Abstract
Stable transformation ofArabidopsis thaliana is a lengthy process that involves up to 3 mo of plant growth and seed selection. We have developed a rapid, 3-wk transient assay system to test the functionality ofcis-regulatory regions controlling expression of a reporter gene in plants before undertaking stable transformation. Two-week-oldArabidopsis seedlings were vacuum-infiltrated withAgrobacterium tumefaciens cultures carrying various upstream regulatory regions controllinguidA (β-glucuronidase [GUS]) expression. Seedlings were fixed and stained for GUS activity 3–5 d following infiltration. Regulatory regions tested in this system include the cauliflower mosaic virus (CaMV)35S promoter, the upstream regulatory region of ribosomal protein geneL23A-1, and a temperature-inducible regulatory region (HSP101B) also fromArabidopsis. The percentage of seedlings positive for GUS activity varied depending on the construct used, with the CaMV35S promoter producing the highest number of GUS-positive seedlings. Temperature induction treatments elicited increased GUS expression in seedlings transformed with theHSP101B regulatory region. Regardless of construct, GUS expression levels were higher in seedlings collected 5 d followingAgrobacterium infiltration than those collected 3–4 d postinfiltration.
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Abbreviations
- CaMV:
-
cauliflower mosaic virus
- GUS:
-
β-glucuronidase
- HSE:
-
heat shock element
- RP:
-
ribosomal protein
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McIntosh, K.B., Hulm, J.L., Young, L.W. et al. A rapidAgrobacterium-mediatedArabidopsis thaliana transient assay system. Plant Mol Biol Rep 22, 53–61 (2004). https://doi.org/10.1007/BF02773349
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DOI: https://doi.org/10.1007/BF02773349