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dc.contributor.authorOoi, Amanda Siok Lee
dc.contributor.authorWong, Aloysius Tze
dc.contributor.authorEsau, Luke
dc.contributor.authorLemtiri-Chlieh, Fouad
dc.contributor.authorGehring, Christoph A
dc.date.accessioned2016-10-12T07:39:21Z
dc.date.available2016-10-12T07:39:21Z
dc.date.issued2016-07-19
dc.identifier.citationOoi A, Wong A, Esau L, Lemtiri-Chlieh F, Gehring C (2016) A Guide to Transient Expression of Membrane Proteins in HEK-293 Cells for Functional Characterization. Front Physiol 7. Available: http://dx.doi.org/10.3389/fphys.2016.00300.
dc.identifier.issn1664-042X
dc.identifier.pmid27486406
dc.identifier.doi10.3389/fphys.2016.00300
dc.identifier.urihttp://hdl.handle.net/10754/620932
dc.description.abstractThe human embryonic kidney 293 (HEK-293) cells are commonly used as host for the heterologous expression of membrane proteins not least because they have a high transfection efficiency and faithfully translate and process proteins. In addition, their cell size, morphology and division rate, and low expression of native channels are traits that are particularly attractive for current-voltage measurements. Nevertheless, the heterologous expression of complex membrane proteins such as receptors and ion channels for biological characterization and in particular for single-cell applications such as electrophysiology remains a challenge. Expression of functional proteins depends largely on careful step-by-step optimization that includes the design of expression vectors with suitable identification tags, as well as the selection of transfection methods and detection parameters appropriate for the application. Here, we use the heterologous expression of a plant potassium channel, the Arabidopsis thaliana guard cell outward-rectifying K+ channel, AtGORK (At5G37500) in HEK-293 cells as an example, to evaluate commonly used transfection reagents and fluorescent detection methods, and provide a detailed methodology for optimized transient transfection and expression of membrane proteins for in vivo studies in general and for single-cell applications in particular. This optimized protocol will facilitate the physiological and cellular characterization of complex membrane proteins.
dc.description.sponsorshipThis research was supported by King Abdullah University of Science and Technology (KAUST) (BAS/1/1013-01-01).
dc.publisherFrontiers Media SA
dc.relation.urlhttp://journal.frontiersin.org/article/10.3389/fphys.2016.00300/full
dc.rightsCopyright © 2016 Ooi, Wong, Esau, Lemtiri-Chlieh and Gehring. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
dc.rights.urihttp://creativecommons.org/licenses/by/4.0/
dc.subjectElectrophysiology
dc.subjectFluorescent imaging
dc.subjectHeterologous expression
dc.subjectHuman embryonic kidney 293 cells
dc.subjectMembrane proteins
dc.subjectTransfection
dc.titleA Guide to Transient Expression of Membrane Proteins in HEK-293 Cells for Functional Characterization
dc.typeArticle
dc.contributor.departmentBiological and Environmental Sciences and Engineering (BESE) Division
dc.contributor.departmentBioscience Program
dc.contributor.departmentComputational Bioscience Research Center (CBRC)
dc.contributor.departmentNGS, qPCR and Single Cell Genomics
dc.contributor.departmentSanger and Third Generation Sequencing
dc.contributor.departmentTechnology Transfer
dc.identifier.journalFrontiers in Physiology
dc.eprint.versionPublisher's Version/PDF
dc.contributor.institutionInstitute of Integrative Biology of the Cell, Centre National de la Recherche Scientifique, Le Commissariat à l'Energie Atomique et aux Energies Alternatives, Paris-Sud University, Gif-Sur-Yvette, France
kaust.personOoi, Amanda Siok Lee
kaust.personWong, Aloysius Tze
kaust.personEsau, Luke
kaust.personLemtiri-Chlieh, Fouad
kaust.personGehring, Christoph A.
kaust.grant.numberBAS/1/1013-01-01
refterms.dateFOA2018-06-13T18:06:21Z


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Copyright © 2016 Ooi, Wong, Esau, Lemtiri-Chlieh and Gehring. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
Except where otherwise noted, this item's license is described as Copyright © 2016 Ooi, Wong, Esau, Lemtiri-Chlieh and Gehring. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.