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ARS Home » Northeast Area » Beltsville, Maryland (BARC) » Beltsville Agricultural Research Center » Food Quality Laboratory » Research » Publications at this Location » Publication #342648

Title: Dominant selectable markers for Penicillium spp. transformation and gene function studies

item Jurick, Wayne
item PENG, HUI - Guangxi Normal University
item Gaskins, Verneta
item VICO, IVANA - University Of Belgrade
item Yu, Jiujiang
item Macarisin, Otilia
item Janisiewicz, Wojciech
item PETER, KARI - Pennsylvania State University

Submitted to: Fungal Genomics and Biology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 8/28/2017
Publication Date: 9/4/2017
Citation: Jurick II, W.M., Peng, H., Gaskins, V.L., Vico, I., Yu, J., Macarisin, O., Janisiewicz, W.J., Peter, K.A. 2017. Dominant selectable markers for Penicillium spp. transformation and gene function studies. Fungal Genomics and Biology. 7:153.

Interpretive Summary: Blue mold, caused by the fungus Penicillium expansum and other Penicillium spp., is an important disease of stored apple and pear fruit. The fungus not only reduces the quantity of fresh fruit available for consumption, but also negatively impacts fruit quality, and produces mycotoxins that contaminate apple and pear fruit juices and sauces that are toxic to humans. To better help control this important disease, the fungus has been studied and transformed by scientists to determine the genes that control fruit decay using a single marker for selection of Penicillium spp. transformantion. However, the current system is limited, and the number of markers for transformation need to be increased to help understand the genes that enable the blue mold fungus to cause decay. Therefore, eight different compounds were tested and it was determined that three could serve as additional markers for Penicillium spp. All three compounds killed the blue mold fungus, at different concentrations, which make them optimal candidates for scientists to use to examine Penicillium spp. gene function involved in fruit decay and in the production of toxins.

Technical Abstract: Penicillium spp. has been genetically manipulated and gene function studies have utilized single gene deletion strains for phenotypic analysis. Fungal transformation experiments have relied on hygromycin and hygromycin phosphotransferase (hph) as the main dominant selectable marker (DSM) system in Penicillium spp. This poses a limitation on the number of loci that can be analyzed and complemented in reverse genetic studies. Additionally, other economically important Penicillium spp. have not been evaluated to determine the utility of additional compounds that can serve as dominant selectable markers. Therefore, six compounds with corresponding cloned and characterized resistance genes, were examined for 15 blue mold strains, and their Minimum Inhibitory Concentrations (MICs) determined. Phleomycin, neomycin and G418 were not effective, as Penicillium spp. growth was observed on amended media containing 1000µg/ml of each compound. The efficacy of bialophos to inhibit fungal growth was intermediate, with MICs ranging from 250 to 1000µg/ml, and was species-dependent. However, chlorimuron ethyl and benlate had the lowest MIC values and minimal variation in efficacy within and between species. Therefore, benlate and chlorimuron ethyl are good candidates for use as selectable markers since corresponding fungal resistance genes have been cloned, characterized, and are available from a variety of public and academic sources.