E. coli barcoded deletion library screen for chemotherapeutic antimetabolite gemcitabine (Repeat)

A genetic screen was performed using barcoded E. coli genome-wide knockout library and sub-inhibitory concentration of antimetabolite drug gemcitabine. This knockout library has ~3,680 mutants, each having a unique DNA barcode, GFP tag and chloramphenicol resistance gene in the knockout locus. In addition there are two versions of the library (Version 1-Oddd and Version 2-Even) in which a corresponding mutant has different DNA barcodes in each version. The library was generated from the parent strain BW38028. Pooled library was grown in the presence and absence of gemcitabine in M9 miminal media as four biological replicates, and the experiment was stopped when the cultures reached OD600 value of 0.6. Genomic DNA was extracted and the barcodes were amplified and sequenced by Illumina Next Generation Sequencing.