Analysis of DNA exchange using Thymidine analogs (ADExTA) in Trypanosoma cruzi
Full metadata record
DC Field | Value | Language |
---|---|---|
dc.contributor | (CeTICS) Centro de Toxinas, Resposta-imune e Sinalização Celular | pt_BR |
dc.contributor | (LCC) Lab. Ciclo Celular | pt_BR |
dc.contributor.author | Silva, Marcelo Santos da | pt_BR |
dc.contributor.author | Marin, Paula Andrea | pt_BR |
dc.contributor.author | Elias, Maria Carolina | pt_BR |
dc.contributor.author | Repolês, Bruno M. | pt_BR |
dc.contributor.author | Machado, Carlos R. | pt_BR |
dc.date.accessioned | 2020-07-09T21:24:06Z | - |
dc.date.available | 2020-07-09T21:24:06Z | - |
dc.date.issued | 2018 | pt_BR |
dc.identifier.citation | Silva 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.uri | https://repositorio.butantan.gov.br/handle/butantan/2757 | - |
dc.description.abstract | Trypanosoma 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.sponsorship | (FAPESP) Fundação de Amparo à Pesquisa do Estado de São Paulo | pt_BR |
dc.description.sponsorship | (CNPq) Conselho Nacional de Desenvolvimento Científico e Tecnológico | pt_BR |
dc.description.sponsorship | (FAPEMIG) Fundação de Amparo à Pesquisa do Estado de Minas Gerais | pt_BR |
dc.description.sponsorship | (CAPES) Coordenação de Aperfeiçoamento de Pessoal de Nível Superior | pt_BR |
dc.format.extent | e3125 | pt_BR |
dc.language.iso | English | pt_BR |
dc.relation.ispartof | Bio-Protocol | pt_BR |
dc.rights | Restricted access | pt_BR |
dc.title | Analysis of DNA exchange using Thymidine analogs (ADExTA) in Trypanosoma cruzi | pt_BR |
dc.type | Article | pt_BR |
dc.identifier.doi | 10.21769/BioProtoc.3125 | pt_BR |
dc.identifier.url | https://doi.org/10.21769/BioProtoc.3125 | pt_BR |
dc.contributor.external | (UFMG) Universidade Federal de Minas Gerais | pt_BR |
dc.identifier.citationvolume | 8 | pt_BR |
dc.identifier.citationissue | 24 | pt_BR |
dc.subject.keyword | DNA exchange | pt_BR |
dc.subject.keyword | Genetic exchange | pt_BR |
dc.subject.keyword | Thymidine analogs | pt_BR |
dc.subject.keyword | CldU | pt_BR |
dc.subject.keyword | IdU | pt_BR |
dc.subject.keyword | DNA replication | pt_BR |
dc.subject.keyword | Trypanosomatids | pt_BR |
dc.subject.keyword | Trypanosoma cruzi | pt_BR |
dc.relation.ispartofabbreviated | Bio Protoc | pt_BR |
dc.identifier.citationabnt | v. 8, n. 24, e3125, dez. 2018 | pt_BR |
dc.identifier.citationvancouver | 2018 Dec;8(24):e3125 | pt_BR |
dc.contributor.butantan | Marin, Paula Andrea|:Aluno|:LCC - Laboratório de Ciclo Celular:Centro de Toxinas, Resposta-imune e Sinalização Celular (CeTICS)|: | pt_BR |
dc.contributor.butantan | Elias, Maria Carolina|:Pesquisador|:LCC - Laboratório de Ciclo Celular:Centro de Toxinas, Resposta-imune e Sinalização Celular (CeTICS)|:Autor de correspondência | pt_BR |
dc.contributor.butantan | Silva, Marcelo Santos da|:Aluno|:Centro de Toxinas, Resposta-imune e Sinalização Celular (CeTICS)|:PrimeiroAutor:Autor de correspondência | pt_BR |
dc.sponsorship.butantan | (CNPq) Conselho Nacional de Desenvolvimento Científico e Tecnológico¦¦870219/1997-9 | pt_BR |
dc.sponsorship.butantan | (CNPq) Conselho Nacional de Desenvolvimento Científico e Tecnológico¦¦304329/2015-0 | pt_BR |
dc.sponsorship.butantan | (CAPES) Coordenação de Aperfeiçoamento de Pessoal de Nível Superior¦¦ | pt_BR |
dc.sponsorship.butantan | Fundação de Amparo à Pesquisa do Estado de Minas Gerais (FAPEMIG)¦¦ | pt_BR |
dc.sponsorship.butantan | (FAPESP) Fundação de Amparo à Pesquisa do Estado de São Paulo¦¦2013/07467-1 | pt_BR |
dc.sponsorship.butantan | (FAPESP) Fundação de Amparo à Pesquisa do Estado de São Paulo¦¦2014/24170-5 | pt_BR |
dc.sponsorship.butantan | (FAPESP) Fundação de Amparo à Pesquisa do Estado de São Paulo¦¦2017/18719-2 | pt_BR |
dc.identifier.bvscc | BR78.1 | pt_BR |
dc.identifier.bvsdb | IBProd | pt_BR |
dc.description.dbindexed | Yes | pt_BR |
item.fulltext | Sem Texto completo | - |
item.openairetype | Article | - |
item.languageiso639-1 | English | - |
item.grantfulltext | none | - |
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