Prediction of prime editing insertion efficiencies using sequence features and DNA repair determinants
knockout HAP1 cell lines were scaled to be identical for >13-nt sequences that are not affected by MMR. The fold changes of the scaled insertion efficiencies between HAP1Validation of nuclease overexpression with individual pegRNAsHEK3
site-targeting pegRNAs endowed with 25- or 34-nt homology arms. At one d before transfection, HEK293T cells were seeded in two 24-well plates at 50,000 cells per well. All transfections were done in replicates and each well was transfected with 500 ng of pCMV_PE2_P2A_PuroR, 150 ng of pTwist nuclease or eGFP overexpression constructs, and 100 ng of pegRNA using Lipofectamine LTX according to the manufacturer’s protocol.
The final model was trained with XGBoost using the features length; normalized secondary structure of the reverse transcriptase template; MMR proficiency; percentage of the nucleotides C, A and T; the number of paired bases between the first 3 nt of the insert and the last 3 nt of the spacer in addition to the first nucleotide of the scaffold; complementarity between the first nucleotide of the insert and the nucleotide at the nicking site; the maximum number of consecutive adenines in the...
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