TY - JOUR
T1 - High-efficiency genomic editing in Epstein-Barr virus-transformed lymphoblastoid B cells using a single-stranded donor oligonucleotide strategy
AU - Johnston, Andrew D.
AU - Simões-Pires, Claudia A.
AU - Suzuki, Masako
AU - Greally, John M.
N1 - Funding Information:
The authors thank the following Einstein core facilities for their expertize: the Epige-nomics Shared Facility, the Flow Cytometry Core Facility and the Genomics Core Facility. The feedback of colleagues at the New York Genome Center, especially the laboratory of Dr. Tuuli Lappalainen, is also gratefully acknowledged. A.D.J. was supported by Einstein’s Medical Scientist Training Program, National Institutes of Health (NIH) NIGMS T32 GM007288. C.A.S.P. was supported by a Marie Sklodowska-Curie grant agreement No 750190.
Publisher Copyright:
© 2019, The Author(s).
PY - 2019/12/1
Y1 - 2019/12/1
N2 - While human lymphoblastoid cell lines represent a valuable resource for population genetic studies, they have usually been regarded as difficult for CRISPR-mediated genomic editing because of very inefficient DNA transfection and retroviral or lentiviral transduction in these cells, which becomes a substantial problem when multiple constructs need to be co-expressed. Here we describe a protocol using a single-stranded donor oligonucleotide strategy for ‘scarless’ editing in lymphoblastoid cells, yielding 12/60 (20%) of clones with homology-directed recombination, when rates of <5–10% are frequently typical for many other cell types. The protocol does not require the use of lentiviruses or stable transfection, permitting lymphoblastoid cell lines to be used for CRISPR-mediated genomic targeting and screening in population genetic studies.
AB - While human lymphoblastoid cell lines represent a valuable resource for population genetic studies, they have usually been regarded as difficult for CRISPR-mediated genomic editing because of very inefficient DNA transfection and retroviral or lentiviral transduction in these cells, which becomes a substantial problem when multiple constructs need to be co-expressed. Here we describe a protocol using a single-stranded donor oligonucleotide strategy for ‘scarless’ editing in lymphoblastoid cells, yielding 12/60 (20%) of clones with homology-directed recombination, when rates of <5–10% are frequently typical for many other cell types. The protocol does not require the use of lentiviruses or stable transfection, permitting lymphoblastoid cell lines to be used for CRISPR-mediated genomic targeting and screening in population genetic studies.
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U2 - 10.1038/s42003-019-0559-3
DO - 10.1038/s42003-019-0559-3
M3 - Article
C2 - 31925152
AN - SCOPUS:85071182351
SN - 2399-3642
VL - 2
JO - Communications Biology
JF - Communications Biology
IS - 1
M1 - 312
ER -