Please use this identifier to cite or link to this item: http://hdl.handle.net/2440/115591
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dc.contributor.authorJolly, L.en
dc.contributor.authorSun, Y.en
dc.contributor.authorCarroll, R.en
dc.contributor.authorHoman, C.en
dc.contributor.authorGecz, J.en
dc.date.issued2018en
dc.identifier.citationJournal of Human Genetics, 2018; 63(9):945-955en
dc.identifier.issn1434-5161en
dc.identifier.issn1435-232Xen
dc.identifier.urihttp://hdl.handle.net/2440/115591-
dc.descriptionPublished online: 20 June 2018en
dc.description.abstractLymphoblastoid cell lines (LCLs) have been by far the most prevalent cell type used to study the genetics underlying normal and disease-relevant human phenotypic variation, across personal to epidemiological scales. In contrast, only few studies have explored the use of LCLs in functional genomics and mechanistic studies. Two major reasons are technical, as (1) interrogating the sub-cellular spatial information of LCLs is challenged by their non-adherent nature, and (2) LCLs are refractory to gene transfection. Methodological details relating to techniques that overcome these limitations are scarce, largely inadequate (without additional knowledge and expertise), and optimisation has never been described. Here we compare, optimise, and convey such methods in-depth. We provide a robust method to adhere LCLs to coverslips, which maintained cellular integrity, morphology, and permitted visualisation of sub-cellular structures and protein localisation. Next, we developed the use of lentiviral-based gene delivery to LCLs. Through empirical and combinatorial testing of multiple transduction conditions, we improved transduction efficiency from 3% up to 48%. Furthermore, we established strategies to purify transduced cells, to achieve sustainable cultures containing >85% transduced cells. Collectively, our methodologies provide a vital resource that enables the use of LCLs in functional cell and molecular biology experiments. Potential applications include the characterisation of genetic variants of unknown significance, the interrogation of cellular disease pathways and mechanisms, and high-throughput discovery of genetic modifiers of disease states among others.en
dc.description.statementofresponsibilityLachlan A. Jolly, Ying Sun, Renée Carroll, Claire C. Homan, Jozef Geczen
dc.language.isoenen
dc.publisherNature Publishing Groupen
dc.rights© The Author(s) under exclusive licence to The Japan Society of Human Genetics 2018en
dc.subjectLymphocytes; Cell Line; Humans; Lentivirus; Transduction, Genetic; Genetic Vectors; Female; Maleen
dc.titleRobust imaging and gene delivery to study human lymphoblastoid cell linesen
dc.typeJournal articleen
dc.identifier.rmid0030091778en
dc.identifier.doi10.1038/s10038-018-0483-2en
dc.relation.granthttp://purl.org/au-research/grants/arc/DE160100620en
dc.relation.granthttp://purl.org/au-research/grants/nhmrc/1041920en
dc.relation.granthttp://purl.org/au-research/grants/nhmrc/1091593en
dc.relation.granthttp://purl.org/au-research/grants/nhmrc/1063808en
dc.identifier.pubid428007-
pubs.library.collectionMolecular and Biomedical Science publicationsen
pubs.library.teamDS03en
pubs.verification-statusVerifieden
pubs.publication-statusPublisheden
dc.identifier.orcidJolly, L. [0000-0003-4538-2658]en
dc.identifier.orcidCarroll, R. [0000-0002-6979-3710]en
dc.identifier.orcidGecz, J. [0000-0002-7884-6861]en
Appears in Collections:Molecular and Biomedical Science publications

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