Cargando…

Efficient gene correction of an aberrant splice site in β‐thalassaemia iPSCs by CRISPR/Cas9 and single‐strand oligodeoxynucleotides

β‐thalassaemia is a prevalent hereditary haematological disease caused by mutations in the human haemoglobin β (HBB) gene. Among them, the HBB IVS2‐654 (C > T) mutation, which is in the intron, creates an aberrant splicing site. Bone marrow transplantation for curing β‐thalassaemia is limited due...

Descripción completa

Detalles Bibliográficos
Autores principales: Xiong, Zeyu, Xie, Yingjun, Yang, Yi, Xue, Yanting, Wang, Ding, Lin, Shouheng, Chen, Diyu, Lu, Dian, He, Lina, Song, Bing, Yang, Yinghong, Sun, Xiaofang
Formato: Online Artículo Texto
Lenguaje:English
Publicado: John Wiley and Sons Inc. 2019
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6850948/
https://www.ncbi.nlm.nih.gov/pubmed/31631510
http://dx.doi.org/10.1111/jcmm.14669
_version_ 1783469539577561088
author Xiong, Zeyu
Xie, Yingjun
Yang, Yi
Xue, Yanting
Wang, Ding
Lin, Shouheng
Chen, Diyu
Lu, Dian
He, Lina
Song, Bing
Yang, Yinghong
Sun, Xiaofang
author_facet Xiong, Zeyu
Xie, Yingjun
Yang, Yi
Xue, Yanting
Wang, Ding
Lin, Shouheng
Chen, Diyu
Lu, Dian
He, Lina
Song, Bing
Yang, Yinghong
Sun, Xiaofang
author_sort Xiong, Zeyu
collection PubMed
description β‐thalassaemia is a prevalent hereditary haematological disease caused by mutations in the human haemoglobin β (HBB) gene. Among them, the HBB IVS2‐654 (C > T) mutation, which is in the intron, creates an aberrant splicing site. Bone marrow transplantation for curing β‐thalassaemia is limited due to the lack of matched donors. The clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR‐associated protein 9 (Cas9), as a widely used tool for gene editing, is able to target specific sequence and create double‐strand break (DSB), which can be combined with the single‐stranded oligodeoxynucleotide (ssODN) to correct mutations. In this study, according to two different strategies, the HBB IVS2‐654 mutation was seamlessly corrected in iPSCs by CRISPR/Cas9 system and ssODN. To reduce the occurrence of secondary cleavage, a more efficient strategy was adopted. The corrected iPSCs kept pluripotency and genome stability. Moreover, they could differentiate normally. Through CRISPR/Cas9 system and ssODN, our study provides improved strategies for gene correction of β‐Thalassaemia, and the expression of the HBB gene can be restored, which can be used for gene therapy in the future.
format Online
Article
Text
id pubmed-6850948
institution National Center for Biotechnology Information
language English
publishDate 2019
publisher John Wiley and Sons Inc.
record_format MEDLINE/PubMed
spelling pubmed-68509482019-12-01 Efficient gene correction of an aberrant splice site in β‐thalassaemia iPSCs by CRISPR/Cas9 and single‐strand oligodeoxynucleotides Xiong, Zeyu Xie, Yingjun Yang, Yi Xue, Yanting Wang, Ding Lin, Shouheng Chen, Diyu Lu, Dian He, Lina Song, Bing Yang, Yinghong Sun, Xiaofang J Cell Mol Med Original Articles β‐thalassaemia is a prevalent hereditary haematological disease caused by mutations in the human haemoglobin β (HBB) gene. Among them, the HBB IVS2‐654 (C > T) mutation, which is in the intron, creates an aberrant splicing site. Bone marrow transplantation for curing β‐thalassaemia is limited due to the lack of matched donors. The clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR‐associated protein 9 (Cas9), as a widely used tool for gene editing, is able to target specific sequence and create double‐strand break (DSB), which can be combined with the single‐stranded oligodeoxynucleotide (ssODN) to correct mutations. In this study, according to two different strategies, the HBB IVS2‐654 mutation was seamlessly corrected in iPSCs by CRISPR/Cas9 system and ssODN. To reduce the occurrence of secondary cleavage, a more efficient strategy was adopted. The corrected iPSCs kept pluripotency and genome stability. Moreover, they could differentiate normally. Through CRISPR/Cas9 system and ssODN, our study provides improved strategies for gene correction of β‐Thalassaemia, and the expression of the HBB gene can be restored, which can be used for gene therapy in the future. John Wiley and Sons Inc. 2019-10-21 2019-12 /pmc/articles/PMC6850948/ /pubmed/31631510 http://dx.doi.org/10.1111/jcmm.14669 Text en © 2019 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine. This is an open access article under the terms of the http://creativecommons.org/licenses/by/4.0/ License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited.
spellingShingle Original Articles
Xiong, Zeyu
Xie, Yingjun
Yang, Yi
Xue, Yanting
Wang, Ding
Lin, Shouheng
Chen, Diyu
Lu, Dian
He, Lina
Song, Bing
Yang, Yinghong
Sun, Xiaofang
Efficient gene correction of an aberrant splice site in β‐thalassaemia iPSCs by CRISPR/Cas9 and single‐strand oligodeoxynucleotides
title Efficient gene correction of an aberrant splice site in β‐thalassaemia iPSCs by CRISPR/Cas9 and single‐strand oligodeoxynucleotides
title_full Efficient gene correction of an aberrant splice site in β‐thalassaemia iPSCs by CRISPR/Cas9 and single‐strand oligodeoxynucleotides
title_fullStr Efficient gene correction of an aberrant splice site in β‐thalassaemia iPSCs by CRISPR/Cas9 and single‐strand oligodeoxynucleotides
title_full_unstemmed Efficient gene correction of an aberrant splice site in β‐thalassaemia iPSCs by CRISPR/Cas9 and single‐strand oligodeoxynucleotides
title_short Efficient gene correction of an aberrant splice site in β‐thalassaemia iPSCs by CRISPR/Cas9 and single‐strand oligodeoxynucleotides
title_sort efficient gene correction of an aberrant splice site in β‐thalassaemia ipscs by crispr/cas9 and single‐strand oligodeoxynucleotides
topic Original Articles
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6850948/
https://www.ncbi.nlm.nih.gov/pubmed/31631510
http://dx.doi.org/10.1111/jcmm.14669
work_keys_str_mv AT xiongzeyu efficientgenecorrectionofanaberrantsplicesiteinbthalassaemiaipscsbycrisprcas9andsinglestrandoligodeoxynucleotides
AT xieyingjun efficientgenecorrectionofanaberrantsplicesiteinbthalassaemiaipscsbycrisprcas9andsinglestrandoligodeoxynucleotides
AT yangyi efficientgenecorrectionofanaberrantsplicesiteinbthalassaemiaipscsbycrisprcas9andsinglestrandoligodeoxynucleotides
AT xueyanting efficientgenecorrectionofanaberrantsplicesiteinbthalassaemiaipscsbycrisprcas9andsinglestrandoligodeoxynucleotides
AT wangding efficientgenecorrectionofanaberrantsplicesiteinbthalassaemiaipscsbycrisprcas9andsinglestrandoligodeoxynucleotides
AT linshouheng efficientgenecorrectionofanaberrantsplicesiteinbthalassaemiaipscsbycrisprcas9andsinglestrandoligodeoxynucleotides
AT chendiyu efficientgenecorrectionofanaberrantsplicesiteinbthalassaemiaipscsbycrisprcas9andsinglestrandoligodeoxynucleotides
AT ludian efficientgenecorrectionofanaberrantsplicesiteinbthalassaemiaipscsbycrisprcas9andsinglestrandoligodeoxynucleotides
AT helina efficientgenecorrectionofanaberrantsplicesiteinbthalassaemiaipscsbycrisprcas9andsinglestrandoligodeoxynucleotides
AT songbing efficientgenecorrectionofanaberrantsplicesiteinbthalassaemiaipscsbycrisprcas9andsinglestrandoligodeoxynucleotides
AT yangyinghong efficientgenecorrectionofanaberrantsplicesiteinbthalassaemiaipscsbycrisprcas9andsinglestrandoligodeoxynucleotides
AT sunxiaofang efficientgenecorrectionofanaberrantsplicesiteinbthalassaemiaipscsbycrisprcas9andsinglestrandoligodeoxynucleotides