A two-plasmid strategy for engineering a dengue virus type 3 infectious clone from primary Brazilian isolate

Detalhes bibliográficos
Autor(a) principal: SANTOS,JEFFERSON J.S.
Data de Publicação: 2014
Outros Autores: CORDEIRO,MARLI T., BERTANI,GIOVANI R., MARQUES,ERNESTO T.A., GIL,LAURA H.V.G.
Tipo de documento: Artigo
Idioma: eng
Título da fonte: Anais da Academia Brasileira de Ciências (Online)
Texto Completo: http://old.scielo.br/scielo.php?script=sci_arttext&pid=S0001-37652014000401749
Resumo: Dengue infections represent one of the most prevalent arthropod-borne diseases worldwide, causing a wide spectrum of clinical outcomes. Engineered infectious clone is an important tool to study Dengue virus (DENV) biology. Functional full-length cDNA clones have been constructed for many positive-strand RNA viruses and have provided valuable tools for studying the molecular mechanisms involved in viral genome replication, virion assembly, virus pathogenesis and vaccine development. We report herein the successful development of an infectious clone from a primary Brazilian isolate of dengue virus 3 (DENV3) of the genotype III. Using a two-plasmid strategy, DENV3 genome was divided in two parts and cloned separately into a yeast-bacteria shuttle vector. All plasmids were assembled in yeast by homologous recombination technique and a full-length template for transcription was obtained by in vitro ligation of the two parts of the genome. Transcript-derived DENV3 is infectious upon transfection into BHK-21 cells and in vitro characterization confirmed its identity. Growth kinetics of transcript-derived DENV3 was indistinguishable from wild type DENV3. This system is a powerful tool that will help shed light on molecular features of DENV biology, as the relationship of specific mutations and DENV pathogenesis.
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spelling A two-plasmid strategy for engineering a dengue virus type 3 infectious clone from primary Brazilian isolatereverse geneticsdengue virusmolecular cloninginfectious cloneDengue infections represent one of the most prevalent arthropod-borne diseases worldwide, causing a wide spectrum of clinical outcomes. Engineered infectious clone is an important tool to study Dengue virus (DENV) biology. Functional full-length cDNA clones have been constructed for many positive-strand RNA viruses and have provided valuable tools for studying the molecular mechanisms involved in viral genome replication, virion assembly, virus pathogenesis and vaccine development. We report herein the successful development of an infectious clone from a primary Brazilian isolate of dengue virus 3 (DENV3) of the genotype III. Using a two-plasmid strategy, DENV3 genome was divided in two parts and cloned separately into a yeast-bacteria shuttle vector. All plasmids were assembled in yeast by homologous recombination technique and a full-length template for transcription was obtained by in vitro ligation of the two parts of the genome. Transcript-derived DENV3 is infectious upon transfection into BHK-21 cells and in vitro characterization confirmed its identity. Growth kinetics of transcript-derived DENV3 was indistinguishable from wild type DENV3. This system is a powerful tool that will help shed light on molecular features of DENV biology, as the relationship of specific mutations and DENV pathogenesis.Academia Brasileira de Ciências2014-12-01info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersiontext/htmlhttp://old.scielo.br/scielo.php?script=sci_arttext&pid=S0001-37652014000401749Anais da Academia Brasileira de Ciências v.86 n.4 2014reponame:Anais da Academia Brasileira de Ciências (Online)instname:Academia Brasileira de Ciências (ABC)instacron:ABC10.1590/0001-3765201420130332info:eu-repo/semantics/openAccessSANTOS,JEFFERSON J.S.CORDEIRO,MARLI T.BERTANI,GIOVANI R.MARQUES,ERNESTO T.A.GIL,LAURA H.V.G.eng2015-10-27T00:00:00Zoai:scielo:S0001-37652014000401749Revistahttp://www.scielo.br/aabchttps://old.scielo.br/oai/scielo-oai.php||aabc@abc.org.br1678-26900001-3765opendoar:2015-10-27T00:00Anais da Academia Brasileira de Ciências (Online) - Academia Brasileira de Ciências (ABC)false
dc.title.none.fl_str_mv A two-plasmid strategy for engineering a dengue virus type 3 infectious clone from primary Brazilian isolate
title A two-plasmid strategy for engineering a dengue virus type 3 infectious clone from primary Brazilian isolate
spellingShingle A two-plasmid strategy for engineering a dengue virus type 3 infectious clone from primary Brazilian isolate
SANTOS,JEFFERSON J.S.
reverse genetics
dengue virus
molecular cloning
infectious clone
title_short A two-plasmid strategy for engineering a dengue virus type 3 infectious clone from primary Brazilian isolate
title_full A two-plasmid strategy for engineering a dengue virus type 3 infectious clone from primary Brazilian isolate
title_fullStr A two-plasmid strategy for engineering a dengue virus type 3 infectious clone from primary Brazilian isolate
title_full_unstemmed A two-plasmid strategy for engineering a dengue virus type 3 infectious clone from primary Brazilian isolate
title_sort A two-plasmid strategy for engineering a dengue virus type 3 infectious clone from primary Brazilian isolate
author SANTOS,JEFFERSON J.S.
author_facet SANTOS,JEFFERSON J.S.
CORDEIRO,MARLI T.
BERTANI,GIOVANI R.
MARQUES,ERNESTO T.A.
GIL,LAURA H.V.G.
author_role author
author2 CORDEIRO,MARLI T.
BERTANI,GIOVANI R.
MARQUES,ERNESTO T.A.
GIL,LAURA H.V.G.
author2_role author
author
author
author
dc.contributor.author.fl_str_mv SANTOS,JEFFERSON J.S.
CORDEIRO,MARLI T.
BERTANI,GIOVANI R.
MARQUES,ERNESTO T.A.
GIL,LAURA H.V.G.
dc.subject.por.fl_str_mv reverse genetics
dengue virus
molecular cloning
infectious clone
topic reverse genetics
dengue virus
molecular cloning
infectious clone
description Dengue infections represent one of the most prevalent arthropod-borne diseases worldwide, causing a wide spectrum of clinical outcomes. Engineered infectious clone is an important tool to study Dengue virus (DENV) biology. Functional full-length cDNA clones have been constructed for many positive-strand RNA viruses and have provided valuable tools for studying the molecular mechanisms involved in viral genome replication, virion assembly, virus pathogenesis and vaccine development. We report herein the successful development of an infectious clone from a primary Brazilian isolate of dengue virus 3 (DENV3) of the genotype III. Using a two-plasmid strategy, DENV3 genome was divided in two parts and cloned separately into a yeast-bacteria shuttle vector. All plasmids were assembled in yeast by homologous recombination technique and a full-length template for transcription was obtained by in vitro ligation of the two parts of the genome. Transcript-derived DENV3 is infectious upon transfection into BHK-21 cells and in vitro characterization confirmed its identity. Growth kinetics of transcript-derived DENV3 was indistinguishable from wild type DENV3. This system is a powerful tool that will help shed light on molecular features of DENV biology, as the relationship of specific mutations and DENV pathogenesis.
publishDate 2014
dc.date.none.fl_str_mv 2014-12-01
dc.type.driver.fl_str_mv info:eu-repo/semantics/article
dc.type.status.fl_str_mv info:eu-repo/semantics/publishedVersion
format article
status_str publishedVersion
dc.identifier.uri.fl_str_mv http://old.scielo.br/scielo.php?script=sci_arttext&pid=S0001-37652014000401749
url http://old.scielo.br/scielo.php?script=sci_arttext&pid=S0001-37652014000401749
dc.language.iso.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv 10.1590/0001-3765201420130332
dc.rights.driver.fl_str_mv info:eu-repo/semantics/openAccess
eu_rights_str_mv openAccess
dc.format.none.fl_str_mv text/html
dc.publisher.none.fl_str_mv Academia Brasileira de Ciências
publisher.none.fl_str_mv Academia Brasileira de Ciências
dc.source.none.fl_str_mv Anais da Academia Brasileira de Ciências v.86 n.4 2014
reponame:Anais da Academia Brasileira de Ciências (Online)
instname:Academia Brasileira de Ciências (ABC)
instacron:ABC
instname_str Academia Brasileira de Ciências (ABC)
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institution ABC
reponame_str Anais da Academia Brasileira de Ciências (Online)
collection Anais da Academia Brasileira de Ciências (Online)
repository.name.fl_str_mv Anais da Academia Brasileira de Ciências (Online) - Academia Brasileira de Ciências (ABC)
repository.mail.fl_str_mv ||aabc@abc.org.br
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