To improve the error rates of the genes assembled from OLS pool 2,
we used ErrASE, a commercially available enzyme cocktail that
detects and corrects mismatched base pairs, to remove errors in the
assembled fluorescent proteins. For each gene, we applied ErrASE
at six different stringencies, reamplified the constructs, cloned the
PCR products and rescreened the cloned genes using flow cytometry.
Improvement of the level of fluorescence progressively increased
with greater ErrASE stringency. At the highest levels of error cor-
rection, the fluorescence levels were 31%, 49% and 26% for mTFP1,
mCitrine and mApple respectively (Fig. 3a and Supplementary
Fig. 8). We also performed the ErrASE procedure on our GFP43
and GFP35 pools from OLS pool 1, resulting in fluorescence levels
of 89% and 92%, respectively (Fig. 3a and Supplementary Fig. 5c).
We sequenced clones of GFP43 and GFP35 and found three errors in
21,510 (1/7,170 bp) and four errors in 20,076 (1/5,019 bp) sequenced
bases, respectively.