% pubman genre = article
@article{item_3487971,
title = {{Detection of unintended on-target effects in CRISPR genome editing by DNA donors carrying diagnostic substitutions}},
author = {Lackner, Martin and Helmbrecht, Nelly and P{\"a}{\"a}bo, Svante and Riesenberg, Stephan},
language = {eng},
issn = {0305-1048; 1362-4962},
doi = {10.1093/nar/gkac1254},
publisher = {Oxford University Press},
year = {2023},
date = {2023-03-21},
abstract = {{CRISPR nucleases can introduce double-stranded DNA breaks in genomes at positions specified by guide RNAs. When repaired by the cell, this may result in the introduction of insertions and deletions or nucleotide substitutions provided by exogenous DNA donors. However, cellular repair can also result in unintended on-target effects, primarily larger deletions and loss of heterozygosity due to gene conversion. Here we present a strategy that allows easy and reliable detection of unintended on-target effects as well as the generation of control cells that carry wild-type alleles but have demonstratively undergone genome editing at the target site. Our {\textquoteleft}sequence-ascertained favorable editing{\textquoteright} (SAFE) donor approach relies on the use of DNA donor mixtures containing the desired nucleotide substitutions or the wild-type alleles together with combinations of additional {\textquoteleft}diagnostic{\textquoteright} substitutions unlikely to have any effects. Sequencing of the target sites then results in that two different sequences are seen when both chromosomes are edited with {\textquoteleft}SAFE{\textquoteright} donors containing different sets of substitutions, while a single sequence indicates unintended effects such as deletions or gene conversion. We analyzed more than 850 human embryonic stem cell clones edited with {\textquoteleft}SAFE{\textquoteright} donors and detect all copy number changes and almost all clones with gene conversion.}},
journal = {{Nucleic Acids Research}},
volume = {51},
number = {5},
eid = {gkac1254},
}