PrimeDesign software for rapid and simplified design of prime editing guide RNAs

Jonathan Y Hsu; Julian Grünewald; Regan Szalay; Justine Shih; Andrew V Anzalone; Kin Chung Lam; Max W Shen; Karl Petri; David R Liu; J Keith Joung; Luca Pinello

doi:10.1038/s41467-021-21337-7

PrimeDesign software for rapid and simplified design of prime editing guide RNAs

Nat Commun. 2021 Feb 15;12(1):1034. doi: 10.1038/s41467-021-21337-7.

Authors

Jonathan Y Hsu^{1

2

3}, Julian Grünewald^{2

3

4}, Regan Szalay^{2

3}, Justine Shih^{2

3}, Andrew V Anzalone^{5

6

7}, Kin Chung Lam^{2

3

4}, Max W Shen^{5

6

7

8}, Karl Petri^{2

3

4}, David R Liu^{5

6

7}, J Keith Joung^{9

10

11}, Luca Pinello^{12

13

14}

Affiliations

¹ Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA.
² Molecular Pathology Unit, Massachusetts General Hospital, Charlestown, MA, USA.
³ Center for Cancer Research and Center for Computational and Integrative Biology, Massachusetts General Hospital, Charlestown, MA, USA.
⁴ Department of Pathology, Harvard Medical School, Boston, MA, USA.
⁵ Merkin Institute of Transformative Technologies in Healthcare, Broad Institute of Harvard and MIT, Cambridge, MA, USA.
⁶ Department of Chemistry and Chemical Biology, Harvard University, Cambridge, MA, USA.
⁷ Howard Hughes Medical Institute, Harvard University, Cambridge, MA, USA.
⁸ Computational and Systems Biology Program, Massachusetts Institute of Technology, Cambridge, MA, USA.
⁹ Molecular Pathology Unit, Massachusetts General Hospital, Charlestown, MA, USA. [email protected].
¹⁰ Center for Cancer Research and Center for Computational and Integrative Biology, Massachusetts General Hospital, Charlestown, MA, USA. [email protected].
¹¹ Department of Pathology, Harvard Medical School, Boston, MA, USA. [email protected].
¹² Molecular Pathology Unit, Massachusetts General Hospital, Charlestown, MA, USA. [email protected].
¹³ Department of Pathology, Harvard Medical School, Boston, MA, USA. [email protected].
¹⁴ Broad Institute of Harvard and MIT, Cambridge, MA, USA. [email protected].

Abstract

Prime editing (PE) is a versatile genome editing technology, but design of the required guide RNAs is more complex than for standard CRISPR-based nucleases or base editors. Here we describe PrimeDesign, a user-friendly, end-to-end web application and command-line tool for the design of PE experiments. PrimeDesign can be used for single and combination editing applications, as well as genome-wide and saturation mutagenesis screens. Using PrimeDesign, we construct PrimeVar, a comprehensive and searchable database that includes candidate prime editing guide RNA (pegRNA) and nicking sgRNA (ngRNA) combinations for installing or correcting >68,500 pathogenic human genetic variants from the ClinVar database. Finally, we use PrimeDesign to design pegRNAs/ngRNAs to install a variety of human pathogenic variants in human cells.

Publication types

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

MeSH terms

Base Pairing
Base Sequence
CRISPR-Associated Protein 9 / genetics
CRISPR-Associated Protein 9 / metabolism
CRISPR-Cas Systems*
Clustered Regularly Interspaced Short Palindromic Repeats
Databases, Genetic
Fabry Disease / genetics
Fabry Disease / metabolism
Fabry Disease / pathology
Gene Editing / methods*
Genome, Human*
Green Fluorescent Proteins / genetics
Green Fluorescent Proteins / metabolism
HEK293 Cells
Hemophilia A / genetics
Hemophilia A / metabolism
Hemophilia A / pathology
Humans
Models, Biological
Muscular Dystrophy, Duchenne / genetics
Muscular Dystrophy, Duchenne / metabolism
Muscular Dystrophy, Duchenne / pathology
Mutation
Nucleic Acid Conformation
Plasmids / chemistry
Plasmids / metabolism
RNA, Guide, CRISPR-Cas Systems / genetics*
RNA, Guide, CRISPR-Cas Systems / metabolism
Recombinant Fusion Proteins / genetics
Recombinant Fusion Proteins / metabolism

Substances

RNA, Guide, CRISPR-Cas Systems
Recombinant Fusion Proteins
enhanced green fluorescent protein
Green Fluorescent Proteins
CRISPR-Associated Protein 9

Abstract

Publication types

MeSH terms

Substances

Grants and funding