Transformation of the entomopathogenic fungus Paecilomyces fumosoroseus with Agrobacterium tumefaciens

Abstract

Aims: To test the suitability of the Agrobacterium tumefaciens‐mediated transformation (AMT) method with Paecilomyces fumosoroseus, a fungal pathogen that causes diseases in a wide range of insects including whiteflies. Methods and Results: Conidia of P. fumosoroseus were successfully transformed to hygromycin B resistance using the hph gene of Escherichia coli as the selectable marker. Transformation frequencies were 58·3 ± 18·5, 98·3 ± 24·8 and 169·7 ± 35·5 (±SEM) transformants per 105, 106 and 107 target conidia respectively. After confirmation by PCR, transformants were subjected to Southern analysis, and the results revealed that 45% (four of nine) of the transformants contained single‐copy integration of the T‐DNA. Conclusions: In our AMT system, we efficiently transformed conidia of P. fumosoroseus. The employment of this method circumvents time‐consuming protoplast preparation and allows the isolation of transformants containing single‐copy integration of the T‐DNA. Significance and Impact of the Study: Considering the efficiency of Ag. tumefaciens‐mediated transformation, this method represents a useful tool for insertional mutagenesis to characterize genes that are important for the pathogenicity of P. fumosoroseus.

Genetic transformation of Fusarium avenaceum by Agrobacterium tumefaciens mediated transformation and the development of a USER-Brick vector construction system

“…The two medium concentrations tested yielded the same number of transformants, while an increase from 5*10 5 to 1*10 6 macroconidia per plate lead to a significant reduction (Figure 6). A similar reduction has been reported for Paecilomyces fumosoroseus [52]. The biological explanation for the reduction is possibly self-inhibition of germination at higher macroconidia concentrations, as described for other fungal species such as Colletotrichum fragariae [53].…”

Tissue specific response of Agrobacterium tumefaciens attachment to Sorghum bicolor (L) Moench

“…However, recent studies have shown that besides infecting its natural host Agrobacterium also transfers DNA to monocotyledons plant species including important cereals like maize (Ishida et al, 1996;Negrotto et al, 2000;Frame et al, 2002), rice (Aldemita and Hodges, 1996;Mohanty et al, 1999;Arockiasamy and Ignacimuthu, 2007) sugarcane (Arencibia et al, 1998;Enriquez-obregon et al, 1998) and sorghum (Zhao et al, 2000). Agrobacterium has also been successfully used for genetic transformation of organisms such as yeast (Piers et al, 1996), fungi (Lee, 2006;lima et al, 2006;Stenlid, 2006) and human cells (Kunik et al, 2001). Although the transformation efficiency in these species is comparatively less when compared to dicotyledonous plant species.…”

WSC1 Regulates the Growth, Development, Patulin Production, and Pathogenicity of Penicillium expansum Infecting Pear Fruits

“…The first report on the genetic transformation of filamentous fungi was from Neurospora scabra. 20 There are currently various methods used to study the gene function of filamentous fungi, such as gene knockout, 21 overexpression, 22,23 transposon labeling, 24,25 and yeast hybridization. 26 Due to its simple tumefaciens as mediators.…”

“…The first report on the genetic transformation of filamentous fungi was from Neurospora scabra . There are currently various methods used to study the gene function of filamentous fungi, such as gene knockout, overexpression, , transposon labeling, , and yeast hybridization . Due to its simple operation and ability to specifically inhibit the expression of target genes, gene knockout is widely used, and most gene knockout protocols use Agrobacterium-mediated homologous recombination methods.…”