{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,23]],"date-time":"2026-10-23T00:07:46Z","timestamp":1792714066273,"version":"4.5.0"},"reference-count":58,"publisher":"Microbiology Society","issue":"7","content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"published-print":{"date-parts":[[2014,7,1]]},"abstract":"<jats:p>Despite tremendous efforts to improve the methodology for constructing flavivirus infectious cDNAs, the manipulation of flavivirus cDNAs remains a difficult task in bacteria. Here, we successfully propagated DNA-launched type 2 dengue virus (DENV2) and Japanese encephalitis virus (JEV) infectious cDNAs by introducing seven repeats of the tetracycline-response element (7\u00d7TRE) and a minimal cytomegalovirus (CMVmin) promoter upstream of the viral genome. Insertion of the 7\u00d7TRE-CMVmin sequence upstream of the DENV2 or JEV genome decreased the cryptic<jats:italic>E. coli<\/jats:italic>promoter (ECP) activity of the viral genome in bacteria, as measured using fusion constructs containing DENV2 or JEV segments and the reporter gene<jats:italic>Renilla<\/jats:italic>luciferase in an empty vector. The growth kinetics of recombinant viruses derived from DNA-launched DENV2 and JEV infectious cDNAs were similar to those of parental viruses. Similarly, RNA-launched DENV2 infectious cDNAs were generated by inserting 7\u00d7TRE-CMVmin, five repeats of the GAL4 upstream activating sequence, or five repeats of<jats:italic>Bam<\/jats:italic>HI linkers upstream of the DENV2 genome. All three tandem repeat sequences decreased the ECP activity of the DENV2 genome in bacteria. Notably, 7\u00d7TRE-CMVmin stabilized RNA-launched JEV infectious cDNAs and reduced the ECP activity of the JEV genome in bacteria. The growth kinetics of recombinant viruses derived from RNA-launched DENV2 and JEV infectious cDNAs displayed patterns similar to those of the parental viruses. These results support a novel methodology for constructing flavivirus infectious cDNAs, which will facilitate research in virology, viral pathogenesis and vaccine development of flaviviruses and other RNA viruses.<\/jats:p>","DOI":"10.1099\/vir.0.064915-0","type":"journal-article","created":{"date-parts":[[2014,4,12]],"date-time":"2014-04-12T04:19:51Z","timestamp":1397276391000},"page":"1493-1503","source":"Crossref","is-referenced-by-count":32,"title":["A novel approach to propagate flavivirus infectious cDNA clones in bacteria by introducing tandem repeat sequences upstream of virus genome"],"prefix":"10.1099","volume":"95","author":[{"given":"Szu-Yuan","family":"Pu","sequence":"first","affiliation":[{"name":"Graduate Institute of Life Sciences, National Defense Medical Center, Taipei, Taiwan, R.O.C."},{"name":"Institute of Biotechnology and Pharmaceutical Research, National Health Research Institutes, Miaoli 350, Taiwan, R.O.C."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ren-Huang","family":"Wu","sequence":"additional","affiliation":[{"name":"Graduate Institute of Life Sciences, National Defense Medical Center, Taipei, Taiwan, R.O.C."},{"name":"Institute of Biotechnology and Pharmaceutical Research, National Health Research Institutes, Miaoli 350, Taiwan, R.O.C."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ming-Han","family":"Tsai","sequence":"additional","affiliation":[{"name":"Institute of Biotechnology and Pharmaceutical Research, National Health Research Institutes, Miaoli 350, Taiwan, R.O.C."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Chi-Chen","family":"Yang","sequence":"additional","affiliation":[{"name":"Graduate Institute of Life Sciences, National Defense Medical Center, Taipei, Taiwan, R.O.C."},{"name":"Institute of Biotechnology and Pharmaceutical Research, National Health Research Institutes, Miaoli 350, Taiwan, R.O.C."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Chung-Ming","family":"Chang","sequence":"additional","affiliation":[{"name":"Institute of Biotechnology and Pharmaceutical Research, National Health Research Institutes, Miaoli 350, Taiwan, R.O.C."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Andrew","family":"Yueh","sequence":"additional","affiliation":[{"name":"Graduate Institute of Life Sciences, National Defense Medical Center, Taipei, Taiwan, R.O.C."},{"name":"Institute of Biotechnology and Pharmaceutical Research, National Health Research Institutes, Miaoli 350, Taiwan, R.O.C."}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"345","reference":[{"key":"r1","doi-asserted-by":"publisher","DOI":"10.1038\/183799a0"},{"key":"r2","doi-asserted-by":"crossref","first-page":"17656","DOI":"10.1016\/S0021-9258(19)84621-5","article-title":"Eukaryotic promoters drive gene expression in Escherichia coli","volume":"264","author":"Antonucci","year":"1989","journal-title":"J Biol Chem"},{"key":"r3","doi-asserted-by":"publisher","DOI":"10.1016\/j.virol.2011.03.008"},{"key":"r4","doi-asserted-by":"publisher","DOI":"10.1128\/JVI.79.13.8339-8347.2005"},{"key":"r5","doi-asserted-by":"publisher","DOI":"10.1089\/vim.2006.19.10"},{"key":"r6","doi-asserted-by":"publisher","DOI":"10.1006\/viro.1994.1053"},{"key":"r7","doi-asserted-by":"publisher","DOI":"10.1099\/vir.0.18860-0"},{"key":"r8","doi-asserted-by":"publisher","DOI":"10.1007\/0-306-46825-5_19"},{"key":"r9","doi-asserted-by":"publisher","DOI":"10.1128\/JVI.79.22.13963-13973.2005"},{"key":"r10","doi-asserted-by":"publisher","DOI":"10.1146\/annurev.mi.44.100190.003245"},{"key":"r11","doi-asserted-by":"crossref","first-page":"6","DOI":"10.1111\/j.1470-8744.1988.tb00001.x","article-title":"Transfer and expression of plasmids containing human cytomegalovirus immediate-early gene 1 promoter-enhancer sequences in eukaryotic and prokaryotic cells","volume":"10","author":"Davis","year":"1988","journal-title":"Biotechnol Appl Biochem"},{"key":"r12","doi-asserted-by":"publisher","DOI":"10.1128\/JVI.03162-12"},{"key":"r13","doi-asserted-by":"crossref","first-page":"2467","DOI":"10.1128\/JVI.65.5.2467-2475.1991","article-title":"Both nonstructural proteins NS2B and NS3 are required for the proteolytic processing of dengue virus nonstructural proteins","volume":"65","author":"Falgout","year":"1991","journal-title":"J Virol"},{"key":"r14","doi-asserted-by":"publisher","DOI":"10.1146\/annurev-genet-072610-155046"},{"key":"r15","doi-asserted-by":"crossref","first-page":"571","DOI":"10.4269\/ajtmh.1989.40.571","article-title":"Aedes aegypti and Aedes aegypti-borne disease control in the 1990s: top down or bottom up. 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