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Combining Single Strand Oligodeoxynucleotides and CRISPR/Cas9 to Correct Gene Mutations in β-Thalassemia-induced Pluripotent Stem Cells
| Content Provider | Scilit |
|---|---|
| Author | Niu, Xiaohua He, Wenyin Song, Bing Ou, Zhanhui Fan, Di Chen, Yuchang Fan, Yong Sun, Xiaofang |
| Copyright Year | 2016 |
| Description | Journal: Journal of Biological Chemistry β-Thalassemia (β-Thal) is one of the most common genetic diseases in the world. The generation of patient-specific β-Thal-induced pluripotent stem cells (iPSCs), correction of the disease-causing mutations in those cells, and then differentiation into hematopoietic stem cells offers a new therapeutic strategy for this disease. Here, we designed a CRISPR/Cas9 to specifically target the Homo sapiens hemoglobin β (HBB) gene CD41/42(−CTTT) mutation. We demonstrated that the combination of single strand oligodeoxynucleotides with CRISPR/Cas9 was capable of correcting the HBB gene CD41/42 mutation in β-Thal iPSCs. After applying a correction-specific PCR assay to purify the corrected clones followed by sequencing to confirm mutation correction, we verified that the purified clones retained full pluripotency and exhibited normal karyotyping. Additionally, whole-exome sequencing showed that the mutation load to the exomes was minimal after CRISPR/Cas9 targeting. Furthermore, the corrected iPSCs were selected for erythroblast differentiation and restored the expression of HBB protein compared with the parental iPSCs. This method provides an efficient and safe strategy to correct the HBB gene mutation in β-Thal iPSCs. |
| Related Links | http://www.jbc.org/content/291/32/16576.full.pdf https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4974373/pdf http://www.jbc.org/article/S0021925820353813/pdf |
| Ending Page | 16585 |
| Page Count | 10 |
| Starting Page | 16576 |
| ISSN | 00219258 |
| e-ISSN | 1083351X |
| DOI | 10.1074/jbc.m116.719237 |
| Journal | Journal of Biological Chemistry |
| Issue Number | 32 |
| Volume Number | 291 |
| Language | English |
| Publisher | Elsevier BV |
| Publisher Date | 2016-08-01 |
| Access Restriction | Open |
| Subject Keyword | Journal: Journal of Biological Chemistry Cell Tissue Engineering Induced Pluripotent Stem Cell (ips Cell) (ipsc) |
| Content Type | Text |
| Resource Type | Article |
| Subject | Cell Biology Biochemistry Molecular Biology |
