Genetically encoded DNA recorders noninvasively convert transient biological events into durable mutations in a cell’s genome, allowing for the later reconstruction of cellular experiences by DNA sequencing. We present a DNA recorder, peCHYRON, that achieves high-information, durable, and temporally resolved multiplexed recording of multiple cellular signals in mammalian cells. In each step of recording, prime editor, a Cas9-reverse transcriptase fusion protein, inserts a variable triplet DNA sequence alongside a constant propagator sequence that deactivates the previous and activates the next step of insertion. Insertions accumulate sequentially in a unidirectional order, editing can continue indefinitely, and high information is achieved by coexpressing a variety of prime editing guide RNAs (pegRNAs), each harboring unique triplet DNA sequences. We demonstrate that the constitutive expression of pegRNA collections generates insertion patterns for the straightforward reconstruction of cell lineage relationships and that the inducible expression of specific pegRNAs results in the accurate recording of exposures to biological stimuli.

基因编码的 DNA 记录器将瞬时生物事件无创地转化为细胞基因组中的持久突变，从而允许稍后通过 DNA 测序重建细胞体验。我们提出了一种 DNA 记录仪 peCHYRON，它可以在哺乳动物细胞中实现多细胞信号的高信息、持久和时间分辨的多重记录。在记录的每个步骤中，Prime Editor（一种 Cas9 逆转录酶融合蛋白）将可变三联体 DNA 序列插入一个恒定的繁殖序列，该序列使前一个序列失活并激活下一步的插入。插入以单向顺序连续积累，编辑可以无限期地持续，并且通过共表达各种引物编辑向导 RNA （pegRNA） 获得高信息，每个 RNA 都含有独特的三联体 DNA 序列。我们证明 pegRNA 集合的组成型表达产生了用于直接重建细胞谱系关系的插入模式，并且特异性 pegRNA 的诱导表达导致准确记录暴露于生物刺激。