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CRISPR 101: Cytosine and Adenine Base Editors

Posted by Mary Gearing on Aug 6, 2020 10:30:00 AM

Originally published Aug 16, 2016 and last updated Aug 6, 2020 by Jennifer Tsang.

When we talk about CRISPR applications, one negative often comes up: the low editing efficiency of homology-directed repair (HDR). Compared to non-homologous end joining, HDR occurs at a relatively low frequency, and in nondividing cells, this pathway is further downregulated. Rather than try to improve HDR, scientists have developed two classes of base editors: cytosine base editors (CBEs) and adenine base editors (ABEs). 

(There are also RNA base editors, but we’ll just be covering DNA base editors here. To learn more about RNA base editors head over to this blog post: CRISPR 101: RNA Editing with Cas13)

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Topics: CRISPR, Base Editing

Four Base Editing Reporters to Monitor and Enrich Editing in Real-time

Posted by Alyssa Cecchetelli on Jul 7, 2020 9:15:00 AM

Base editors create specific point mutations in the genome, but they’re inefficient compared to CRISPR/Cas9 edits that rely on double strand DNA breaks. Due to this inefficiency it is crucial for scientists to not only easily identify base editing events in real-time but also enrich for base-edited cells in their experiments. In the past few years, scientists have created an array of base editing reporters that can help you do just that.

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Topics: CRISPR, Base Editing

A New Generation of Adenine Base Editors Improves Editing in Primary Human Cells

Posted by Susanna Bachle on May 7, 2020 9:15:00 AM

Adenine base editors (ABE) mediate A•T-to-G•C base changes (Figure 1), but it can be challenging to make these base changes, especially in primary human cells. Now, scientists at Beam Therapeutics have found a way to improve editing in primary human cells (Gaudelli et al., 2020).

One of the widely used base editing systems, ABE7.10 (and the starting point for a new generation of ABEs), consists of 3 components: 

  • a deaminase (TadA, originally from E.coli, named TadA7.10 in ABE7.10) 
  • a catalytically impaired Cas protein (dCas or Cas nickase) 
  • a guide RNA that targets the complex of TadA and dCas to the genomic DNA of interest 
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Topics: CRISPR, Base Editing

Expanding the Targeting Scope and Editing Efficiency of Adenine Base Editors

Posted by Susanna Bachle on Mar 17, 2020 9:32:55 AM

David Liu’s lab created the first base editor in 2016 (Komor et al., 2016) and since then has been trying to expand their precision editing capabilities. Base editors make specific DNA base changes and consist of a catalytically impaired Cas protein (dCas or Cas nickase) fused to a DNA-modifying enzyme, in this case a deaminase. Base changes from C•G-to-T•A are mediated by cytosine base editors (CBEs) and base changes from A•T-to-G•C are mediated by adenine base editors (ABEs). How does this work? Through molecular biology teamwork. The guide RNA (gRNA) specifies the editing target site on the DNA, the Cas domain directs the modifying enzyme to the target site, and the deaminase induces the DNA base change without a DNA double-strand break. But base editors aren’t perfect. They may be slow, can only target certain sites, or make only a subset of base substitutions. 

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Topics: CRISPR, Base Editing

Prime Editing: Adding Precision and Flexibility to CRISPR Editing

Posted by Jennifer Tsang on Oct 24, 2019 9:26:53 AM

Updated June 5, 2020.

There are over 75,000 pathogenic genetic variants that have been identified in humans and catalogued in the ClinVar database. Previously developed genome editing methods using nucleases and base editors have the potential to correct only a minority of those variants in most cell types. A new technique from David Liu’s lab at the Broad Institute could add more precision and flexibility to the CRISPR editing world.

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Topics: CRISPR, Cas Proteins, CRISPR gRNAs, Base Editing

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