Fig. 4: Evidence of a g/P linkage in batch TXTL (cell-free transcription-translation) reactions.
From: PHEIGES: all-cell-free phage synthesis and selection from engineered genomes
a One pot assembly of T7 phages from five parts leading to two possible phage genomes carrying a tail fiber mutation or not. Batch TXTL expression of this equimolar co-assembly leads to six types of phages, phages encapsulating a mutant tail fiber genome or not and displaying combinations of mutant and wild-type tail fibers (mut: mutant genotype, MUT: mutant phenotype, wt: wild type genotype, WT: wild type phenotype, MIX: mixture of MUT and WT) b Titer of the co-expression on rfaC versus E. coli B determines the mut/MUT + mut/MIX proportion of phages in the co-expression. c Presence of purified ReLPS leads to selective inhibition of phages with mutant tail fibers. E. coli B titer of the phage/LPS mixture determines the wt/WT + mut/WT initial proportion of phage in the co-expression. Subsequent amplification on E. coli B transforms mut/WT phages in mut/MUT phages. Titer on rfaC and E. coli B determines the mut/WT over wt/WT proportion of phages in the ejection mixtures. d Table compiling the proportion of phage types measured in the co-expression and the predicted proportion for a random tail fiber mix with the hypothesis that the six trimer tail fibers of a phage results from the random combinations of three monomers. e Co-expression of mutant and WT tail-fiber phages with various decreasing proportion of mutant tail fiber. The random tail mix corresponds to the H0 hypothesis while other hypotheses regarding the random assembly of tail fibers are also considered (Supplementary Fig. 16). n = 3 biologically independent experiments. Data are presented as mean values +/− SD. Source data are provided as a Source Data file.
