title: Infectious clone construction of dengue virus type 2, strain Jamaican 1409, and characterization of a conditional E6 mutation.
abstract: A full-length infectious cDNA clone (ic) was constructed from the genome of the dengue virus type 2 (DENV-2) Jamaica83 1409 strain, pBAC1409ic, by using a bacterial artifical chromosome plasmid system. Infectious virus was generated and characterized for growth in cell culture and for infection in Aedes aegypti mosquitoes. During construction, an isoleucine to methionine (Ile-->Met) change was found at position 6 in the envelope glycoprotein sequence between low- and high-passage DENV-2 1409 strains. In vitro-transcribed genomic RNA of 1409ic with E6-Ile produced infectious virions following electroporation in mosquito cells, but not mammalian cells, while 1409ic RNA with an E6-Met mutation produced virus in both cell types. Moreover, DENV-2 1409 with the E6-Ile residue produced syncytia in C6/36 cell culture, whereas viruses with E6-Met did not. However, in vitro cell culture-derived growth-curve data and in vivo mosquito-infection rates revealed that none of the analysed DENV-2 strains differed from each other.
authors.affiliation: Arthropod-borne and Infectious Disease Laboratory, Department of Microbiology, Immunology and Pathology, Colorado State University, Foothills Research Campus, Fort Collins, CO 80523, USA.
authors.forename: Dennis J
authors.initials: DJ
authors.lastname: Pierro
authors.affiliation: Arthropod-borne and Infectious Disease Laboratory, Department of Microbiology, Immunology and Pathology, Colorado State University, Foothills Research Campus, Fort Collins, CO 80523, USA.
authors.forename: Ma Isabel
authors.initials: MI
authors.lastname: Salazar
authors.affiliation: Arthropod-borne and Infectious Disease Laboratory, Department of Microbiology, Immunology and Pathology, Colorado State University, Foothills Research Campus, Fort Collins, CO 80523, USA.
authors.forename: Barry J
authors.initials: BJ
authors.lastname: Beaty
authors.affiliation: Arthropod-borne and Infectious Disease Laboratory, Department of Microbiology, Immunology and Pathology, Colorado State University, Foothills Research Campus, Fort Collins, CO 80523, USA.
authors.forename: Ken E
authors.initials: KE
authors.lastname: Olson
mesh_terms: D000330:Aedes / Q000821:virology; D019943:Amino Acid Substitution; D000818:Animals; D002460:Cell Line; D022202:Chromosomes, Artificial, Bacterial; D003001:Cloning, Molecular; D018076:DNA, Complementary; D004279:DNA, Viral / Q000235:genetics; D003716:Dengue Virus / Q000235:genetics* / Q000502:physiology*; D016679:Genome, Viral; D015726:Giant Cells / Q000821:virology; D008253:Macaca mulatta; D009154:Mutation; D012367:RNA, Viral; D017422:Sequence Analysis, DNA; D014162:Transfection; D014759:Viral Envelope Proteins / Q000235:genetics*
chemical_list: D018076:DNA, Complementary; D004279:DNA, Viral; C051821:E-glycoprotein, Dengue virus type 2; D012367:RNA, Viral; D014759:Viral Envelope Proteins