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Series GSE135064 Query DataSets for GSE135064
Status Public on Dec 04, 2019
Title Expanding the editable genome and CRISPR-Cas9 versatility using DNA cutting-free gene targeting based on in trans paired nicking
Organism Homo sapiens
Experiment type Other
Summary Genome editing typically involves recombination between donor nucleic acids and genomic sequences subjected to double-stranded DNA breaks (DSBs) made by programmable nucleases (e.g. CRISPR-Cas9). Yet, amongst other deleterious by-products, DSBs yield translocations, off-target mutations and, most pervasively, unpredictable on-target allelic disruptions. Remarkably, hitherto, the untoward phenotypic consequences of on-target disruptions at allelic and non-allelic (e.g. pseudogene) sequences have received scant scrutiny and, crucially, remain to be addressed. Here, we demonstrate that gene-edited cells can lose fitness due to on-target DSBs and report that simultaneous single-stranded DNA break formation at donor and target DNA by CRISPR-Cas9 “nickases” overcomes, to a great extent, such genotype-phenotype disrupting events. Moreover, in trans paired nicking gene editing can efficiently and precisely add large DNA segments (i.e. live-cell reporter tags) into essential and multiple-copy genomic sequences while circumventing most of the allelic and non-allelic collateral mutations and chromosomal rearrangements characteristic of nuclease-dependent gene editing procedures.
Overall design We employed an saCas9:RAG1.1 gRNA bait DNA double strand break to map genome-wide repair outcomes in the presence or absence of the spCas9:AAVS1 gRNA nuclease or the D10A nickase variant using LAM-HTGTS
Contributor(s) Chen X, Tasca F, Frock RL, Goncalves MA
Citation(s) 31799604
Submission date Jul 29, 2019
Last update date Dec 06, 2019
Contact name Richard L Frock
Organization name Stanford University
Department Radiation Oncology
Lab Frock Lab
Street address 269 Campus Drive
City Stanford
State/province California
ZIP/Postal code 94305
Country USA
Platforms (1)
GPL15520 Illumina MiSeq (Homo sapiens)
Samples (12)
GSM3984757 saC9_R1_Rep1
GSM3984758 saC9_R1_spC9D10A_AAVS1_Rep1
GSM3984759 saC9_R1_spC9_AAVS1_Rep1
BioProject PRJNA557250
SRA SRP216762

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Supplementary file Size Download File type/resource
GSE135064_iPSC_saC9_R1.txt.gz 405.6 Kb (ftp)(http) TXT
GSE135064_iPSC_saC9_R1_spC9D10A_AAVS1.txt.gz 141.5 Kb (ftp)(http) TXT
GSE135064_iPSC_saC9_R1_spC9_AAVS1.txt.gz 166.3 Kb (ftp)(http) TXT
GSE135064_saC9_R1_s3.txt.gz 1.9 Mb (ftp)(http) TXT
GSE135064_saC9_R1_spC9D10A_AAVS1_s3.txt.gz 1.3 Mb (ftp)(http) TXT
GSE135064_saC9_R1_spC9_AAVS1_s3.txt.gz 2.1 Mb (ftp)(http) TXT
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Processed data are available on Series record

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