Efficient, continuous mutagenesis in human cells using a pseudo-random DNA editor

Nat Biotechnol. 2020 Feb;38(2):165-168. doi: 10.1038/s41587-019-0331-8. Epub 2019 Dec 16.

Abstract

Here we describe TRACE (T7 polymerase-driven continuous editing), a method that enables continuous, targeted mutagenesis in human cells using a cytidine deaminase fused to T7 RNA polymerase. TRACE induces high rates of mutagenesis over multiple cell generations in genes under the control of a T7 promoter integrated in the genome. We used TRACE in a MEK1 inhibitor-resistance screen, and identified functionally correlated mutations.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • DNA / genetics*
  • DNA-Directed RNA Polymerases / metabolism
  • Gene Editing / methods*
  • Genetic Loci
  • HEK293 Cells
  • Humans
  • Mutagenesis / genetics*
  • Mutation / genetics
  • Promoter Regions, Genetic
  • Viral Proteins / metabolism

Substances

  • Viral Proteins
  • DNA
  • bacteriophage T7 RNA polymerase
  • DNA-Directed RNA Polymerases