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Prime Editing: Adding Precision and Flexibility to CRISPR Editing

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

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

CRISPR 101: CRISPR-mediated Plant Base Editors

Posted by Guest Blogger on Jan 3, 2019 8:35:29 AM

This post was contributed by Kutubuddin Molla, a Fulbright Visiting Scholar at the Pennsylvania State University.

Imagine you are dealing with a defective gene, Xm, the sequence of which is identical to the correct gene, Xw, except for a single base. If you heard about CRISPR, one question probably comes to mind: can CRISPR be applied to fix the defective base precisely?

Until 2016, precise single base changes were only possible through exploiting the homology-directed repair (HDR) pathway which occurs in cells at low frequencies and relies on the efficient delivery of donor DNA to facilitate repair. Since the development of CRISPR-mediated base editing (BE), these types of repairs can now be done more efficiently than before. A base editor precisely changes a single base with an efficiency typically ranging from 25-75%, while the success of precise change via HDR limited to 0-5%. This blog post covers a brief review of different basic BE technologies and their adaptation for plant genome editing.
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Topics: CRISPR, Plant Biology, CRISPR 101, Base Editing

Single Base Editing with CRISPR

Posted by Mary Gearing on Aug 16, 2016 10:30:00 AM

This post was updated on Nov 27, 2017.

When we talk about CRISPR applications, one negative always 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, Addgene depositor David Liu’s lab created new Cas9 fusion proteins that act as base editorsThese fusions contain dCas9 or Cas9 nickase and a cytidine deaminase, and they can convert cytosine to uracil without cutting DNA. Uracil is then subsequently converted to thymine through DNA replication or repair. Later work has improved base editor targeting flexibility and specificity. New adenine base editors also allow you to change adenine to inosine, which is then converted to guanine.

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Topics: CRISPR, Base Editing

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