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dc.date.accessioned2017-03-03T08:43:54Z
dc.date.available2017-03-03T08:43:54Z
dc.date.created2012-02-06T10:00:27Z
dc.date.issued2011
dc.identifier.citationWälchli, Sebastien Løset, Geir Åge Kumari, Shraddha Johansen, Jorunn Yang, Weiwen Sandlie, Inger Olweus, Johanna . A Practical Approach to T-Cell Receptor Cloning and Expression. PLoS ONE. 2011, 6(11)
dc.identifier.urihttp://hdl.handle.net/10852/54165
dc.description.abstractAlthough cloning and expression of T-cell Receptors (TcRs) has been performed for almost two decades, these procedures are still challenging. For example, the use of T-cell clones that have undergone limited expansion as starting material to limit the loss of interesting TcRs, must be weighed against the introduction of mutations by excess PCR cycles. The recent interest in using specific TcRs for cancer immunotherapy has, however, increased the demand for practical and robust methods to rapidly clone and express TcRs. Two main technologies for TcR cloning have emerged; the use of a set of primers specifically annealing to all known TcR variable domains, and 59-RACE amplification. We here present an improved 59-RACE protocol that represents a fast and reliable way to identify a TcR from 105 cells only, making TcR cloning feasible without a priori knowledge of the variable domain sequence. We further present a detailed procedure for the subcloning of TcRa and b chains into an expression system. We show that a recombination-based cloning protocol facilitates simple and rapid transfer of the TcR transgene into different expression systems. The presented comprehensive method can be performed in any laboratory with standard equipment and with a limited amount of starting material. We finally exemplify the straightforwardness and reliability of our procedure by cloning and expressing several MART-1-specific TcRs and demonstrating their functionality. © 2011 Wälchli et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.en_US
dc.languageEN
dc.publisherPublic Library of Science (PLoS)
dc.titleA Practical Approach to T-Cell Receptor Cloning and Expressionen_US
dc.typeJournal articleen_US
dc.creator.authorWälchli, Sebastien
dc.creator.authorLøset, Geir Åge
dc.creator.authorKumari, Shraddha
dc.creator.authorJohansen, Jorunn
dc.creator.authorYang, Weiwen
dc.creator.authorSandlie, Inger
dc.creator.authorOlweus, Johanna
cristin.unitcode185,53,2,11
cristin.unitnameSenter for immunregulering
cristin.ispublishedtrue
cristin.fulltextoriginal
cristin.qualitycode1
dc.identifier.cristin905353
dc.identifier.bibliographiccitationinfo:ofi/fmt:kev:mtx:ctx&ctx_ver=Z39.88-2004&rft_val_fmt=info:ofi/fmt:kev:mtx:journal&rft.jtitle=PLoS ONE&rft.volume=6&rft.spage=&rft.date=2011
dc.identifier.jtitlePLoS ONE
dc.identifier.volume6
dc.identifier.issue11
dc.identifier.pagecount11
dc.identifier.doihttp://dx.doi.org/10.1371/journal.pone.0027930
dc.identifier.urnURN:NBN:no-57293
dc.type.documentTidsskriftartikkelen_US
dc.type.peerreviewedPeer reviewed
dc.source.issn1932-6203
dc.identifier.fulltextFulltext https://www.duo.uio.no/bitstream/handle/10852/54165/1/31.%2Bpone.0027930.pdf
dc.type.versionPublishedVersion
cristin.articleide27930


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