Theoretical and Applied Genetics

, Volume 105, Issue 4, pp 505-514

First online:

A constitutive gene expression system derived from the tCUP cryptic promoter elements

  •  K. MalikAffiliated withEastern Cereal and Oilseed Research Centre, Agriculture and Agri-Food Canada, Room 2091, KW Neatby Bldg, CEF, Ottawa, Ontario K1A 0C6, Canada
  • ,  K. WuAffiliated withEastern Cereal and Oilseed Research Centre, Agriculture and Agri-Food Canada, Room 2091, KW Neatby Bldg, CEF, Ottawa, Ontario K1A 0C6, Canada
  • ,  X.-Q. LiAffiliated withPotato Research Centre, Agriculture and Agri-Food Canada, P.O. Box 20280, Fredericton, New Brunswick E3B 4Z7 Canada
  • ,  T. Martin-HellerAffiliated withEastern Cereal and Oilseed Research Centre, Agriculture and Agri-Food Canada, Room 2091, KW Neatby Bldg, CEF, Ottawa, Ontario K1A 0C6, Canada
  • ,  M. HuAffiliated withEastern Cereal and Oilseed Research Centre, Agriculture and Agri-Food Canada, Room 2091, KW Neatby Bldg, CEF, Ottawa, Ontario K1A 0C6, Canada
  • ,  E. FosterAffiliated withEastern Cereal and Oilseed Research Centre, Agriculture and Agri-Food Canada, Room 2091, KW Neatby Bldg, CEF, Ottawa, Ontario K1A 0C6, Canada
  • ,  L. TianAffiliated withSouthern Crop Protection and Food Research Centre, Agriculture and Agri-Food Canada, London, Ontario N5V 4T3
  • ,  C. WangAffiliated withPotato Research Centre, Agriculture and Agri-Food Canada, P.O. Box 20280, Fredericton, New Brunswick E3B 4Z7 Canada
  • ,  K. WardAffiliated withCereal Research Centre, Agriculture and Agri-Food Canada, Winnipeg, Manitoba R3T 2M9 Canada
    • ,  M. JordanAffiliated withCereal Research Centre, Agriculture and Agri-Food Canada, Winnipeg, Manitoba R3T 2M9 Canada
    • ,  D. BrownAffiliated withSouthern Crop Protection and Food Research Centre, Agriculture and Agri-Food Canada, London, Ontario N5V 4T3
    • ,  S. GleddieAffiliated withEastern Cereal and Oilseed Research Centre, Agriculture and Agri-Food Canada, Room 2091, KW Neatby Bldg, CEF, Ottawa, Ontario K1A 0C6, Canada
    • ,  D. SimmondsAffiliated withEastern Cereal and Oilseed Research Centre, Agriculture and Agri-Food Canada, Room 2091, KW Neatby Bldg, CEF, Ottawa, Ontario K1A 0C6, Canada
    • ,  S. ZhengAffiliated withEastern Cereal and Oilseed Research Centre, Agriculture and Agri-Food Canada, Room 2091, KW Neatby Bldg, CEF, Ottawa, Ontario K1A 0C6, Canada
    • ,  J. SimmondsAffiliated withEastern Cereal and Oilseed Research Centre, Agriculture and Agri-Food Canada, Room 2091, KW Neatby Bldg, CEF, Ottawa, Ontario K1A 0C6, Canada
    • ,  B. MikiAffiliated withEastern Cereal and Oilseed Research Centre, Agriculture and Agri-Food Canada, Room 2091, KW Neatby Bldg, CEF, Ottawa, Ontario K1A 0C6, Canada

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Abstract.

A limited number of constitutive promoters have been used to direct transgene expression in plants and they are often derived from non-plant sources. Here, we describe novel gene-regulatory elements which are associated with a cryptic constitutive promoter from tobacco, tCUP, and modifications that were made to create a strong gene-expression system that is effective across all living cell types from a wide range of plant species, including several important crops (Arabidopsis, canola, flax, alfalfa, tobacco). The tCUP 5′ untranslated region was mutated to eliminate translational interference by upstream ATGs, and the influence of the Kozak consensus sequence on the levels of a β-glucuronidase (GUS) reporter gene activity was demonstrated. These modifications resulted in expression that was greatly enhanced in all organs. A TATA consensus sequence was added to the core promoter to complement an existing Initiator (Inr) sequence. Although this addition was known to elevate core promoter activity by 3-fold the additive effect on the overall gene-expression system was marginal in all of the transgenic plants tested. Two transcriptional enhancers were identified and the region containing them were oligomerized, yielding a significant increase in marker gene-expression in some but not all plant species. In general, the enhanced tCUP gene-expression system generated levels of GUS activity which exceeded that of the 35S promoter in most plant species and the elevation in activity occurred uniformly among the various plant organs. The potential benefit of cryptic elements for the construction of gene-expression systems for crop species is discussed

Constitutive promoter Cryptic elements Enhancer elements Transgenic crop plants