. If transcriptional activity fosters silencing, then transgenes with inducible promoters may be less susceptible to silencing. To test this idea, a doxycycline‐inducible luciferase transgene was transformed into an NT1 tobacco suspension culture cell line that constitutively expressed the tetracycline repressor. The inducible luciferase gene was flanked by tobacco Rb7 matrix attachment regions (MAR) or spacer control sequences in order to test the effects of MARs in conjunction with regulated transcription. Transformed lines were grown under continuous doxycycline (CI), or delayed doxycycline induction (DI) conditions. Delayed induction resulted in higher luciferase expression initially, but continued growth in the presence of doxycycline resulted in a reduction of expression to levels similar to those found in continuously induced lines. In both DI and CI treatments, the Rb7 MAR significantly reduced the percentage of silenced lines and increased transgene expression levels. These data demonstrate that active transcription increases silencing, especially in the absence of the Rb7 MAR. Importantly, the Rb7 MAR lines showed higher expression levels under both CI and DI conditions and avoided silencing that may occur in the absence of active transcription such as what would be expected as a result of condensed chromatin spreading.}, number={5}, journal={PLANT BIOTECHNOLOGY JOURNAL}, author={Abranches, R and Shultz, RW and Thompson, WF and Allen, GC}, year={2005}, month={Sep}, pages={535–543} } @article{ascenzi_ulker_todd_sowinski_schimeneck_allen_weissinger_thompson_2003, title={Analysis of trans-silencing interactions using transcriptional silencers of varying strength and targets with and without flanking nuclear matrix attachment regions}, volume={12}, ISSN={["1573-9368"]}, DOI={10.1023/A:1023310118231}, abstractNote={We investigated the effect of the Rb7 matrix attachment region (MAR) on trans-silencing in tobacco plants, comparing the effects of three transgene silencer loci on ten target loci. Two of the silencer loci, C40 and C190, contain complex and rearranged transgene arrays consisting of 35S:GUS or NOS:NPTII containing plasmids. The third silencer locus, V271, was previously characterized as a complex locus containing rearranged 35S:RiN sequences. Each of these silencers can reduce 35S promoter-driven expression at other loci, albeit with varying efficiencies. The presence of MARs at a target locus does not prevent trans-silencing by the V271 silencer. However, four of seven MAR-containing loci were at least partially resistant to silencing by the C40 and C190 loci. One MAR locus was unaffected by C40, our weakest silencer, and three were silenced only when the silencer locus was maternally inherited. Silencing is progressive in the F1 and F2 generations; two days after germination there is little or no difference between seedlings derived from crosses to silencing or control lines, but seedlings containing silencer loci slowly lose expression during subsequent development. These observations are compatible with the hypothesis that a product of the silencer locus must accumulate before unlinked loci can be affected. However, our silencer loci are themselves silenced for GUS transcription, and coding region homology is not required for their effects on target loci. Our results are consistent with a model in which transcriptional silencing is triggered by transcription of sequences during the early stages of embryo or seedling development.}, number={3}, journal={TRANSGENIC RESEARCH}, author={Ascenzi, R and Ulker, B and Todd, JJ and Sowinski, DA and Schimeneck, CR and Allen, GC and Weissinger, AK and Thompson, WF}, year={2003}, month={Jun}, pages={305–318} } @article{mankin_allen_phelan_spiker_thompson_2003, title={Elevation of transgene expression level by flanking matrix attachment regions (MAR) is promoter dependent: a study of the interactions of six promoters with the RB7 3 ' MAR}, volume={12}, ISSN={["0962-8819"]}, DOI={10.1023/A:1022194120518}, abstractNote={We have analyzed effects of a matrix attachment region (MAR) from the tobacco RB7 gene on transgene expression from six different promoters in stably transformed tobacco cell cultures. The presence of MARs flanking the transgene increased expression of constructs based on the constitutive CaMV 35S, NOS, and OCS promoters. Expression from an induced heat shock promoter was also increased and MARs did not cause expression in the absence of heat shock. There was also no effect of MARs on the pea ferredoxin promoter, which is not normally expressed in this cell line. Importantly, most transgenes flanked by RB7 MAR elements showed a large reduction in the number of low expressing GUS transformants relative to control constructs without MARs.}, number={1}, journal={TRANSGENIC RESEARCH}, author={Mankin, SL and Allen, GC and Phelan, T and Spiker, S and Thompson, WF}, year={2003}, month={Feb}, pages={3–12} } @article{love_allen_gatz_thompson_2002, title={Differential Top10 promoter regulation by six tetracycline analogues in plant cells}, volume={53}, ISSN={["1460-2431"]}, DOI={10.1093/jxb/erf050}, abstractNote={The effects of five tetracycline analogues, anhydrotetracycline, doxycycline, minocycline, oxytetracycline, and tetracycline, on Top10 promoter activity in NT1 tobacco tissue culture cells have been analysed. The concentration that repressed Top10 promoter activity, the level of transgene repression and the kinetics of transgene de-repression were determined for each analogue, and could not be predicted from in vitro binding affinity to the tetracycline repressor or from comparison with animal cells. Doxycycline had the most potent effect on the Top10 promoter and completely inhibited transgene expression at 4 nmol l(-1). Tetracycline was the most versatile of the analogues tested; tetracycline inhibited the Top10 promoter at 10 nmol l(-1) and was easily washed out to restore Top10-driven expression in 12-24 h. A study was also made of the suitability for plant research of a novel tetracycline analogue, GR33076X. In animal cells, GR33076X de-repressed Top10 promoter activity in the presence of inhibitory concentrations of anhydrotetracycline. In NT1, it is shown that GR 33076X can antagonize repression of the Top10 promoter in the presence of tetracycline, but not of anhydrotetracycline or of doxycycline. Different tetracycline analogues can therefore be used to regulate the Top10 promoter in plant cells and this property may be exploited in planning an optimum course of transgene regulation.}, number={376}, journal={JOURNAL OF EXPERIMENTAL BOTANY}, author={Love, J and Allen, GC and Gatz, C and Thompson, WF}, year={2002}, month={Sep}, pages={1871–1877} } @article{callaway_abranches_scroggs_allen_thompson_2002, title={High-throughput transgene copy number estimation by competitive PCR}, volume={20}, ISSN={["0735-9640"]}, DOI={10.1007/BF02782462}, number={3}, journal={PLANT MOLECULAR BIOLOGY REPORTER}, author={Callaway, AS and Abranches, R and Scroggs, J and Allen, GC and Thompson, WF}, year={2002}, month={Sep}, pages={265–277} } @misc{thompson_allen_mankin_2000, title={Increasing expression of transgenes in plant cells using insulator elements}, volume={6,100,448}, number={2000 Aug. 8}, publisher={Washington, DC: U.S. Patent and Trademark Office}, author={Thompson, W. and Allen, G. and Mankin, S.}, year={2000} } @misc{thompson_allen_mankin_2000, title={Method for reducing expression variability of transgenes in plant cells}, volume={6,037,525}, number={2000 Mar. 14}, publisher={Washington, DC: U.S. Patent and Trademark Office}, author={Thompson, W. and Allen, G. and Mankin, S.}, year={2000} } @misc{allen_spiker_thompson_2000, title={Use of matrix attachment regions (MARs) to minimize transgene silencing}, volume={43}, ISSN={["0167-4412"]}, DOI={10.1023/A:1006424621037}, abstractNote={Matrix attachment regions (MARs) are operationally defined as DNA elements that bind specifically to the nuclear matrix in vitro. It is possible, although unproven, that they also mediate binding of chromatin to the nuclear matrix in vivo and alter the topology of the genome in interphase nuclei. When MARs are positioned on either side of a transgene their presence usually results in higher and more stable expression in transgenic plants or cell lines, most likely by minimizing gene silencing. Our review explores current data and presents several plausible models to explain MAR effects on transgene expression.}, number={2-3}, journal={PLANT MOLECULAR BIOLOGY}, author={Allen, GC and Spiker, S and Thompson, WF}, year={2000}, month={Jun}, pages={361–376} } @article{ulker_allen_thompson_spiker_weissinger_1999, title={A tobacco matrix attachment region reduces the loss of transgene expression in the progeny of transgenic tobacco plants}, volume={18}, ISSN={["1365-313X"]}, DOI={10.1046/j.1365-313X.1999.00453.x}, abstractNote={SummaryThe RB7 matrix attachment region (MAR), when flanking a uidA (GUS) reporter gene, has been previously shown to increase uidA gene expression by 60‐fold in stably transformed tobacco suspension cell lines. We have now used the same co‐transformation procedure to determine the effect of flanking MARs on uidA gene expression in tobacco plants. The neomycin phosphotransferase selection gene and uidA reporter gene on separate plasmids were co‐transformed into seedlings by microprojectile bombardment. In primary transgenic plants, the average uidA expression in plants with MARs was twofold greater than in control plants without MARs, but there was no effect on variation of expression. GUS activity was not proportional to the number of integrated uidA transgenes over the entire range of copy numbers. However, in the lower part of the copy number range, MAR lines show a tendency for expression to increase with copy number. Transgene expression in backcross progenies of the MAR‐containing lines averaged threefold higher than in control progenies. MARs also reduced the loss of transgene expression in the BC1 generation. Sixty‐three per cent of the 21 MAR‐containing primary transformants, but only 20% of the 14 control primary transformants, produced backcross progenies in which no loss of transgene expression was observed. These observations are discussed in the context of homology‐dependent gene silencing.}, number={3}, journal={PLANT JOURNAL}, author={Ulker, B and Allen, GC and Thompson, WF and Spiker, S and Weissinger, AK}, year={1999}, month={May}, pages={253–263} } @article{michalowski_allen_hall_thompson_spiker_1999, title={Characterization of randomly-obtained matrix attachment regions (MARs) from higher plants}, volume={38}, ISSN={["0006-2960"]}, DOI={10.1021/bi991142c}, abstractNote={Matrix attachment regions (MARs) can be operationally defined as DNA fragments that bind to the nuclear matrix. We have created a library of randomly obtained MARs from tobacco (Nicotiana tobacum) by cloning DNA fragments that co-isolate with nuclear matrixes prepared by a method involving lithium diiodosalicylate. The interactions of several of the cloned MARs with nuclear matrixes were tested by an in vitro binding assay in which genomic DNA was used as competitor. Based on this assay, the MARs were classified as strong, medium, and weak binders. Examples of each of the binding classes were further studied by in vitro binding using self- and cross-competition. Estimates of dissociation constants for several MARs ranged from 6 to 11 nM and correlated inversely with binding strength. The number of binding sites per matrix for several MARs ranged from 4 x 10(5) to 9 x 10(5) and correlated directly with binding strength. We conclude that binding strength, as we have measured it, is a function of both numbers of binding sites and affinity for the sites. The tobacco MARs were sequenced and analyzed for overall AT content, for distribution of AT-rich regions, and for the abundance of several MAR-related motifs. Previously identified MAR motifs correlate to various degrees with binding strength. Notably, the Drosophila topoisomerase II motif does not correlate with binding strength of the tobacco MARs. A newly identified motif, the "90%AT Box," correlates better with binding strength than any of the previously identified motifs we investigated.}, number={39}, journal={BIOCHEMISTRY}, author={Michalowski, SM and Allen, GC and Hall, GE and Thompson, WF and Spiker, S}, year={1999}, month={Sep}, pages={12795–12804} } @article{vain_worland_kohli_snape_christou_allen_thompson_1999, title={Matrix attachment regions increase transgene expression levels and stability in transgenic rice plants and their progeny}, volume={18}, ISSN={["0960-7412"]}, DOI={10.1046/j.1365-313X.1999.00446.x}, abstractNote={SummaryTo investigate the effect of matrix attachment regions (MARs) on transgene expression levels and stability in cereal crops, we generated 83 independent transgenic rice callus lines containing a gusA expression cassette either as a simple expression unit, or flanked with MARs from tobacco (Rb7) or yeast (ARS1). Transgenic rice plants were regenerated from these callus lines and analysed at the structural and expression levels over two generations. In the first generation (T0), both Rb7 and ARS1 MARs significantly increased transgene expression levels. In the populations of plants containing MARs, we observed a significant reduction in the number of non‐expressing lines compared to the population of plants without MARs. However, variation in β‐glucuronidase (GUS) expression levels between independent lines was similar both in the presence and absence of flanking MARs. In the presence of MARs, GUS activity increased in proportion to transgene copy number up to 20 copies, but was generally reduced in lines carrying a higher copy number. In the population of plants without MARs, there was no correlation between expression level and transgene copy number. In the second generation (T1), transgene expression levels were significantly correlated with those of the T0 parents. The Rb7 MARs significantly improved the stability of transgene expression levels over two generations, and therefore appear to offer protection against transgene silencing. Our study shows that the exploitation of MARs may be an important strategy for stabilising transgene expression levels in genetically engineered cereals.}, number={3}, journal={PLANT JOURNAL}, author={Vain, P and Worland, B and Kohli, A and Snape, JW and Christou, P and Allen, GC and Thompson, WF}, year={1999}, month={May}, pages={233–242} } @article{petracek_dickey_nguyen_gatz_sowinski_allen_thompson_1998, title={Ferredoxin-1 mRNA is destabilized by changes in photosynthetic electron transport}, volume={95}, ISSN={["0027-8424"]}, DOI={10.1073/pnas.95.15.9009}, abstractNote={ In transgenic tobacco, pea Ferredoxin-1 ( Fed-1 ) mRNA accumulates rapidly in response to photosynthesis even when the transgene is driven by a constitutive promoter. To investigate the role of photosynthesis on Fed-1 mRNA stability, we used the tetracycline repressible Top10 promoter system to specifically shut off transcription of the Fed-1 transgene. The Fed-1 mRNA has a half-life of approximately 2.4 hr in the light and a half-life of only 1.2 hr in the dark or in the presence of the photosynthetic electron transport inhibitor 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU). These data indicate that cessation of photosynthesis, either by darkness or DCMU results in a destabilization of the Fed-1 mRNA. Furthermore, the Fed-1 mRNA half-life is reduced immediately upon transfer to darkness, suggesting that Fed-1 mRNA destabilization is a primary response to photosynthesis rather than a secondary response to long-term dark adaptation. Finally, the two different methods for efficient tetracycline delivery reported here generally should be useful for half-life measurements of other mRNAs in whole plants. }, number={15}, journal={PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA}, author={Petracek, ME and Dickey, LF and Nguyen, TT and Gatz, C and Sowinski, DA and Allen, GC and Thompson, WF}, year={1998}, month={Jul}, pages={9009–9013} } @misc{method of increasing expression for foreign genes in plant cells_1998, volume={5,773,689}, number={1998 June 30}, publisher={Washington, DC: U.S. Patent and Trademark Office}, year={1998} } @misc{plant nuclear scaffold attachment region and method for increasing gene expression in transgenic cells_1998, volume={5,773,695}, number={1998 June 30}, publisher={Washington, DC: U.S. Patent and Trademark Office}, year={1998} } @inbook{petracek_dickey_hansen_sowinski_nguyen_allen_thompson_1998, title={The dependence of Fed-1 light regulation on translation}, booktitle={A look beyond transcription: Mechanisms determining mRNAstability and translation in plants}, publisher={Washington, DC: American Society of Plant Physiologists}, author={Petracek, M. E. and Dickey, L. F. and Hansen, E. R. and Sowinski, D. A. and Nguyen, T. and Allen, G. C. and Thompson, G. F.}, editor={J. Bailey-Serres and Gallie, D. R.Editors}, year={1998}, pages={96–101} } @article{mankin_allen_thompson_1997, title={Introduction of a plant intron into the luciferase gene of Photinus pyralis}, volume={15}, DOI={10.1007/bf02812270}, number={2}, journal={Plant Molecular Biology Reporter}, author={Mankin, S. L. and Allen, G. C and Thompson, William}, year={1997}, pages={186–196} } @article{allen_hall_michalowski_newman_spiker_weissinger_thompson_1996, title={High-Level Transgene Expression in Plant Cells: Effects of a Strong Scaffold Attachment Region from Tobacco}, volume={8}, ISSN={1040-4651}, url={http://dx.doi.org/10.2307/3870291}, DOI={10.2307/3870291}, number={5}, journal={The Plant Cell}, publisher={JSTOR}, author={Allen, George C. and Hall, Gerald and Michalowski, Susan and Newman, Winnell and Spiker, Steven and Weissinger, Arthur K. and Thompson, William F.}, year={1996}, month={May}, pages={899} } @article{allen_hall_childs_weissinger_spiker_thompson_1993, title={Scaffold attachment regions increase reporter gene expresssion in stably transformed plant cells}, volume={5}, DOI={10.2307/3869803}, number={6}, journal={Plant Cell}, author={Allen, G.C. and Hall, G.E. and Childs, L.C. and Weissinger, A.K. and Spiker, S.L. and Thompson, W.F.}, year={1993}, pages={603–613} } @article{allen_grimm_elkan_1991, title={Oxygen uptake and hydrogen-stimulated nitrogenase activity from Azorhizobium caulinodans ORS571 grown in a succinate-limited chemostat}, volume={57}, number={11}, journal={Applied and Environmental Microbiology}, author={Allen, G. C. and Grimm, D. T. and Elkan, G. H.}, year={1991}, pages={3220} }