Baculoviral delivery of CRISPR/Cas9 facilitates efficient genome editing in human cells

PLoS One. 2017 Jun 22;12(6):e0179514. doi: 10.1371/journal.pone.0179514. eCollection 2017.

Abstract

The CRISPR/Cas9 system is a highly effective tool for genome editing. Key to robust genome editing is the efficient delivery of the CRISPR/Cas9 machinery. Viral delivery systems are efficient vehicles for the transduction of foreign genes but commonly used viral vectors suffer from a limited capacity in the genetic information they can carry. Baculovirus however is capable of carrying large exogenous DNA fragments. Here we investigate the use of baculoviral vectors as a delivery vehicle for CRISPR/Cas9 based genome-editing tools. We demonstrate transduction of a panel of cell lines with Cas9 and an sgRNA sequence, which results in efficient knockout of all four targeted subunits of the chromosomal passenger complex (CPC). We further show that introduction of a homology directed repair template into the same CRISPR/Cas9 baculovirus facilitates introduction of specific point mutations and endogenous gene tags. Tagging of the CPC recruitment factor Haspin with the fluorescent reporter YFP allowed us to study its native localization as well as recruitment to the cohesin subunit Pds5B.

MeSH terms

  • Baculoviridae / genetics*
  • CRISPR-Cas Systems / genetics*
  • Cell Line, Tumor
  • Gene Editing / methods*
  • Gene Knockout Techniques
  • Genome, Human / genetics
  • Humans
  • Intracellular Signaling Peptides and Proteins / genetics
  • Point Mutation
  • Protein Serine-Threonine Kinases / genetics

Substances

  • Intracellular Signaling Peptides and Proteins
  • HASPIN protein, human
  • Protein Serine-Threonine Kinases

Grants and funding

This work was financially supported by The Dutch Cancer Society, KWF Kankerbestrijding (UU2011-5134 to S.M.A.L.) and The Netherlands Organization for Scientific Research (NWO-Vici 918.12.610 to S.M.A.L. and NWO-Veni 722.013.003 to M.A.H.).