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Transient expression of gfp in the obligate biotrophic oomycete Plasmopara halstedii using electroporation and a mechanoperforation method
Author(s) -
Hammer T. R.,
Thines M.,
Spring O.
Publication year - 2007
Publication title -
plant pathology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.928
H-Index - 85
eISSN - 1365-3059
pISSN - 0032-0862
DOI - 10.1111/j.1365-3059.2006.01494.x
Subject(s) - oomycete , biology , downy mildew , obligate , transformation (genetics) , obligate parasite , hypha , botany , sporangium , electroporation , phytophthora capsici , green fluorescent protein , sunflower , microbiology and biotechnology , genetics , pathogen , spore , host (biology) , gene , phytophthora , horticulture
Studying plant biotrophic oomycetes is difficult, because they depend on living plant cells for nutrition, which necessitates investigations in planta . Internal optical labelling is a powerful tool for analysing intra‐ and interspecific interactions. Different approaches are described for establishing a gfp ‐transformation system for Plasmopara halstedii , the causal agent of sunflower downy mildew. A vector containing gfp and the oomycete‐specific ham34 regulators from Bremia lactucae was constructed for transformation experiments. Particle bombardment of infected sunflower cotyledons during different developmental stages of the pathogen did not result in successful transformation. In contrast, transient gfp expression in sporangia was achieved using electroporation. However, gfp expression was lost during the subsequent round of infection. A novel transformation method for biotrophic organisms in planta employing mechanoperforation – so‐called Löchern–resulted in sporangiophores of P. halstedii transiently expressing gfp and emerging distant from the site of transformation. The new technique is advantageous compared with others as the transformed hyphae can recover during their vegetative growth, before reproductive structures occur. This first step lays an important foundation for further investigations of obligate biotrophic organisms.