Figure 3.

Permuted gene abundances are independent of orientation. (A) The relative abundances of every possible cognate P (purple) and AP (green) variant mapped adjacent to one another on a circle as a function of the distance from the start codon, which is shown as a closed black symbol. Within the unselected library, the relative abundance of identical genes in P and AP orientations is similar. (B) For each unique P (purple) and AP (green) sequence, we evaluated the number of degenerate in frame sequences observed for each variant and plotted these as stacked bars. In the unselected library, 159 of the P variants and 148 of the AP variants were observed in one or more reads of the deep mutational scanning data. (C) Following selection, the relative abundances of every possible gene in the P and AP orientation differed as well as (D) the number of degenerate sequences. Among the selected sequence reads, 144 of the P variants and 85 of the AP variants were observed. Among both the naïve and selected libraries, a total of 171 unique P variants were observed out of 223.