Skip to main content Accessibility help

BAC Library Development and Clone Characterization for Dormancy-Responsive DREB4A, DAM, and FT from Leafy Spurge (Euphorbia esula) Identifies Differential Splicing and Conserved Promoter Motifs

  • David P. Horvath (a1), David Kudrna (a2), Jayson Talag (a2), James V. Anderson (a1), Wun S. Chao (a1), Rod Wing (a2), Michael E. Foley (a1) and Münevver Doğramacı (a1)...


We developed two leafy spurge bacterial artificial chromosome (BAC) libraries that together represent approximately 5× coverage of the leafy spurge genome. The BAC libraries have an average insert size of approximately 143 kb, and copies of the library and filters for hybridization-based screening are publicly available through the Arizona Genomics Institute. These libraries were used to clone full-length genomic copies of an AP2/ERF transcription factor of the A4 subfamily of DEHYDRATION-RESPONSIVE ELEMENT-BINDING PROTEINS (DREB) known to be differentially expressed in crown buds of leafy spurge during endodormancy, a DORMANCY ASSOCIATED MADS-BOX (DAM) gene, and several FLOWERING LOCUS T (FT) genes. Sequencing of these BAC clones revealed the presence of multiple FT genes in leafy spurge. Sequencing also provided evidence that two different DAM transcripts expressed in crown buds of leafy spurge during endo- and eco-dormancy result from alternate splicing of a single DAM gene. Sequence data from the FT promoters was used to identify several conserved elements previously recognized in Arabidopsis, as well as potential novel transcription factor binding sites that may regulate FT. These leafy spurge BAC libraries represent a new genomics-based tool that complements existing genomics resources for the study of plant growth and development in this model perennial weed. Furthermore, phylogenetic footprinting using genes identified with this resource demonstrate the usefulness of studying weedy species to further our general knowledge of agriculturally important genes.


Corresponding author

Corresponding author's E-mail:


Hide All
Alabadi, D., Oyama, T., Yanovsky, M. J., Harmon, F. G., Mas, P., and Kay, S. A. 2001. Reciprocal regulation between TOC1 and LHY/CCA1 within the Arabidopsis circadian clock. Science 293: 880883.
Anderson, J. V. 2008. Emerging technologies: an opportunity for weed biology research. Weed Sci. 56: 281282.
Anderson, J. V., Horvath, D. P., Chao, W. S., Foley, M. E., Hernandez, A. G., Thimmapuram, J., Liu, L., Gong, G. L., Band, M., Kim, R., and Mikel, M. A. 2007. Characterization of an EST database for the perennial weed leafy spurge: an important resource for weed biology research. Weed Sci. 55: 193203.
Bailey, T. L. and Elkan, C. 1994. Fitting a mixture model by expectation maximization to discover motifs in biopolymers. Pp 2836 in Proceedings of the Second International Conference on Intelligent Systems for Molecular Biology. Menlo Park, CA: AAAI Press.
Blanchette, M., Schwikowski, B., and Tompa, M. 2002. Discovery of regulatory elements by a computational method for phylogenetic footprinting. J. Comp. Biol. 9: 211223.
Bohlenius, H., Huang, T., Charbonnel-Campaa, L., Brunner, A. M., Jansson, S., Strauss, S. H., and Nilsson, O. 2006. CO/FT regulatory module controls timing of flowering and seasonal growth cessation in trees. Science 312: 10401043.
Chao, W. S., Foley, M. E., Doğramacı, M., Anderson, J. V., and Horvath, D. P. 2011. Alternating temperature breaks dormancy in leafy spurge seeds and impacts signaling networks associated with HY5. Funct. Integr. Genomics 11: 637649.
Chao, W. S., Horvath, D. P., Anderson, J. V., and Foley, M. E. 2005. Potential model weeds to study genomics, ecology, and physiology in the 21st century. Weed Sci. 53: 929937.
Doğramacı, M., Horvath, D. P., Chao, W. S., Foley, M. E., Christoffers, M. J., and Anderson, J. V. 2010. Extended low temperature impacts dormancy status, flowering competence, and transcript profiles in crown buds of leafy spurge. Plant Mol. Biol. 73: 207226.
Doğramacı, M., Horvath, D. P., Christoffers, M. J., and Anderson, J. V. 2011. Dehydration and vernalization treatments identify overlapping molecular networks impacting endodormancy maintenance in leafy spurge crown buds. Funct. Integr. Genomics 11: 611626.
Dong, M. A., Farré, E. M., and Thomashow, M. F. 2011. CIRCADIAN CLOCK-ASSOCIATED 1 and LATE ELONGATED HYPOCOTYL regulate expression of the C-REPEAT BINDING FACTOR (CBF) pathway in Arabidopsis. Proc. Nat. Acad. Sci. U. S. A. 108: 72417246.
Foley, M. E., Anderson, J. V., Chao, W. S., Doğramacı, M., and Horvath, D. P. 2010. Initial changes in the transcriptome of Euphorbia esula seeds induced to germinate with a combination of constant and diurnal alternating temperatures. Plant Mol. Biol. 73: 131142.
Henson, J., Tischler, G., and Ning, Z. 2012. Next-generation sequencing and large genome assemblies. Pharmacogenomics 13: 901915.
Horvath, D. P. 2009a. Genomics for weed science. Curr. Genomics 11: 4751.
Horvath, D. P. 2009b. Common mechanisms regulate flowering and dormancy. Plant Sci. 177: 523531.
Horvath, D. P., Chao, W. S., Suttle, J. C., Thimmapuram, J., and Anderson, J. V. 2008. Transcriptome analysis identifies novel responses and potential regulatory genes involved in seasonal dormancy transitions of leafy spurge (Euphorbia esula L.). BMC Genomics. 9: 536.
Horvath, D. P., Sung, S., Kim, D., Chao, W. S., and Anderson, J. V. 2010. Cloning, characterization, and regulation of DORMANCY-ASSOCIATED MADS-BOX genes from leafy spurge. Plant Mol. Biol. 73: 169179.
Hsu, C.-Y., Adams, J. P., Klim, H., No, K., Ma, C., Strauss, S. H., Drnevich, J., Vandervelde, L., Ellis, J. D., Rice, M. D., Wickett, N., Gunter, L. E., Tuskan, G. A., Brunner, A. M., Page, G. P., Barakat, A., Carlson, J. E., dePamphilis, C. W., Luthe, D. S., and Yuceer, C. 2011. FLOWERING LOCUS T duplication coordinates reproductive and vegetative growth in perennial poplar. Proc. Nat. Acad. Sci. U. S. A. 108: 1075610761.
Jeffares, D. C., Mourier, T., and Penny, D. 2006. The biology of intron gain and loss. Trends Genet. 22: 1622.
Kahn, M. S. 2011. The role of DREB transcription factors in abiotic stress tolerance of plants. Agric. Environ. Biotech. 25: 24332442.
Koren, S., Schatz, M. C., Walenz, B. P., Martin, J., Howard, J. T., Ganapathy, G., Wang, Z., Rasko, D. A., McCombie, W. R., Jarvis, E. D., and Phillippy, A. M. 2012. Hybrid error correction and de novo assembly of single-molecule sequencing reads. Nat. Biotech. 30: 693700.
Lescot, M., Déhais, P., Thijs, G., Marchal, K., Moreau, Y., Van de Peer, Y., Rouzé, P., and Rombauts, S. 2002. PlantCARE, a database of plant cis-acting regulatory elements and a portal to tools for in silico analysis of promoter sequences. Nucleic Acids Res. 30: 325327.
Loman, N. J., Misra, R. V., Dallman, T. J., Constantinidou, C., Gharbia, S. E., Wain, J., and Pallen, M. 2012. Performance comparison of benchtop high-throughput sequencing platforms. Nat. Biotech. 30: 434439.
Luo, M. and Wing, R. 2003. An improved method for plant BAC library construction. Methods Mol. Biol. 236: 320.
Nakano, T., Suzuki, K., Fujimura, T., and Shinshi, H. 2006. Genome-wide analysis of the ERF gene family in Arabidopsis and rice. Plant Physiol. 140: 411432.
Pin, P. A. and Nilsson, O. 2012. The multifaceted roles of FLOWERING LOCUS T in plant development. Plant Cell Environ. 35: 17421755.
Rinne, P. L. H., Welling, A., Vahala, J., Ripel, L., Ruonala, R., Kangasjarvi, J., and van der Schoot, C. 2011. Chilling of dormant buds hyperinduces FLOWERING LOCUS T and recruits GA-inducible 1,3-beta-glucanases to reopen signal conduits and release dormancy in Populus . Plant Cell 23: 130146.
Salome, P. A., To, J. P. C., Kieber, J. J., and McClung, C. R. 2006. Arabidopsis response regulators ARR3 and ARR4 play cytokinin-independent roles in the control of circadian period. Plant Cell 18: 5569.
Schaller, G. E., Mathews, D. E., Gribskov, M., and Walker, J. C. 2002. Two-component signaling elements and histidyl-aspartyl phosphorelays. in Somerville, C. R., and Meyerowitz, E. M., eds. The Arabidopsis Book. Rockville, MD: American Society of Plant Biologists. doi/10.1199/tab.0044. Accessed January 3, 2013.
Schulz-Schaeffer, J. and Gerhardt, S. 1989. Cytotaxonomic analysis of the Euphorbia spp. (leafy spurge) complex. II. Comparative study of the chromosome morphology. Biol. Zentralbl. 108: 6476.
Stahevitch, A. E., Crompton, C. W., and Wojtas, W. A. 1988. Cytogenetic studies of leafy spurge, Euphorbia esula, and its allies (Euphorbiaceae). Can. J. Bot. 66: 22472257.
Stewart, C. N. Jr., Tranel, P. J., Horvath, D. P., Anderson, J. V., Rieseberg, L. H., Westwood, J. H., Mallory-Smith, C. A., Zapiola, M. L., and Dlugosch, K. M. 2009. Evolution of weediness and invasiveness: charting the course for weed genomics. Weed Sci. 57: 451462.
Tiwari, S. B., Shen, Y., Chang, H.-C., Hou, Y., Harris, A., Ma, S. F., McPartland, M., Hymus, G. J., Adam, L., Marion, C., Belachew, A., Repetti, P. P., Reuber, T. L., and Ratcliffe, O. J. 2010. The flowering time regulator CONSTANS is recruited to the FLOWERING LOCUS T promoter via a unique cis-element. New Phytol. 187: 5766.



Full text views

Total number of HTML views: 0
Total number of PDF views: 0 *
Loading metrics...

Abstract views

Total abstract views: 0 *
Loading metrics...

* Views captured on Cambridge Core between <date>. This data will be updated every 24 hours.

Usage data cannot currently be displayed