Reconstruction of evolving gene variants and fitness from short sequencing reads.
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| Abstract | Directed evolution can generate proteins with tailor-made activities. However, full-length genotypes, their frequencies and fitnesses are difficult to measure for evolving gene-length biomolecules using most high-throughput DNA sequencing methods, as short read lengths can lose mutation linkages in haplotypes. Here we present Evoracle, a machine learning method that accurately reconstructs full-length genotypes (R = 0.94) and fitness using short-read data from directed evolution experiments, with substantial improvements over related methods. We validate Evoracle on phage-assisted continuous evolution (PACE) and phage-assisted non-continuous evolution (PANCE) of adenine base editors and OrthoRep evolution of drug-resistant enzymes. Evoracle retains strong performance (R = 0.86) on data with complete linkage loss between neighboring nucleotides and large measurement noise, such as pooled Sanger sequencing data (~US$10 per timepoint), and broadens the accessibility of training machine learning models on gene variant fitnesses. Evoracle can also identify high-fitness variants, including low-frequency 'rising stars', well before they are identifiable from consensus mutations. |
| Year of Publication | 2021
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| Journal | Nature chemical biology
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| Volume | 17
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| Issue | 11
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| Pages | 1188-1198
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| Date Published | 11/2021
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| ISSN | 1552-4469
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| DOI | 10.1038/s41589-021-00876-6
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| PubMed ID | 34635842
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