{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,29]],"date-time":"2026-09-29T23:27:39Z","timestamp":1790724459807,"version":"4.1.0"},"reference-count":29,"publisher":"Springer Science and Business Media LLC","issue":"1","license":[{"start":{"date-parts":[[2017,10,25]],"date-time":"2017-10-25T00:00:00Z","timestamp":1508889600000},"content-version":"tdm","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0"},{"start":{"date-parts":[[2017,10,25]],"date-time":"2017-10-25T00:00:00Z","timestamp":1508889600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0"}],"content-domain":{"domain":["link.springer.com"],"crossmark-restriction":false},"short-container-title":["Sci Rep"],"abstract":"<jats:title>Abstract<\/jats:title><jats:p>Reverse genetics is a critical tool to decrypt the biological properties of arboviruses. However, whilst reverse genetics methods have been usually applied to vertebrate cells, their use in insect cells remains uncommon due to the conjunction of laborious molecular biology techniques and of specific difficulties surrounding the transfection of such cells. To leverage reverse genetics studies in both vertebrate and mosquito cells, we designed an improved DNA transfection protocol for insect cells and then demonstrated that the simple and flexible ISA (Infectious Subgenomic Amplicons) reverse-genetics method can be efficiently applied to both mammalian and mosquito cells to generate in days recombinant infectious positive-stranded RNA viruses belonging to genera <jats:italic>Flavivirus<\/jats:italic> (Japanese encephalitis, Yellow fever, West Nile and Zika viruses) and <jats:italic>Alphavirus<\/jats:italic> (Chikungunya virus). This method represents an effective option to potentially overcome technological issues related to the study of arboviruses.<\/jats:p>","DOI":"10.1038\/s41598-017-14522-6","type":"journal-article","created":{"date-parts":[[2017,10,19]],"date-time":"2017-10-19T09:09:58Z","timestamp":1508404198000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":34,"title":["New reverse genetics and transfection methods to rescue arboviruses in mosquito cells"],"prefix":"10.1038","volume":"7","author":[{"given":"Th\u00e9r\u00e8se","family":"Atieh","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Antoine","family":"Nougair\u00e8de","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-7777-0308","authenticated-orcid":false,"given":"Rapha\u00eblle","family":"Klitting","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Fabien","family":"Aubry","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Anna-Bella","family":"Failloux","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7895-2720","authenticated-orcid":false,"given":"Xavier","family":"de Lamballerie","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7102-3654","authenticated-orcid":false,"given":"St\u00e9phane","family":"Priet","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"297","published-online":{"date-parts":[[2017,10,25]]},"reference":[{"key":"14522_CR1","doi-asserted-by":"publisher","first-page":"330","DOI":"10.1016\/S0188-4409(02)00378-8","volume":"33","author":"DJ Gubler","year":"2002","unstructured":"Gubler, D. J. The global emergence\/resurgence of arboviral diseases as public health problems. Arch Med Res \n                           33, 330\u2013342 (2002).","journal-title":"Arch Med Res"},{"key":"14522_CR2","doi-asserted-by":"publisher","first-page":"328","DOI":"10.1016\/j.antiviral.2009.10.008","volume":"85","author":"SC Weaver","year":"2010","unstructured":"Weaver, S. C. & Reisen, W. K. Present and future arboviral threats. Antiviral Res \n                           85, 328\u2013345, https:\/\/doi.org\/10.1016\/j.antiviral.2009.10.008 (2010).","journal-title":"Antiviral Res"},{"key":"14522_CR3","doi-asserted-by":"publisher","first-page":"360","DOI":"10.1016\/j.tim.2013.03.003","volume":"21","author":"SC Weaver","year":"2013","unstructured":"Weaver, S. C. Urbanization and geographic expansion of zoonotic arboviral diseases: mechanisms and potential strategies for prevention. Trends Microbiol \n                           21, 360\u2013363, https:\/\/doi.org\/10.1016\/j.tim.2013.03.003 (2013).","journal-title":"Trends Microbiol"},{"key":"14522_CR4","doi-asserted-by":"publisher","first-page":"67","DOI":"10.1016\/j.antiviral.2014.12.007","volume":"114","author":"F Aubry","year":"2015","unstructured":"Aubry, F., Nougairede, A., Gould, E. A. & de Lamballerie, X. Flavivirus reverse genetic systems, construction techniques and applications: a historical perspective. Antiviral Res \n                           114, 67\u201385, https:\/\/doi.org\/10.1016\/j.antiviral.2014.12.007 (2015).","journal-title":"Antiviral Res"},{"key":"14522_CR5","doi-asserted-by":"publisher","first-page":"2531","DOI":"10.3390\/v6072531","volume":"6","author":"CC Stobart","year":"2014","unstructured":"Stobart, C. C. & Moore, M. L. RNA virus reverse genetics and vaccine design. Viruses \n                           6, 2531\u20132550, https:\/\/doi.org\/10.3390\/v6072531 (2014).","journal-title":"Viruses"},{"key":"14522_CR6","doi-asserted-by":"publisher","first-page":"e08347","DOI":"10.7554\/eLife.08347","volume":"4","author":"MU Kraemer","year":"2015","unstructured":"Kraemer, M. U. et al. The global distribution of the arbovirus vectors Aedes aegypti and Ae. albopictus. Elife \n                           4, e08347, https:\/\/doi.org\/10.7554\/eLife.08347 (2015).","journal-title":"Elife"},{"key":"14522_CR7","doi-asserted-by":"publisher","first-page":"817","DOI":"10.1016\/j.vaccine.2007.11.082","volume":"26","author":"JE Blaney Jr","year":"2008","unstructured":"Blaney, J. E. Jr et al. Dengue virus type 3 vaccine candidates generated by introduction of deletions in the 3\u2032 untranslated region (3\u2032-UTR) or by exchange of the DENV-3 3\u2032-UTR with that of DENV-4. Vaccine \n                           26, 817\u2013828, https:\/\/doi.org\/10.1016\/j.vaccine.2007.11.082 (2008).","journal-title":"Vaccine"},{"key":"14522_CR8","doi-asserted-by":"publisher","first-page":"3010","DOI":"10.1016\/j.vaccine.2014.03.062","volume":"32","author":"GD Gromowski","year":"2014","unstructured":"Gromowski, G. D., Firestone, C. Y., Hanson, C. T. & Whitehead, S. S. Japanese encephalitis virus vaccine candidates generated by chimerization with dengue virus type 4. Vaccine \n                           32, 3010\u20133018, https:\/\/doi.org\/10.1016\/j.vaccine.2014.03.062 (2014).","journal-title":"Vaccine"},{"key":"14522_CR9","doi-asserted-by":"publisher","unstructured":"Junglen, S. et al. Host Range Restriction of Insect-Specific Flaviviruses Occurs at Several Levels of the Viral Life Cycle. mSphere \n                           2, doi:https:\/\/doi.org\/10.1128\/mSphere.00375-16 (2017).","DOI":"10.1128\/mSphere.00375-16"},{"key":"14522_CR10","doi-asserted-by":"publisher","first-page":"153","DOI":"10.1016\/j.virusres.2014.10.010","volume":"195","author":"T Ishikawa","year":"2015","unstructured":"Ishikawa, T., Abe, M. & Masuda, M. Construction of an infectious molecular clone of Japanese encephalitis virus genotype V and its derivative subgenomic replicon capable of expressing a foreign gene. Virus Res \n                           195, 153\u2013161, https:\/\/doi.org\/10.1016\/j.virusres.2014.10.010 (2015).","journal-title":"Virus Res"},{"key":"14522_CR11","doi-asserted-by":"publisher","first-page":"139","DOI":"10.1016\/j.virusres.2006.10.013","volume":"124","author":"KN Tsai","year":"2007","unstructured":"Tsai, K. N., Tsang, S. F., Huang, C. H. & Chang, R. Y. Defective interfering RNAs of Japanese encephalitis virus found in mosquito cells and correlation with persistent infection. Virus Res \n                           124, 139\u2013150, https:\/\/doi.org\/10.1016\/j.virusres.2006.10.013 (2007).","journal-title":"Virus Res"},{"key":"14522_CR12","doi-asserted-by":"publisher","first-page":"2457","DOI":"10.1099\/vir.0.011486-0","volume":"90","author":"H Kroschewski","year":"2009","unstructured":"Kroschewski, H., Sagripanti, J. L. & Davidson, A. D. Identification of amino acids in the dengue virus type 2 envelope glycoprotein critical to virus infectivity. J Gen Virol \n                           90, 2457\u20132461, https:\/\/doi.org\/10.1099\/vir.0.011486-0 (2009).","journal-title":"J Gen Virol"},{"key":"14522_CR13","doi-asserted-by":"publisher","first-page":"511","DOI":"10.1016\/j.virol.2012.06.028","volume":"432","author":"AM Groat-Carmona","year":"2012","unstructured":"Groat-Carmona, A. M. et al. 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