Both TALENs and CRISPR/Cas9 directly target the HBB IVS2–654 (C > T) mutation in β-thalassemia-derived iPSCs
| Autor: | Peng Xu, Bo-yu Wang, Xiang Lv, Li Cheng, Xiu-zhen Liu, Tingting Wang, Yue Huang, Ying Tong, De-Pei Liu |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 2015 |
| Předmět: |
Transcription
Genetic Induced Pluripotent Stem Cells Karyotype Molecular Sequence Data beta-Globins Biology Article 03 medical and health sciences 0302 clinical medicine Gene Order CRISPR Humans Cell Self Renewal Induced pluripotent stem cell Homologous Recombination Gene 030304 developmental biology Genetics 0303 health sciences Transcription activator-like effector nuclease Multidisciplinary Base Sequence Cas9 Point mutation beta-Thalassemia Gene targeting Cell Differentiation Introns 3. Good health Genetic Loci 030220 oncology & carcinogenesis Gene Targeting Mutation CRISPR-Cas Systems Homologous recombination Sequence Alignment |
| Zdroj: | Scientific Reports |
| ISSN: | 2045-2322 |
| DOI: | 10.1038/srep12065 |
| Popis: | β-Thalassemia is one of the most common genetic blood diseases and is caused by either point mutations or deletions in the β-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. |
| Databáze: | OpenAIRE |
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