Evaluation of nat1 and nptII as selectable markers in Lentinula edodes

A large variety of selectable marker genes is desirable to apply diverse experimental techniques in the shiitake mushroom ( Lentinula edodes ). In this study, we evaluated the performance of the genes for nourseothricin resistance ( nat1 ) and G418 (geneticin) resistance ( nptII ) as selectable marker genes in L. edodes. Plasmids containing either nat1 or nptII , each driven by the actin promoter derived from the basidiomycetous yeast Cryptococcus neoformans , were introduced into the L. edodes strain M2. Transformation with nat1 and nptII yielded 3.7 and 14.7 transformants per experiment, respectively. Split read-based insertion site analysis suggested that one to three copies of the introduced sequences were integrated into the chromosomes of a single strain, and the insertion sites differed among strains. Overall, this study presents two new selectable marker genes that can facilitate genetic engineering techniques for shiitake mushrooms.