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2D and 3D Chromosome Painting in Malaria Mosquitoes

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dc.contributor.authorGeorge, Phillipen
dc.contributor.authorSharma, Atashien
dc.contributor.authorSharakhov, Igor V.en
dc.contributor.departmentEntomologyen
dc.date.accessioned2017-01-05T23:44:06Zen
dc.date.available2017-01-05T23:44:06Zen
dc.date.issued2014-01-01en
dc.description.abstractFluorescent in situ hybridization (FISH) of whole arm chromosome probes is a robust technique for mapping genomic regions of interest, detecting chromosomal rearrangements, and studying three-dimensional (3D) organization of chromosomes in the cell nucleus. The advent of laser capture microdissection (LCM) and whole genome amplification (WGA) allows obtaining large quantities of DNA from single cells. The increased sensitivity of WGA kits prompted us to develop chromosome paints and to use them for exploring chromosome organization and evolution in non-model organisms. Here, we present a simple method for isolating and amplifying the euchromatic segments of single polytene chromosome arms from ovarian nurse cells of the African malaria mosquito Anopheles gambiae. This procedure provides an efficient platform for obtaining chromosome paints, while reducing the overall risk of introducing foreign DNA to the sample. The use of WGA allows for several rounds of re-amplification, resulting in high quantities of DNA that can be utilized for multiple experiments, including 2D and 3D FISH. We demonstrated that the developed chromosome paints can be successfully used to establish the correspondence between euchromatic portions of polytene and mitotic chromosome arms in An. gambiae. Overall, the union of LCM and single-chromosome WGA provides an efficient tool for creating significant amounts of target DNA for future cytogenetic and genomic studies.en
dc.description.versionPublished versionen
dc.format.extent11 pagesen
dc.format.mimetypeapplication/pdfen
dc.identifier.doihttps://doi.org/10.3791/51173en
dc.identifier.issn1940-087Xen
dc.identifier.issue83en
dc.identifier.urihttp://hdl.handle.net/10919/73981en
dc.language.isoenen
dc.publisherJournal of Visualized Experimentsen
dc.relation.urihttp://gateway.webofknowledge.com/gateway/Gateway.cgi?GWVersion=2&SrcApp=PARTNER_APP&SrcAuth=LinksAMR&KeyUT=WOS:000348513500056&DestLinkType=FullRecord&DestApp=ALL_WOS&UsrCustomerID=930d57c9ac61a043676db62af60056c1en
dc.rightsCreative Commons Attribution-NonCommercial-NoDerivs 3.0 Unporteden
dc.rights.urihttp://creativecommons.org/licenses/by-nc-nd/3.0/en
dc.subjectimmunologyen
dc.subjectissue 83en
dc.subjectmicrodissectionen
dc.subjectwhole genome amplificationen
dc.subjectmalaria mosquitoen
dc.subjectpolytene chromosomeen
dc.subjectmitotic chromosomesen
dc.subjectfluorescence in situ hybridizationen
dc.subjectchromosome paintingen
dc.subjectwhole-genome amplificationen
dc.subjectlaser capture microdissectionen
dc.subjectpolytene chromosomesen
dc.subjectsex-chromosomesen
dc.subjectsystemic reorganizationen
dc.subjectphylogenetic levelsen
dc.subjectevolutionen
dc.subjectprobesen
dc.subjectcellsen
dc.subjectdnaen
dc.title2D and 3D Chromosome Painting in Malaria Mosquitoesen
dc.title.serialJove-Journal of Visualized Experimentsen
dc.typeArticle - Refereeden
dc.type.dcmitypeTexten
pubs.organisational-group/Virginia Techen
pubs.organisational-group/Virginia Tech/Agriculture & Life Sciencesen
pubs.organisational-group/Virginia Tech/Agriculture & Life Sciences/CALS T&R Facultyen
pubs.organisational-group/Virginia Tech/Agriculture & Life Sciences/Entomologyen
pubs.organisational-group/Virginia Tech/All T&R Facultyen
pubs.organisational-group/Virginia Tech/Faculty of Health Sciencesen

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