Schematic of TN-seq in C. neoformans. Top left: Transposon insertions (orange arrow) into nonessential genes results in viable cells. In contrast, insertions into essential genes will result in dead and nonrecoverable cells. Top right: TN-seq works by generating a library of cells where each cell has a single independent transposon insertion in a random location. As in A, those insertions into essential regions cause the cells to die and are nonrecoverable. As a result, the total library (bottom) is depleted in insertions in essential regions. Bottom: The Ac/Ds transposon was split into an Ac transposase and a Ds transposon containing a neomycin resistance marker. This Ds transposon was integrated into an intron of URA5 and the Ac transposase was integrated into the safe haven locus. The resulting stain is ura− and neomycin resistant. Upon initiating transposition via growth on galactose, the strain becomes URA+ and mutant at another locus (depicted here as YFG1).
#Fungal infections are hard to treat due to #DrugResistance. @blakebillmyre.bsky.social &co use a high-throughput #TNseq system in #Cryptococcus neoformans to identify >1400 essential genes & reveal a role for #mitochondrial genes in #fluconazole sensitivity @plosbiology.org 🧪 plos.io/4dz3iVm