Efficient transfection protocol of rainbow trout gills epithelial (RTgill-W1) cell line through nucleofection system. v1

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dc.catalogadorgjm
dc.contributor.authorEscobar Aguirre, Sebastián Gonzalo
dc.contributor.authorEscorza Sius, Amanda Paz
dc.contributor.authorEscobar Aguirre, Matías
dc.date.accessioned2024-05-06T16:06:25Z
dc.date.available2024-05-06T16:06:25Z
dc.date.issued2024
dc.description.abstractIn this protocol, we optimize the experimental condition to express plasmid DNA in salmonid fish rainbow trout gills epithelial (RTgill-W1), using a nucleofection system. The principle of nucleofection is a combination of electrical parameters that ensures efficient DNA delivery into the nucleus, combined with low toxicity and high cell viability. Using the Neon Transfection System (Invitrogen‱ MPK5000; code: 10431915) and the Invitrogen‱ Neon‱ Transfection System Kit 10 μl (brand: Invitrogen‱ MPK1096; code: 10124334) we were able to express over 60% of GFP in RTgill W 1 line cells. These results were achieved after trying different optimization steps voltage (ranges between 850 - 1600 volts), pulse exposure (times 10 30 ms), and number of pulses, being the condition 1600 volts, 20 ms 1 pulse, and 1 μg μL of plasmid the most effective compared to the other conditions, presenting a higher percentage of GFP expression and lower mortality rate.
dc.fechaingreso.objetodigital2024-05-06
dc.format.extent8 páginas
dc.fuente.origenORCID
dc.identifier.doi10.17504/protocols.io.8epv5x4yng1b/v1
dc.identifier.urihttps://doi.org/10.17504/protocols.io.8epv5x4yng1b/v1
dc.identifier.urihttps://repositorio.uc.cl/handle/11534/85459
dc.information.autorucFacultad de Agronomía e Ingenieria Forestal; Escobar Aguirre, Sebastián Gonzalo; 0000-0001-8826-2096; 1092624
dc.information.autorucFacultad de Ciencias Biológicas; Escorza Sius, Amanda Paz; S/I; 221074
dc.language.isoen
dc.nota.accesocontenido completo
dc.rightsacceso abierto
dc.rights.licenseCC BY 4.0 DEED Attribution 4.0 International
dc.rights.urihttps://creativecommons.org/licenses/by/4.0/
dc.subjectFish cell lines
dc.subjectRT gill
dc.subjectElectroporation
dc.subjectGFP
dc.titleEfficient transfection protocol of rainbow trout gills epithelial (RTgill-W1) cell line through nucleofection system. v1
dc.typepreprint
sipa.codpersvinculados1092624
sipa.codpersvinculados221074
sipa.trazabilidadORCID;2024-05-06
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