دورية أكاديمية
CRISPR/Cas9-based targeting of fluorescent reporters to human iPSCs to isolate atrial and ventricular-specific cardiomyocytes
| العنوان: | CRISPR/Cas9-based targeting of fluorescent reporters to human iPSCs to isolate atrial and ventricular-specific cardiomyocytes |
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| المؤلفون: | Orlando Chirikian, William R. Goodyer, Elda Dzilic, Vahid Serpooshan, Jan W. Buikema, Wesley McKeithan, HaoDi Wu, Guang Li, Soah Lee, Markus Merk, Francisco Galdos, Aimee Beck, Alexandre J. S. Ribeiro, Sharon Paige, Mark Mercola, Joseph C. Wu, Beth L. Pruitt, Sean M. Wu |
| المصدر: | Scientific Reports, Vol 11, Iss 1, Pp 1-10 (2021) |
| بيانات النشر: | Nature Portfolio, 2021. |
| سنة النشر: | 2021 |
| المجموعة: | LCC:Medicine LCC:Science |
| مصطلحات موضوعية: | Medicine, Science |
| الوصف: | Abstract Generating cardiomyocytes (CMs) from human induced pluripotent stem cells (hiPSCs) has represented a significant advance in our ability to model cardiac disease. Current differentiation protocols, however, have limited use due to their production of heterogenous cell populations, primarily consisting of ventricular-like CMs. Here we describe the creation of two chamber-specific reporter hiPSC lines by site-directed genomic integration using CRISPR-Cas9 technology. In the MYL2-tdTomato reporter, the red fluorescent tdTomato was inserted upstream of the 3′ untranslated region of the Myosin Light Chain 2 (MYL2) gene in order faithfully label hiPSC-derived ventricular-like CMs while avoiding disruption of endogenous gene expression. Similarly, in the SLN-CFP reporter, Cyan Fluorescent Protein (CFP) was integrated downstream of the coding region of the atrial-specific gene, Sarcolipin (SLN). Purification of tdTomato+ and CFP+ CMs using flow cytometry coupled with transcriptional and functional characterization validated these genetic tools for their use in the isolation of bona fide ventricular-like and atrial-like CMs, respectively. Finally, we successfully generated a double reporter system allowing for the isolation of both ventricular and atrial CM subtypes within a single hiPSC line. These tools provide a platform for chamber-specific hiPSC-derived CM purification and analysis in the context of atrial- or ventricular-specific disease and therapeutic opportunities. |
| نوع الوثيقة: | article |
| وصف الملف: | electronic resource |
| اللغة: | English |
| تدمد: | 2045-2322 |
| العلاقة: | https://doaj.org/toc/2045-2322Test |
| DOI: | 10.1038/s41598-021-81860-x |
| الوصول الحر: | https://doaj.org/article/d5033e1c1320405bbdfe19a5a5ff8a89Test |
| رقم الانضمام: | edsdoj.5033e1c1320405bbdfe19a5a5ff8a89 |
| قاعدة البيانات: | Directory of Open Access Journals |
Full Text Finder | تدمد: | 20452322 |
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| DOI: | 10.1038/s41598-021-81860-x |