Analysis of DNA Exchange Using Thymidine Analogs (ADExTA) in Trypanosoma cruzi

Full metadata record
DC FieldValueLanguage
dc.contributorCentro de Toxinas, Resposta-imune e Sinalização Celular (CeTICS)pt_BR
dc.contributorLCC - Laboratório de Ciclo Celularpt_BR
dc.contributor.authorSilva, Marcelo Santos dapt_BR
dc.contributor.authorMarin, Paula Andreapt_BR
dc.contributor.authorElias, Maria Carolinapt_BR
dc.contributor.authorRepolês, Bruno M.pt_BR
dc.contributor.authorMachado, Carlos R.pt_BR
dc.date.accessioned2020-07-09T21:24:06Z-
dc.date.available2020-07-09T21:24:06Z-
dc.date.issued2018pt_BR
dc.identifier.citationSilva MS, Marin PA, Elias MC, Repolês BM., Machado CR.. Analysis of DNA Exchange Using Thymidine Analogs (ADExTA) in Trypanosoma cruzi. Bio Protoc.. 2018 Dec;8(24):e3125. doi:10.21769/BioProtoc.3125.pt_BR
dc.identifier.urihttps://repositorio.butantan.gov.br/handle/butantan/2757-
dc.description.abstractTrypanosoma cruzi is a protozoan parasite belonging to the Trypanosomatidae family. Although the trypanosomatids multiply predominantly by clonal generation, the presence of DNA exchange in some of them has been puzzling researchers over the years, mainly because it may represent a novel form that these organisms use to gain variability. Analysis of DNA Exchange using Thymidine Analogs (ADExTA) is a method that allows the in vitro detection and measurement of rates of DNA exchange, particularly in trypanosomatid cells, in a rapid and simple manner by indirect immunofluorescence assay (IFA). The method can be used to detect DNA exchange within one trypanosomatid lineage or among different lineages by paired analysis. The principle of this assay is based on the incorporation of two distinguishable halogenated thymidine analogs called 5'-chloro-2'-deoxyuridine (CldU) and 5'-iodo-2'-deoxyuridine (IdU) during DNA replication. After mixing the two cell cultures that had been previously incorporated with CldU and IdU separately, the presence of these unusual deoxynucleosides in the genome can be detected by specific antibodies. For this, a DNA denaturation step is required to expose the sites of thymidine analogs incorporated. Subsequently, a secondary reaction using fluorochrome-labeled antibodies will generate distinct signals under fluorescence analysis. By using this method, DNA exchange verification (i.e., the presence of both CldU and IdU in the same cell) is possible using a standard fluorescence microscope. It typically takes 2-3 days from the thymidine analogs incorporation to results. Of note, ADExTA is relatively cheap and does not require transfections or harsh genetic manipulation. These features represent an advantage when compared to other time-consuming protocols that demand DNA manipulation to introduce distinct drug-resistance markers in different cells for posterior selection.pt_BR
dc.description.sponsorshipFundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)pt_BR
dc.description.sponsorshipConselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)pt_BR
dc.description.sponsorshipFundação de Amparo à Pesquisa do Estado de Minas Gerais (FAPEMIG)pt_BR
dc.description.sponsorshipCoordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)pt_BR
dc.format.extente3125pt_BR
dc.language.isoEnglishpt_BR
dc.relation.ispartofBio-Protocolpt_BR
dc.titleAnalysis of DNA Exchange Using Thymidine Analogs (ADExTA) in Trypanosoma cruzipt_BR
dc.typeArticlept_BR
dc.identifier.doi10.21769/BioProtoc.3125pt_BR
dc.identifier.urlhttps://doi.org/10.21769/BioProtoc.3125pt_BR
dc.contributor.externalUniversidade Federal de Minas Gerais (UFMG)¦¦Brasilpt_BR
dc.identifier.citationvolume8pt_BR
dc.identifier.citationissue24pt_BR
dc.subject.keywordDNA exchangept_BR
dc.subject.keywordGenetic exchangept_BR
dc.subject.keywordThymidine analogspt_BR
dc.subject.keywordCldUpt_BR
dc.subject.keywordIdUpt_BR
dc.subject.keywordDNA replicationpt_BR
dc.subject.keywordTrypanosomatidspt_BR
dc.subject.keywordTrypanosoma cruzipt_BR
dc.relation.ispartofabbreviatedBio Protocpt_BR
dc.identifier.citationabntv. 8, n. 24, e3125, dez. 2018pt_BR
dc.identifier.citationvancouver2018 Dec;8(24):e3125pt_BR
dc.contributor.butantanMarin, Paula Andrea|:Aluno|:LCC - Laboratório de Ciclo Celular:Centro de Toxinas, Resposta-imune e Sinalização Celular (CeTICS)|:pt_BR
dc.contributor.butantanElias, Maria Carolina|:Pesquisador|:LCC - Laboratório de Ciclo Celular:Centro de Toxinas, Resposta-imune e Sinalização Celular (CeTICS)|:Autor de correspondênciapt_BR
dc.contributor.butantanSilva, Marcelo Santos da|:Aluno|:Centro de Toxinas, Resposta-imune e Sinalização Celular (CeTICS)|:PrimeiroAutor:Autor de correspondênciapt_BR
dc.sponsorship.butantanConselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)¦¦870219/1997-9pt_BR
dc.sponsorship.butantanConselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)¦¦304329/2015-0pt_BR
dc.sponsorship.butantanCoordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)¦¦pt_BR
dc.sponsorship.butantanFundação de Amparo à Pesquisa do Estado de Minas Gerais (FAPEMIG)¦¦pt_BR
dc.sponsorship.butantanFundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)¦¦2013/07467-1pt_BR
dc.sponsorship.butantanFundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)¦¦2014/24170-5pt_BR
dc.sponsorship.butantanFundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)¦¦2017/18719-2pt_BR
dc.identifier.bvsccBR78.1pt_BR
dc.identifier.bvsdbIBProdpt_BR
dc.description.dbindexedYespt_BR
item.openairetypeArticle-
item.fulltextCom Texto completo-
item.grantfulltextopen-
item.languageiso639-1English-
crisitem.author.dept#PLACEHOLDER_PARENT_METADATA_VALUE#-
crisitem.author.dept#PLACEHOLDER_PARENT_METADATA_VALUE#-
crisitem.author.dept#PLACEHOLDER_PARENT_METADATA_VALUE#-
crisitem.author.dept#PLACEHOLDER_PARENT_METADATA_VALUE#-
crisitem.author.dept#PLACEHOLDER_PARENT_METADATA_VALUE#-
crisitem.author.orcid#PLACEHOLDER_PARENT_METADATA_VALUE#-
crisitem.author.orcid#PLACEHOLDER_PARENT_METADATA_VALUE#-
crisitem.author.orcid#PLACEHOLDER_PARENT_METADATA_VALUE#-
crisitem.author.orcid#PLACEHOLDER_PARENT_METADATA_VALUE#-
crisitem.author.orcid#PLACEHOLDER_PARENT_METADATA_VALUE#-
crisitem.author.parentorg#PLACEHOLDER_PARENT_METADATA_VALUE#-
crisitem.author.parentorg#PLACEHOLDER_PARENT_METADATA_VALUE#-
crisitem.journal.journalissn#PLACEHOLDER_PARENT_METADATA_VALUE#-
crisitem.journal.journaleissn#PLACEHOLDER_PARENT_METADATA_VALUE#-
Appears in Collections:Artigos de periódicos


Files in This Item:

10.21769BioProtoc.3125.pdf
Size: 2.31 MB
Format: Adobe PDF
View/Open
Show simple item record

The access to the publications deposited in this repository respects the licenses from journals and publishers.