FX174 Attenuation by Whole Genome Codon Deoptimization

James T. Van Leuven; Martina M. Ederer; Katelyn Burleigh; LuAnn Scott; Randall A. Hughes; Vlad Codrea; Andrew D. Ellington; Holly Wichman; Craig Miller

We codon deoptimized the genes of ΦX174 by recoding nearly all synonymous residues to the least preferred E. coli codons. We did not recode regions of genes that overlapped with other genes nor the first six residues of each gene since these residues are known to have strong effects on gene expression (87) . Four of the six recoded variants were less fit than wild type ΦX174 ( fig. 3a) . The construct containing the fully deoptimized G gene could not be recovered, even after growing the strain overnight on the susceptible host cells in an attempt to 8 . CC-BY-NC 4.0 International license author/funder. It is made available under a The copyright holder for this preprint (which was not peer-reviewed) is the . https://doi.org/10.1101/2020.02.10.942847 doi: bioRxiv preprint obtain a recovery mutation. Recoding highly expressed genes (J, F, and G) resulted in larger fitness decreases than recoding lowly expressed genes (A and H). Although the number of variants built was small, the fitness effects of deoptimization were correlated to the proportion of codons edited and the CAI of the recoded genes ( fig. 3b and To further investigate how deleterious effects combine, we employed a statistical framework for calculating the best-fitting model of epistasis.