Data/hora: 03/06/2024 19:00:32

Provedor de dados: | 42
País: | Brazil
Título: | Isoenzyme and molecular approach for authenticating and monitoring of animal cell lines
Autores: | ARAÚJO,SHIRLEY B. DE; PATRICIO,GIANCARLO F.; SIMONI,ISABELA C.; RIVAS,ELIANA B.; FERNANDES,MARIA JUDITE B.
Data: | 2019-01-01
Ano: | 2019
Palavras-chave: | Authentication; Cell line; DNA barcode; Isoenzyme.
Resumo: | Abstract: Authentication of cell lines is of paramount importance to validate the results from their use in biomedical research. Although isoenzyme polymorphism is the standard method, molecular methods based on mitochondrial DNA (mtDNA) have been developed to replace it. The aim of this study was the improvement of our isoenzyme electrophoretic analysis and the validation of one molecular technique targeted at mtDNA for the authentication of our animal cell lines. The combined method of cellular lysing through osmotic shock, followed by freezing-thawing in N2 to obtain isoenzyme extracts, and with 42 × 106 cells maintained the best efficiency. The superior electrophoretic conditions were PAGE run at 200 V. All cell lines had isoenzymatic mobility corresponding to their species to lactate dehydrogenase, malate-dehydrogenase, and glucose-6-phosphate dehydrogenase isoenzymes, and could be distinguished from each other. Two molecular techniques based on mtDNA were tested, one on the cytochrome b gene and other on cytochrome c oxidase I subunit gene. Due to difficulties in distinguishing all cell lines using only one these techniques, we merged the primers of two methods in such a way that there was a sufficient differentiation of all DNA fragments. The sequencing of these PCR products was also performed to validate these data.
Tipo: | Info:eu-repo/semantics/article
Idioma: | Inglês
Identificador: | http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0001-37652019000500625
Editor: | Academia Brasileira de Ciências
Relação: | 10.1590/0001-3765201920180487
Formato: | text/html
Fonte: | Anais da Academia Brasileira de Ciências  v.91 n.3 2019
Fonte: | info:eu-repo/semantics/openAccess