Construction of a human hTERT RPE-1 cell line with inducible Cre for editing of endogenous genes.

Autor:	Hindul NL; Department of Molecular and Cell Biology, Henry Wellcome Building, University of Leicester, University Road, Leicester LE1 7RH, UK., Jhita A; Department of Molecular and Cell Biology, Henry Wellcome Building, University of Leicester, University Road, Leicester LE1 7RH, UK., Oprea DG; Department of Molecular and Cell Biology, Henry Wellcome Building, University of Leicester, University Road, Leicester LE1 7RH, UK., Hussain TA; Department of Molecular and Cell Biology, Henry Wellcome Building, University of Leicester, University Road, Leicester LE1 7RH, UK., Gonchar O; Department of Molecular and Cell Biology, Henry Wellcome Building, University of Leicester, University Road, Leicester LE1 7RH, UK., Campillo MAM; Department of Molecular and Cell Biology, Henry Wellcome Building, University of Leicester, University Road, Leicester LE1 7RH, UK., O'Regan L; Department of Molecular and Cell Biology, Henry Wellcome Building, University of Leicester, University Road, Leicester LE1 7RH, UK., Kanemaki MT; Department of Chromosome Science, National Institute of Genetics, Research Organization of Information and Systems (ROIS), Yata 1111, Mishima, Shizuoka 411-8540, Japan.; Department of Genetics, SOKENDAI, Mishima, Shizuoka 411-8540, Japan., Fry AM; Department of Molecular and Cell Biology, Henry Wellcome Building, University of Leicester, University Road, Leicester LE1 7RH, UK., Hirota K; Department of Chemistry, Graduate School of Science, Tokyo Metropolitan University, Minamiosawa 1-1, Hachioji-shi, Tokyo, 192-0397, Japan., Tanaka K; Department of Molecular and Cell Biology, Henry Wellcome Building, University of Leicester, University Road, Leicester LE1 7RH, UK.
Jazyk:	angličtina
Zdroj:	Biology open [Biol Open] 2022 Feb 15; Vol. 11 (2). Date of Electronic Publication: 2022 Feb 16.
DOI:	10.1242/bio.059056
Abstrakt:	The human retinal pigment epithelial RPE-1 cell line immortalized with hTERT retains a stable karyotype with a modal chromosome number of 46 and has been widely used to study physiological events in human cell culture systems. To facilitate inducible knock-out or knock-in experiments in this cell line, we have modified the AAVS1 locus to harbour a DNA fragment encoding ERT2-Cre-ERT2 fusion protein under regulation of a Tet-On expression system. In the generated cell line, active Cre recombinase was induced by simple addition of doxycycline and tamoxifen to the culture medium. As proof of concept, we successfully introduced an oncogenic point mutation to the endogenous KRAS gene locus of this cell line. The cell line will serve as a powerful tool to conduct functional analyses of human genes. Competing Interests: Competing interests The authors declare no competing or financial interests. (© 2022. Published by The Company of Biologists Ltd.)
Databáze:	MEDLINE
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