Genome editing mediated by SpCas9 variants with broad non-canonical PAM compatibility in plants

Dec 31, 2020Molecular plant

Genome editing in plants using SpCas9 variants that recognize a wider range of DNA targets

AI simplified

CRISPR Gene Editing on OpenScience ↗PubMed ↗DOI ↗

Abstract

Five engineered SpCas9 variants can target non-canonical PAM sites in rice.

SpCas9 variants were modified to enable recognition of non-canonical PAMs, expanding their editing capabilities.
Targeted mutagenesis was assessed at various PAM sites in rice through stable transformation.
Base editors derived from these variants successfully converted C∙G to T∙A at non-canonical PAM sites.
The toolbox created could enhance gene disruption and precise editing in plants.

AI simplified

Streptococcus pyogenes Cas9 (SpCas9) is the most widely used genome editing tool in plants. The editing induced by SpCas9 strictly requires a canonical NGG protospacer-adjacent motif (PAM), significantly limiting its scope of application. Recently, five SpCas9 variants, SpCas9-NRRH, SpCas9-NRCH, SpCas9-NRTH, SpG, and SpRY, were developed to recognize non-canonical PAMs in human cells. In this study, these variants were engineered for plant genome editing, and their targeted mutagenesis capabilities were comprehensively examined at various canonical and non-canonical PAM sites in rice (Oryza sativa) by stable transformation. Moreover, both cytosine base editors using a rat APOBEC1 or a human APOBEC3a and adenine base editors using a directly evolved highly compatible TadA∗-8e deaminase were developed from these SpCas9 variants. Our results demonstrated that the developed SpCas9 variants-based base editors readily generated conversions between C∙G and T∙A in the target sites with non-canonical PAMs in transgenic rice lines. Collectively, the toolbox developed in this study substantially expands the scope of SpCas9-mediated genome editing and will greatly facilitate gene disruption and precise editing in plants.

Full Text

Full text is available at the source.

View full text (XML) ↗View full text ↗

Genome editing mediated by SpCas9 variants with broad non-canonical PAM compatibility in plants

Abstract

Full Text

You found one interesting study. We’ll send the next 7.

what lands in your inbox each week:

Recent issues from the crispr gene editing brief

Abstract

Full Text

Related papers

You found one interesting study. We’ll send the next 7.

what lands in your inbox each week:

Recent issues from the crispr gene editing brief