CRISPR 101: Any Base Transversion Editors

By Emily P. Bentley

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The nucleobases are shown with arrows describing conversions between them. In step one, a cytosine deaminase converts cytosine to uracil; this is catalyzed directly by the base editor. Next, base excision of uracil (also by the base editor) is repaired in two different ways by the cell, shown by an arrow that splits into two outcomes. Repair in E. coli leads to adenine, while repair in mammalian cells leads to guanine.
A cartoon depiction of cytidine base editing. A base editor, consisting of a cytidine deaminase fused to Cas9, is shown binding to DNA using its guide RNA. The guide RNA base pairs to target DNA, leaving the opposite strand of DNA free to be contacted by the cytidine deaminase, which converts a C to a U within this single-stranded sequence. This deamination yields DNA with a G:U mismatch without creating a double-strand break. Mismatch repair preserves the edit IF the modified strand is used as the template, converting the mismatched G to an A and yielding a single-base-pair edit.
Schematic showing that base editing converts the blue fluorescent protein expressing plasmid into one that expresses green fluorescent protein.
Workflow of phage-assisted evolution of base-editing activity. Fresh host cells contain Cas, mutated gIII, and mutagenesis plasmids. Phage infection delivers the deaminase plasmid. After mutagenesis, if the base editor is inactive, no pIII is produced, and the progeny phage are not infectious. If the base editor is active, however, pIII is produced and the progeny phage are infectious, allowing them to continue propagating.
Plant base editor platforms and their molecular components.

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