DRiF-Seq analysis of gene function in bloodstream-form Trypanosoma brucei lines

Direct-RNAi fragment sequencing (DRiF-Seq) provides quantitative information on the fitness cost of gene knockout through time. To characterise the consequence of gene knockdown in bloodstream-form Trypanosoma brucei, libraries of >240,000 RNAi mutants were generated in Lister 427 (2 libraries) and EATRO1125 (1 library) strain cells and the abundance of each mutant after induction of RNAi was followed through time by sequencing. Lister 427 libraries were sampled at days 0, 1, 2 and 3 post-induction. EATRO1125 libraries were sampled at days 0, 1, 2, 3, 4 and 6. RNAi fragments were amplified from total genomic DNA by linear quantitative PCR and sequenced from both ends on an Illumina platform.