List of base-pair substitutions found in Mutation Accumulation lines of Escherichia coli K-12 PFM2 and its derivative strains (DNA repair)

List of base-pair substitutions found in mutation accumulation of the following E. coli strains: 1. K12 PFM2 (LB) 2. K12 PFM2 (minimal) 3. ED1a 4. IAI1 5. PFM101 6. PFM133 7. PFM35 8. PFM40 9. PFM88 10. PFM180 11. PFM22 12. PFM91 13. PFM61 14. PFM6 15. PFM94Multidisciplinary University Research Initiative Award W911NF-09-1-0444 from the US Army Research OfficePlease refer to Foster et al. PNAS (2015)Microsoft Excel for Mac 2011 Version 14.3.8 (130913

Determinants of Base-Pair Substitution Patterns Revealed by Whole-Genome Sequencing of DNA Mismatch Repair Defective Escherichia coli

Mismatch repair (MMR) is a major contributor to replication fidelity, but its impact varies with sequence context and the nature of the mismatch. Mutation accumulation experiments followed by whole-genome sequencing of MMR-defective Escherichia coli strains yielded ≈30,000 base-pair substitutions (BPSs), revealing mutational patterns across the entire chromosome. The BPS spectrum was dominated by A:T to G:C transitions, which occurred predominantly at the center base of 5'NAC3'+5'GTN3' triplets. Surprisingly, growth on minimal medium or at low temperature attenuated these mutations. Mononucleotide runs were also hotspots for BPSs, and the rate at which these occurred increased with run length. Comparison with ≈2000 BPSs accumulated in MMR-proficient strains revealed that both kinds of hotspots appeared in the wild-type spectrum and so are likely to be sites of frequent replication errors. In MMR-defective strains transitions were strand biased, occurring twice as often when A and C rather than T and G were on the lagging-strand template. Loss of nucleotide diphosphate kinase increases the cellular concentration of dCTP, which resulted in increased rates of mutations due to misinsertion of C opposite A and T. In an mmr ndk double mutant strain, these mutations were more frequent when the template A and T were on the leading strand, suggesting that lagging-strand synthesis was more error-prone, or less well corrected by proofreading, than was leading strand synthesis