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. 2022 Jul 6;608(7921):98–107. doi: 10.1038/s41586-022-04922-8

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© The Author(s) 2022

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 2 — a, Schematic of five transfection programmes over 8 or 16 epochs. For programmes 1 and 2, pegRNAs with single barcodes were introduced in each epoch for 16 epochs.The specific orders aimed to maximize (programme 1) or minimize (programme 2) the edit distances between temporally adjacent transfections. For programme 3, pegRNAs with two different barcodes were introduced at a 1:1 ratio for 16 epochs, with one barcode always shared between adjacent epochs (and between epochs 1 and 16). For programmes 4 and 5, pegRNAs with two different barcodes were introduced either at a constant ratio (1:3) or at varying ratios in each epoch (1:1, 1:2, 1:4 or 1:8) for eight epochs, respectively. b, Barcode frequencies across five insertion sites in 5×TAPE-1 in programmes 1 and 2 following epoch 16. Barcodes introduced in early epochs are more frequently observed at the first site. c–g, Bigram transition matrices for programmes 1 (c), 2 (d), 3 (e), 4 (f) and 5 (g). Barcodes are ordered from early (left/top) to late (right/bottom). h, Calculated versus intended relative frequencies between programmes 4 and 5. Programme ratios were calculated by combining sequencing reads from n = 3 independent transfection experiments.