ИСТИНА

Yeast serves as a convenient model organism for genetic screening. Yeast KnockOut Collection (YKOC) comprises about 5,000 knockout strains, each of which contains a unique nucleotide barcode in the genome [1, 2]. Treatment of YKOC strain pool with chemicals enables identification of the strains sensitive and resistant to this specific chemical [3]. In order to do this, total DNA is isolated from the YKOC pool before and after the treatment. Barcode regions are amplified and sequenced using next generation sequencing techniques. However, there is no standard analysis pipeline of such barcode sequencing (BarSeq) experiment results. In this work we performed a BarSeq experiment aimed to identify genes, whose deletions either prevent or enhance the clonal expansion of mutant mitochondrial DNA variants in heteroplasmic yeast cells. We introduce a pipeline for analyzing BarSeq results which identifies barcodes, clarifies ambiguous reads, offers statistics on the BarSeq experiment, and produces ranked lists of genes. We validated our approach by contrasting the fitness of yeast strains in non-fermentable versus fermentable carbon sources in a control experiment. Our findings showed a correlation between single gene knockout strains fitnesses in our BarSeq experiment and previously published genetic screenings. The pipeline made it possible to identify candidate genes whose deletion prevents the clonal expansion of selfish mtDNA in heteroplasmic yeast cells. The list includes MOT3, TRX2 and SIR3 genes as well as genes involved in vesicular transport although the finding requires further validation in independently generated knockouts. The study is supported by RSCF grant # 22-14-00108