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- Title
Generation of ΔF508-CFTR T84 cell lines by CRISPR/Cas9-mediated genome editing.
- Authors
Chung, Woo; Kim, Hyongbum; Lee, Min; Kim, Joo; Song, Myungjae; Park, Jinhong; Namkung, Wan; Lee, Jinu
- Abstract
Objectives: To provide a simple method to make a stable ΔF508-CFTR-expressing T84 cell line that can be used as an efficient screening model system for ΔF508-CFTR rescue. Results: CFTR knockout cell lines were generated by Cas9 with a single-guide RNA (sgRNA) targeting exon 1 of the CFTR genome, which produced indels that abolished CFTR protein expressions. Next, stable ΔF508-CFTR expression was achieved by genome integration of ΔF508-CFTR via the lentivirus infection system. Finally, we showed functional rescue of ΔF508-CFTR not only by growing the cells at a low temperature, but also incubating with VX-809, a ΔF508-CFTR corrector, in the established T84 cells expressing ΔF508-CFTR. Conclusions: This cell system provides an appropriate screening platform for rescue of ΔF508-CFTR, especially related to protein folding, escaped from endoplasmic-reticulum-associated protein degradation, and membrane transport.
- Subjects
CYSTIC fibrosis transmembrane conductance regulator; CELL lines; CRISPRS; GENOME editing; GENETIC mutation; EPITHELIAL cells; GENE knockout; LENTIVIRUSES
- Publication
Biotechnology Letters, 2016, Vol 38, Issue 12, p2023
- ISSN
0141-5492
- Publication type
Article
- DOI
10.1007/s10529-016-2190-4