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[{"key": "dc.contributor.author", "value": "Myllynen, Mira", "language": null, "element": "contributor", "qualifier": "author", "schema": "dc"}, {"key": "dc.date.accessioned", "value": "2014-09-05T10:33:08Z", "language": "", "element": "date", "qualifier": "accessioned", "schema": "dc"}, {"key": "dc.date.available", "value": "2014-09-05T10:33:08Z", "language": "", "element": "date", "qualifier": "available", "schema": "dc"}, {"key": "dc.date.issued", "value": "2014", "language": null, "element": "date", "qualifier": "issued", "schema": "dc"}, {"key": "dc.identifier.other", "value": "oai:jykdok.linneanet.fi:1445144", "language": null, "element": "identifier", "qualifier": "other", "schema": "dc"}, {"key": "dc.identifier.uri", "value": "https://jyx.jyu.fi/handle/123456789/44199", "language": "", "element": "identifier", "qualifier": "uri", "schema": "dc"}, {"key": "dc.description.abstract", "value": "Echovirus 1 (EV1) kuuluu Pikornavirusperheeseen ja tarkemmin luokiteltuna Enterovirusten sukuun. EV1 on pieni ja vaipaton RNA-virus, jonka proteiinikuori koostuu nelj\u00e4st\u00e4 proteiinista VP1-VP4. Infektion on osoitettu alkavan, kun EV1 tarttuu \u03b12\u03b21-integriiniin solun pinnalla, jonka j\u00e4lkeen virus otetaan solun sis\u00e4\u00e4n kaveoliiniv\u00e4litteist\u00e4 endosytoosia sek\u00e4 makropinosytoosia muistuttavalla mekanismilla. Toisaalta sit\u00e4, kuinka virus avautuu sek\u00e4 vapauttaa RNA:nsa, ei viel\u00e4 tarkalleen tiedet\u00e4. T\u00e4ss\u00e4 tutkimuksessa EV1 puhdistettiin ensin sakkaroosigradientilla, jonka j\u00e4lkeen virus leimattiin fluoresoivalla amiinireaktiivisella v\u00e4rill\u00e4. Tulokset osoittivat, ett\u00e4 leimauksen onnistumiseksi tarvitaan suhteellisen v\u00e4h\u00e4inen leimam\u00e4\u00e4r\u00e4, ja ett\u00e4 v\u00e4h\u00e4inen leimam\u00e4\u00e4r\u00e4 riitt\u00e4\u00e4 fluoresenssin havaitsemiseksi sek\u00e4 on t\u00e4rke\u00e4 viruksen infektiivisyyden s\u00e4ilytt\u00e4miseksi. Leimattua EV1:st\u00e4 k\u00e4ytettiin solukokeissa, joissa Fosfolipaasi C inhibiittoria (PLC-inhibiittori) k\u00e4ytettiin viruksen endosytoosin est\u00e4miseen, jolloin asettamamme hypoteesin mukaan endosytoosia ei tapahdu, vaan virus j\u00e4\u00e4 solun pinnalle. Immunofluoresenssikokeiden ja kvantitatiivisen analysoinnin perusteella endosytoosi estyi, vaikkakin ero kontrollisoluihin oli ainoastaan 10 %. Lis\u00e4ksi toinen immunofluoresenssikoe osoitti, ett\u00e4 EV1:n infektiivisyys laski k\u00e4ytett\u00e4ess\u00e4 PLC-inhibiittoria. Immunofluoresenssikokeissa ongelmia aiheuttivat tyhj\u00e4t ja rikkin\u00e4iset viruspartikkelit, jotka havaittiin EV1:n karakterisoinnin aikana l\u00e4p\u00e4isyelektronimikroskoopilla. Leimautuneet tyhj\u00e4t ja rikkin\u00e4iset partikkelit aiheuttivat todenn\u00e4k\u00f6isesti v\u00e4\u00e4r\u00e4\u00e4 signaalia immunofluoresenssikokeissa, joiden perusteella voitiin todeta, ett\u00e4 viruksen puhdistaminen ja leimaaminen vaativat lis\u00e4\u00e4 optimointia. Suora kapsidileima voisi onnistuessaan toimia ty\u00f6kaluna tulevissa viruksen avautumiskokeissa ja livetutkimuksissa, jolloin vasta-aineleimauksiin liittyv\u00e4 taustafluoresenssi saataisiin minimoitua. Lis\u00e4ksi PLC-inhibiittori voisi mahdollistaa viruksen avautumisen tutkimisen solun pinnalla endosomaalisten rakenteiden sijaan.", "language": "fi", "element": "description", "qualifier": "abstract", "schema": "dc"}, {"key": "dc.description.abstract", "value": "Echovirus 1 (EV1) belongs to the family of Picornaviridae and more specifically to the genus Enterovirus. EV1 is a small, non-enveloped RNA-virus which has a capsid consisting of four proteins VP1-VP4. EV1 has been shown to bind to \u03b12\u03b21-integrin on the cell surface and then endocytosed inside the cell by a mechanism that resembles both caveolin-mediated endocytosis and macropinocytosis. Instead, uncoating and RNA release are poorly understood. Here, we have successfully purified EV1 with sucrose gradient and labeled the virus with an amine reactive dye. The results showed that for successful labeling, considerably low amount of dye is sufficient for signal detection, and according to end-point dilution assay, is necessary for maintaining virus infectivity. Capsid labeled EV1 was used in cellular studies where Phospholipase C-inhibitor (PLC-inhibitor) was tested as an endocytosis blocker. According to our hypothesis PLC-inhibitor blocks endocytosis, and as a result, virus remains on the cell surface. Indeed, based on immunofluorescence studies and quantification with Bioimage XD software, endocytosis was blocked, although difference to control cells was only 10%. Additionally, another immunofluorescence study showed that PLC-inhibitor decreased EV1 infection. Immunofluorescence studies were hindered by empty and disassembled virus particles, which were detected during EV1 characterization with transmission electron microcopy. Labeled empty and disassembled particles probably caused false signal in immunofluorescence studies, and thus, it was concluded that virus purification and labeling has to be more optimized for these kinds of studies. Fluorescent capsid label could provide us a tool for future uncoating and live imaging studies by minimizing non-specificity that is usually related to immunolabeling. In addition, PLC-inhibitor could enable uncoating studies to be carried out on the cell surface instead of endosomes.", "language": "en", "element": "description", "qualifier": "abstract", "schema": "dc"}, {"key": "dc.description.provenance", "value": "Submitted using Plone Publishing form by Mira Myllynen (mianmyll) on 2014-09-05 10:33:08.006952. Form: Pro gradu -lomake (https://kirjasto.jyu.fi/julkaisut/julkaisulomakkeet/pro-gradu-lomake). JyX data: [jyx_publishing-allowed (fi) =True]", "language": "en", "element": "description", "qualifier": "provenance", "schema": "dc"}, {"key": "dc.description.provenance", "value": "Submitted by jyx lomake-julkaisija (jyx-julkaisija@noreply.fi) on 2014-09-05T10:33:08Z\r\nNo. of bitstreams: 2\r\nURN:NBN:fi:jyu-201409052732.pdf: 1468849 bytes, checksum: 6f82d90c94ea9dba15601397f2a08371 (MD5)\r\nlicense.html: 4859 bytes, checksum: 5b8e1ee17a753161ea5475d57977bdf2 (MD5)", "language": "en", "element": "description", "qualifier": "provenance", "schema": "dc"}, {"key": "dc.description.provenance", "value": "Made available in DSpace on 2014-09-05T10:33:08Z (GMT). No. of bitstreams: 2\r\nURN:NBN:fi:jyu-201409052732.pdf: 1468849 bytes, checksum: 6f82d90c94ea9dba15601397f2a08371 (MD5)\r\nlicense.html: 4859 bytes, checksum: 5b8e1ee17a753161ea5475d57977bdf2 (MD5)\r\n Previous issue date: 2014", "language": "en", "element": "description", "qualifier": "provenance", "schema": "dc"}, {"key": "dc.format.extent", "value": "1 verkkoaineisto (46 sivua)", "language": null, "element": "format", "qualifier": "extent", "schema": "dc"}, {"key": "dc.format.mimetype", "value": "application/pdf", "language": null, "element": "format", "qualifier": "mimetype", "schema": "dc"}, {"key": "dc.language.iso", "value": "eng", "language": null, "element": "language", "qualifier": "iso", "schema": "dc"}, {"key": "dc.rights", "value": "In Copyright", "language": "en", "element": "rights", "qualifier": null, "schema": "dc"}, {"key": "dc.subject.other", "value": "Echovirus 1", "language": "", "element": "subject", "qualifier": "other", "schema": "dc"}, {"key": "dc.subject.other", "value": "uncoating", "language": "", "element": "subject", "qualifier": "other", "schema": "dc"}, {"key": "dc.subject.other", "value": "endocytosis inhibitor", "language": "", "element": "subject", "qualifier": "other", "schema": "dc"}, {"key": "dc.subject.other", "value": "fluorescent labeling", "language": "", "element": "subject", "qualifier": "other", "schema": "dc"}, {"key": "dc.title", "value": "The effect of PLC-inhibitor on echovirus 1 internalization probed with fluorescently labeled echovirus 1", "language": null, "element": "title", "qualifier": null, "schema": "dc"}, {"key": "dc.type", "value": "master thesis", "language": null, "element": "type", "qualifier": null, "schema": "dc"}, {"key": "dc.identifier.urn", "value": "URN:NBN:fi:jyu-201409052732", "language": null, "element": "identifier", "qualifier": "urn", "schema": "dc"}, {"key": "dc.type.ontasot", "value": "Pro gradu -tutkielma", "language": "fi", "element": "type", "qualifier": "ontasot", "schema": "dc"}, {"key": "dc.type.ontasot", "value": "Master\u2019s thesis", "language": "en", "element": "type", "qualifier": "ontasot", "schema": "dc"}, {"key": "dc.contributor.faculty", "value": "Matemaattis-luonnontieteellinen tiedekunta", "language": "fi", "element": "contributor", "qualifier": "faculty", "schema": "dc"}, {"key": "dc.contributor.faculty", "value": "Faculty of Sciences", "language": "en", "element": "contributor", "qualifier": "faculty", "schema": "dc"}, {"key": "dc.contributor.department", "value": "Bio- ja ymp\u00e4rist\u00f6tieteiden laitos", "language": "fi", "element": "contributor", "qualifier": "department", "schema": "dc"}, {"key": "dc.contributor.department", "value": "Department of Biological and Environmental Science", "language": "en", "element": "contributor", "qualifier": "department", "schema": "dc"}, {"key": "dc.contributor.organization", "value": "University of Jyv\u00e4skyl\u00e4", "language": "en", "element": "contributor", "qualifier": "organization", "schema": "dc"}, {"key": "dc.contributor.organization", "value": "Jyv\u00e4skyl\u00e4n yliopisto", "language": "fi", "element": "contributor", "qualifier": "organization", "schema": "dc"}, {"key": "dc.subject.discipline", "value": "Solu- ja molekyylibiologia", "language": "fi", "element": "subject", "qualifier": "discipline", "schema": "dc"}, {"key": "dc.subject.discipline", "value": "Cell and molecular biology", "language": "en", "element": "subject", "qualifier": "discipline", "schema": "dc"}, {"key": "dc.date.updated", "value": "2014-09-05T10:33:09Z", "language": "", "element": "date", "qualifier": "updated", "schema": "dc"}, {"key": "dc.type.coar", "value": "http://purl.org/coar/resource_type/c_bdcc", "language": null, "element": "type", "qualifier": "coar", "schema": "dc"}, {"key": "dc.rights.accesslevel", "value": "openAccess", "language": "fi", "element": "rights", "qualifier": "accesslevel", "schema": "dc"}, {"key": "dc.type.publication", "value": 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