Differential vulnerability CRISPRi screen in Rif Sensitive and Rif Resistant strains of M. tuberculosis

A genome-wide CRISPRi library (RLC0011) was transformed into an M. tuberculosis bioA deletion strain (Rif Sensitive) and an M. tuberculosis rpoB S450L bioA deletion strain (Rif Resistant). The libraries were serially passaged in triplicate in either the presence (CRISPRi on) or absence (CRISPRi off) of ATc. Genomic DNA was harvested at the following generations (cumulative) and subjected to deep sequencing: 0, 2.5, 5, 7.5, 10, 15, 20, 25, and 30. Genetic vulnerability was then calculated, similar to what was described in Bosch & DeJesus et al. (PMID: 34297925). Differential vulnerability of each gene in the S450L mutant strain was calculated with respect to the parental, wild-type M. tuberculosis strain.