TY - JOUR
T1 - Engineering CRISPR-Cpf1 crRNAs and mRNAs to maximize genome editing efficiency
AU - Li, Bin
AU - Zhao, Weiyu
AU - Luo, Xiao
AU - Zhang, Xinfu
AU - Li, Chenglong
AU - Zeng, Chunxi
AU - Dong, Yizhou
N1 - Publisher Copyright:
© 2017 Macmillan Publishers Limited, part of Springer Nature. All rights reserved.
PY - 2017/5/10
Y1 - 2017/5/10
N2 - Cpf1, a type-V CRISPR-Cas effector endonuclease, exhibits gene-editing activity in human cells through a single RNA-guided approach. Here, we report the design and assessment of an array of 42 types of engineered Acidaminococcus sp. Cpf1 (AsCpf1) CRISPR RNA (crRNA) and 5 types of AsCpf1 mRNA in human cell lines. We show that the top-performing modified crRNA (cr3′5F, containing five 2′-fluoro ribose at the 3′ terminus) and AsCpf1 mRNA (full ψ-modification) improved gene-cutting efficiency by, respectively, 127% and 177%, with respect to unmodified crRNA and plasmid-encoding AsCpf1. We also show that the combination of cr3′5F and ψ-modified AsCpf1 or Lachnospiraceae bacterium Cpf1 (LbCpf1) mRNA augmented gene-cutting efficiency by over 300% with respect to the same control, and discovered that 11 out of 16 crRNAs from Cpf1 orthologues enabled genome editing in the presence of AsCpf1. Engineered CRISPR-Cpf1 systems should facilitate a broad range of genome editing applications.
AB - Cpf1, a type-V CRISPR-Cas effector endonuclease, exhibits gene-editing activity in human cells through a single RNA-guided approach. Here, we report the design and assessment of an array of 42 types of engineered Acidaminococcus sp. Cpf1 (AsCpf1) CRISPR RNA (crRNA) and 5 types of AsCpf1 mRNA in human cell lines. We show that the top-performing modified crRNA (cr3′5F, containing five 2′-fluoro ribose at the 3′ terminus) and AsCpf1 mRNA (full ψ-modification) improved gene-cutting efficiency by, respectively, 127% and 177%, with respect to unmodified crRNA and plasmid-encoding AsCpf1. We also show that the combination of cr3′5F and ψ-modified AsCpf1 or Lachnospiraceae bacterium Cpf1 (LbCpf1) mRNA augmented gene-cutting efficiency by over 300% with respect to the same control, and discovered that 11 out of 16 crRNAs from Cpf1 orthologues enabled genome editing in the presence of AsCpf1. Engineered CRISPR-Cpf1 systems should facilitate a broad range of genome editing applications.
UR - http://www.scopus.com/inward/record.url?scp=85029927053&partnerID=8YFLogxK
U2 - 10.1038/s41551-017-0066
DO - 10.1038/s41551-017-0066
M3 - Article
AN - SCOPUS:85029927053
SN - 2157-846X
VL - 1
JO - Nature Biomedical Engineering
JF - Nature Biomedical Engineering
IS - 5
M1 - 0066
ER -