Institut for Biomedicin

Lars Bolund

Chromatin accessibility and guide sequence secondary structure affect CRISPR-Cas9 gene editing efficiency

Publikation: Bidrag til tidsskrift/Konferencebidrag i tidsskrift /Bidrag til avisTidsskriftartikelForskningpeer review

DOI

  • Kristopher Torp Jensen, Cambridge University
  • ,
  • Lasse Fløe
  • ,
  • Trine Skov Petersen
  • Jinrong Huang, BGI-Shenzhen, 518083 Shenzhen, China; China National GeneBank-Shenzhen, BGI-Shenzhen, 518083 Shenzhen, China.
  • ,
  • Fengping Xu, Københavns Universitet
  • ,
  • Lars Bolund
  • Yonglun Luo
  • Lin Lin

CRISPR-Cas9 systems have emerged as the method of choice for genome editing, but large variations in on-target efficiencies continue to limit their applicability. Here, we investigate the effect of chromatin accessibility on Cas9-mediated gene editing efficiency for 20 gRNAs targeting 10 genomic loci in HEK293T cells using both SpCas9 and the eSpCas9(1.1) variant. Our study indicates that gene editing is more efficient in euchromatin than in heterochromatin, and we validate this finding in HeLa cells and in human fibroblasts. Furthermore, we investigate the gRNA sequence determinants of CRISPR-Cas9 activity using a surrogate reporter system and find that the efficiency of Cas9-mediated gene editing is dependent on guide sequence secondary structure formation. This knowledge can aid in the further improvement of tools for gRNA design. This article is protected by copyright. All rights reserved.

OriginalsprogEngelsk
TidsskriftFEBS Letters
Vol/bind591
Nummer13
Sider (fra-til)1892-1901
Antal sider10
ISSN0014-5793
DOI
StatusUdgivet - 5 jun. 2017

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