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Both TALENs and CRISPR/Cas9 directly target the HBB IVS2-654 (C? ?T) mutation in
?-thalassemia-derived iPSCs
#MMPMID26156589
Xu P
; Tong Y
; Liu XZ
; Wang TT
; Cheng L
; Wang BY
; Lv X
; Huang Y
; Liu DP
Sci Rep
2015[Jul]; 5
(?): 12065
PMID26156589
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?-Thalassemia is one of the most common genetic blood diseases and is caused by
either point mutations or deletions in the ?-globin (HBB) gene. The generation of
patient-specific induced pluripotent stem cells (iPSCs) and subsequent correction
of the disease-causing mutations may be a potential therapeutic strategy for this
disease. Due to the low efficiency of typical homologous recombination,
endonucleases, including TALENs and CRISPR/Cas9, have been widely used to enhance
the gene correction efficiency in patient-derived iPSCs. Here, we designed TALENs
and CRISPR/Cas9 to directly target the intron2 mutation site IVS2-654 in the
globin gene. We observed different frequencies of double-strand breaks (DSBs) at
IVS2-654 loci using TALENs and CRISPR/Cas9, and TALENs mediated a higher
homologous gene targeting efficiency compared to CRISPR/Cas9 when combined with
the piggyBac transposon donor. In addition, more obvious off-target events were
observed for CRISPR/Cas9 compared to TALENs. Finally, TALENs-corrected iPSC
clones were selected for erythroblast differentiation using the OP9 co-culture
system and detected relatively higher transcription of HBB than the uncorrected
cells. This comparison of using TALENs or CRISPR/Cas9 to correct specific HBB
mutations in patient-derived iPSCs will guide future applications of TALENs- or
CRISPR/Cas9-based gene therapies in monogenic diseases.
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