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Long QT syndrome type 3 (LQT3) is one of the more malignant congenital long QT syndrome (LQTS) variants with an\n                    <jats:italic>SCN5A<\/jats:italic>\n                    gain-of-function effect on the gated sodium channel. Moreover, the predominant pathogenic variants in LQTS genes are single nucleotide substitutions (missense) and small insertion\/deletions (INDEL). CRISPR\/Cas9 genome editing has been utilised to create isogenic hiPSCs to control for an identical genetic background and to isolate the pathogenicity of a single nucleotide change. In this study, we described an optimized and rapid protocol to introduce a heterozygous LQT3-specific variant into healthy control hiPSCs using ribonucleoprotein (RNP) and single-stranded oligonucleotide (ssODN). Based on this protocol, we successfully screened hiPSCs carrying a heterozygous LQT3 pathogenic variant (\n                    <jats:italic>SCN5A<\/jats:italic>\n                    <jats:sup>\u00b1<\/jats:sup>\n                    ) with high efficiency (6 out of 69) and confirmed no off-target effect, normal karyotype, high alkaline phosphatase activity, unaffected pluripotency, and\n                    <jats:italic>in vitro<\/jats:italic>\n                    embryonic body formation capacity within 2\u00a0weeks. In addition, we also provide protocols to robustly differentiate hiPSCs into cardiomyocytes and evaluate the electrophysiological characteristics using Multi-electrode Array. This protocol is also applicable to introduce and\/or correct other disease-specific variants into hiPSCs for future pharmacological screening and gene therapeutic development.\n                  <\/jats:p>\n                  <jats:p>\n                    <jats:bold>Graphical Abstract<\/jats:bold>\n                  <\/jats:p>","DOI":"10.1007\/s12015-023-10602-5","type":"journal-article","created":{"date-parts":[[2023,8,31]],"date-time":"2023-08-31T15:02:57Z","timestamp":1693494177000},"page":"2774-2789","update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":7,"title":["Using Ribonucleoprotein-based CRISPR\/Cas9 to Edit Single Nucleotide on Human Induced Pluripotent Stem Cells to Model Type 3 Long QT Syndrome (SCN5A\u00b1)"],"prefix":"10.1007","volume":"19","author":[{"given":"Ning","family":"Ge","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Min","family":"Liu","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Rui","family":"Li","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Nicholas M.","family":"Allen","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Joseph","family":"Galvin","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Sanbing","family":"Shen","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Timothy","family":"O\u2019Brien","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3824-636X","authenticated-orcid":false,"given":"Terence W.","family":"Prendiville","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"297","published-online":{"date-parts":[[2023,8,31]]},"reference":[{"issue":"12","key":"10602_CR1","doi-asserted-by":"publisher","first-page":"2827","DOI":"10.1253\/circj.cj-14-0905","volume":"78","author":"Y Mizusawa","year":"2014","unstructured":"Mizusawa, Y., Horie, M., & Wilde, A. A. (2014). Genetic and clinical advances in congenital long QT syndrome. Circulation Journal, 78(12), 2827\u20132833. https:\/\/doi.org\/10.1253\/circj.cj-14-0905","journal-title":"Circulation Journal"},{"issue":"8","key":"10602_CR2","doi-asserted-by":"publisher","first-page":"1308","DOI":"10.1016\/j.hrthm.2011.05.020","volume":"8","author":"MJ Ackerman","year":"2011","unstructured":"Ackerman, M. J., Priori, S. G., Willems, S., Berul, C., Brugada, R., Calkins, H., Camm, A. J., Ellinor, P. T., Gollob, M., Hamilton, R., Hershberger, R. E., Judge, D. P., Le Marec, H., McKenna, W. J., Schulze-Bahr, E., Semsarian, C., Towbin, J. A., Watkins, H., Wilde, A., \u2026 Zipes, D. P. (2011). HRS\/EHRA expert consensus statement on the state of genetic testing for the channelopathies and cardiomyopathies this document was developed as a partnership between the Heart Rhythm Society (HRS) and the European Heart Rhythm Association (EHRA). Heart Rhythm, 8(8), 1308\u20131339. https:\/\/doi.org\/10.1016\/j.hrthm.2011.05.020","journal-title":"Heart Rhythm"},{"issue":"4","key":"10602_CR3","doi-asserted-by":"publisher","first-page":"374","DOI":"10.1016\/j.jacep.2016.10.013","volume":"3","author":"A Porta-S\u00e1nchez","year":"2017","unstructured":"Porta-S\u00e1nchez, A., Spillane, D. R., Harris, L., Xue, J., Dorsey, P., Care, M., Chauhan, V., Gollob, M. H., & Spears, D. A. (2017). T-wave morphology analysis in congenital long QT syndrome discriminates patients from healthy individuals. JACC Clin Electrophysiol., 3(4), 374\u2013381. https:\/\/doi.org\/10.1016\/j.jacep.2016.10.013","journal-title":"JACC Clin Electrophysiol."},{"issue":"1","key":"10602_CR4","doi-asserted-by":"publisher","first-page":"12997","DOI":"10.1038\/s41598-019-49450-0","volume":"9","author":"HJ Kim","year":"2019","unstructured":"Kim, H. J., Kim, B. G., Park, J. E., Ki, C. S., Huh, J., Youm, J. B., Kang, J. S., & Cho, H. (2019). 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