Green fluorescent protein (GFP) as a reporter gene for the plant pathogenic oomycete Phytophthora palmivora

Abstract

Transgenic Phytophthora palmivora strains that produce green fluorescent protein (GFP) or beta-glucuronidase (GUS) constitutively were obtained after stable DNA integration using a polyethylene-glycol and CaCl2-based transformation protocol. GFP and GUS production were monitored during several stages of the life cycle of P. palmivora to evaluate their use in molecular and physiological studies. 40% of the GFP transformants produced the GFP to a level detectable by a confocal laser scanning microscope, whereas 75% of the GUS transformants produced GUS. GFP could be visualised readily in swimming zoospores and other developmental stages of P. palmivora cells. For high magnification microscopic studies, GFP is better visualised and was superior to GUS. In contrast, for macroscopic examination, GUS was superior. Our findings indicate that both GFP and GUS can be used successfully as reporter genes in P. palmivora.

The CBEL glycoprotein ofPhytophthora parasiticavar-nicotianaeis involved in cell wall deposition and adhesion to cellulosic substrates

“…Both strategies only allowed recovery of strains in which CBEL expression was suppressed. This is in accordance with data obtained in P. infestans, where silencing resulted from both sense and antisense constructs (van West et al, 1999a).…”

“…The feasibility to suppress gene expression was first illustrated in P. infestans where a transgenic strain expressing the β-glucuronidase (gus) reporter gene was further transformed with gus antisense constructs resulting in suppression of gus expression (Judelson et al, 1993b). So far, this technology has been applied only once to inactivate an endogenous gene: the inf1 gene of P. infestans, which encodes a peptide elicitor from the elicitin protein family (Kamoun et al, 1998;van West et al, 1999a). However, the presence of multiple inf genes in the pathogen did not allow complete suppression of elicitin production.…”

“…These two expressing (CBEL + ) and the two non-expressing (CBEL -) transformants were selected for further analyses. By analogy to suppression of gene expression in sense and antisense transformants of P. infestans (van West et al, 1999a), it can be supposed that the CBEL -strains resulted from homology-dependent gene silencing. It is worth noting that the transformants were highly impaired in zoospore production independently of CBEL gene expression.…”

“…strains with altered levels of CBEL production. So far, only a few Phytophthora (P.) species, comprising P. infestans, P. sojae, P. capsici and P. palmivora, have been transformed (Judelson et al, 1991;Judelson et al, 1993a;Ersek et al, 1994;van West et al, 1999b). We formerly established that P.p.n.…”

Transformation of Antisense Chalcone Synthase (CHS) Gene into Lotus (Nelumbo Nucifera Gaertn.) by Particle Bombardment

“…Rachinee were similar (72 and 75 pieces, respectively; Table 1). The GFP gene was also reported as a reporter gene in transgenic Phytothora palmivora and provided better visualization and was superior to GUS [17]. GFP can partially replace antibiotic selection and is particularly important when organogenesis or conversion of transformation procedures is inefficient under antibiotic or herbicide selection.…”