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Complete Knockout o...
Complete Knockout of Endogenous Mdr1 (Abcb1) in MDCK Cells by CRISPR-Cas9
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- Simoff, Ivailo (författare)
- Uppsala universitet,Institutionen för farmaci,Science for Life Laboratory, SciLifeLab
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- Karlgren, Maria (författare)
- Uppsala universitet,Institutionen för farmaci
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- Backlund, Maria (författare)
- Uppsala universitet,Institutionen för farmaci,Uppsala University Drug Optimization and Pharmaceutical Profiling Platform (UDOPP), Uppsala, Sweden
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- Lindström, Anne-Christine (författare)
- Uppsala universitet,Science for Life Laboratory, SciLifeLab,Institutionen för immunologi, genetik och patologi
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- Gaugaz, Fabienne Z. (författare)
- Uppsala universitet,Institutionen för farmaci
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- Matsson, Pär (författare)
- Uppsala universitet,Institutionen för farmaci
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- Artursson, Per (författare)
- Uppsala universitet,Institutionen för farmaci,Science for Life Laboratory, SciLifeLab,Uppsala University Drug Optimization and Pharmaceutical Profiling Platform (UDOPP), Uppsala, Sweden
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(creator_code:org_t)
- 2016
- 2016
- Engelska.
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Ingår i: Journal of Pharmaceutical Sciences. - 0022-3549 .- 1520-6017. ; 105:2, s. 1017-1021
- Relaterad länk:
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https://urn.kb.se/re...
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https://doi.org/10.1...
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Abstract
Ämnesord
Stäng
- Madin-Darby canine kidney II cells transfected with one or several transport proteins are commonly used models to study drug transport. In these cells, however, endogenous transporters such as canine Mdr1/P-glycoprotein (Abcb1) complicate the interpretation of transport studies. The aim of this investigation was to establish a Madin-Darby canine kidney II cell line using CRISPR-Cas9 gene-editing technology to knock out endogenous canine Mdr1 (cMdr1) expression. CRISPR-Cas9-mediated Abcb1 homozygous disruption occurred at frequencies of around 20% and resulted in several genotypes. We selected 1 clonal cell line, cMdr1 KO Cl2, for further examination. Consistent with an on-target effect of CRISPR-Cas9 in specific regions of the endogenous canine Abcb1 gene, we obtained a cell clone with Abcb1 gene alterations and without any cMdr1 expression, as confirmed by genome sequencing and quantitative protein analysis. Functional studies of these cells, using digoxin and other prototypic MDR1 substrates, showed close to identical transport in the apical-to-basolateral and basolateral-to-apical directions, resulting in efflux ratios indistinguishable from unity.
Ämnesord
- MEDICIN OCH HÄLSOVETENSKAP -- Medicinska och farmaceutiska grundvetenskaper -- Farmaceutiska vetenskaper (hsv//swe)
- MEDICAL AND HEALTH SCIENCES -- Basic Medicine -- Pharmaceutical Sciences (hsv//eng)
Nyckelord
- MDCK cells
- CRISPR-Cas9
- ABC transporters
- efflux pumps
- P-glycoprotein
- permeability
- membrane transporter
- drug transport
- genomics
- proteomics
Publikations- och innehållstyp
- ref (ämneskategori)
- art (ämneskategori)
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