Generation of ΔF508-CFTR T84 cell lines by CRISPR/Cas9-mediated genome editing

Woo Young Chung, Myungjae Song, Jinhong Park, Wan Namkung, Jinu Lee, Hyongbum Kim, Min Goo Lee, Joo Young Kim

Research output: Contribution to journalArticle

3 Citations (Scopus)

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.

Original languageEnglish
Pages (from-to)2023-2034
Number of pages12
JournalBiotechnology Letters
Volume38
Issue number12
DOIs
Publication statusPublished - 2016 Dec 1

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Clustered Regularly Interspaced Short Palindromic Repeats
Genes
Cells
Cell Line
Screening
Lentivirus Infections
Guide RNA
Endoplasmic Reticulum-Associated Degradation
Genome
Proteins
Protein folding
Cystic Fibrosis Transmembrane Conductance Regulator
Protein Folding
RNA
Proteolysis
Exons
Membranes
Degradation
Temperature
Gene Editing

All Science Journal Classification (ASJC) codes

  • Biotechnology

Cite this

Chung, Woo Young ; Song, Myungjae ; Park, Jinhong ; Namkung, Wan ; Lee, Jinu ; Kim, Hyongbum ; Lee, Min Goo ; Kim, Joo Young. / Generation of ΔF508-CFTR T84 cell lines by CRISPR/Cas9-mediated genome editing. In: Biotechnology Letters. 2016 ; Vol. 38, No. 12. pp. 2023-2034.
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Generation of ΔF508-CFTR T84 cell lines by CRISPR/Cas9-mediated genome editing. / Chung, Woo Young; Song, Myungjae; Park, Jinhong; Namkung, Wan; Lee, Jinu; Kim, Hyongbum; Lee, Min Goo; Kim, Joo Young.

In: Biotechnology Letters, Vol. 38, No. 12, 01.12.2016, p. 2023-2034.

Research output: Contribution to journalArticle

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AU - Chung, Woo Young

AU - Song, Myungjae

AU - Park, Jinhong

AU - Namkung, Wan

AU - Lee, Jinu

AU - Kim, Hyongbum

AU - Lee, Min Goo

AU - Kim, Joo Young

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AB - 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.

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