Expression of Recombinant Aequorin as an Intracellular Calcium Reporter in the Phytopathogenic Fungus Phyllosticta ampelicida Academic Article uri icon

abstract

  • Conidia of Phyllosticta ampelicida germinate only after they have made contact with a substratum. Previous work has shown that external free calcium must be available to the spore for germination to be initiated. Transgenic strains of P. ampelicida expressing apo-aequorin, a calcium-sensitive luminescent protein, were developed to monitor cytoplasmic free Ca(2+) ([Ca(2+)]c). Transformants were verified by PCR and Southern hybridization. Apo-aequorin production was quantified for each of 21 transformants. The transformant that emitted the most light per unit of protein was found to contain 0.59 mg apo-aequorin/g total protein. To ascertain the feasibility of aequorin-based [Ca(2+)]c quantification, [Ca(2+)]c changes were measured in mycelia during various physiologically perturbing treatments: exposure to high concentrations of external Ca(2+), hypoosmotic shock, and mechanical perturbation. This is the first report of a plant pathogenic fungus for which aequorin-based Ca(2+) measurement protocols have been developed.

altmetric score

  • 3

author list (cited authors)

  • Shaw, B. D., Kozlova, O., Read, N. D., Turgeon, B. G., & Hoch, H. C.

citation count

  • 10

publication date

  • December 2001