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Hypercompact adenine base editors based on transposase B guided by engineered RNA.

Do Yon Kim1, Yuhee Chung1, Yujin Lee2,3

  • 1GenKOre, Daejeon, Republic of Korea.

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|August 1, 2022
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Summary

Researchers developed TaRGET (TnpB-augment RNA-based Genome Editing Technology), a new system for precise genome editing using Transposon-associated transposase B (TnpB). This technology enables efficient adenine base editing in mammalian cells, advancing CRISPR-based gene therapy.

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Area of Science:

  • Molecular Biology
  • Genetics
  • Biotechnology

Background:

  • Transposon-associated transposase B (TnpB) is an ancestral protein related to Cas12 enzymes.
  • Previous work engineered guide RNAs for Cas12f1 with high indel efficiency in human cells.

Purpose of the Study:

  • To develop a novel genome editing technology utilizing TnpB.
  • To establish TnpB-based adenine base editors (ABEs) for precise genome modification.

Main Methods:

  • Engineered guide RNAs were designed to confer endonuclease activity to TnpB.
  • A specific TnpB mutant (V106W, D108Q dimer fused to dTnpB D354A) was created for adenine base editing.
  • Engineered TnpB variants and optimized deaminases were used to expand targetable sites.

Main Results:

  • The new technology, TaRGET (TnpB-augment RNA-based Genome Editing Technology), demonstrated high-efficiency programmable endonuclease activity for TnpB.
  • The developed TnpB-based adenine base editors (ABEs) achieved significant A-to-G conversion rates.
  • Delivery of TaRGET-ABE resulted in potent A-to-G conversion in mammalian genomes.

Conclusions:

  • TaRGET-ABE represents a significant advancement in precise genome editing tools.
  • The technology is amenable to delivery via adeno-associated viruses (AAVs).
  • This innovation holds promise for the development of clustered regularly interspaced short palindromic repeats (CRISPR)-based gene therapies.