RT Journal Article
T1 Validation of a New Multicistronic Plasmid for the Efficient and Stable Expression of Transgenes in Microalgae
A1 Molina Márquez, Ana María
A1 Vila Spinola, Marta
A1 Rengel, Rocío
A1 Fernández, Emilio
A1 García Maroto, Federico
A1 Vigara Fernández, Javier
A1 León Bañares, Rosa María
AB Low stability of transgenes and high variability of their expression levels among theobtained transformants are still pending challenges in the nuclear genetic transformation ofmicroalgae. We have generated a new multicistronic microalgal expression plasmid, calledPhyco69, to make easier the large phenotypic screening usually necessary for the selection ofhigh‐expression stable clones. This plasmid contains a polylinker region (PLK) where any gene ofinterest (GOI) can be inserted and get linked, through a short viral self‐cleaving peptide to theamino terminus of the aminoglycoside 3’‐phosphotransferase (APHVIII) from Streptomyces rimosus,which confers resistance to the antibiotic paromomycin. The plasmid has been validated byexpressing a second antibiotic resistance marker, the ShBLE gene, which confers resistance tophleomycin. It has been shown, by RT‐PCR and by phenotypic studies, that the fusion of the GOI tothe selective marker gene APHVIII provides a simple method to screen and select the transformantswith the highest level of expression of both the APHVIII gene and the GOI among the obtainedtransformants. Immunodetection studies have shown that the multicistronic transcript generatedfrom Phyco69 is correctly processed, producing independent gene products from a commonpromoter.
PB MDPI
SN 1661-6596
YR 2020
FD 2020-01
LK http://hdl.handle.net/10272/17593
UL http://hdl.handle.net/10272/17593
LA eng
NO Molina Márquez, A., Vila, M., Rengel, R.,  Fernández, E., García Maroto, F.,  Vigara Fernández, J.,  León Bañares, R. (2020). Validation of a New Multicistronic Plasmid for the Efficient and Stable Expression of Transgenes in Microalgae. International Journal of Molecular Sciences, 21(3), 718. DOI: https://doi.org/10.3390/ijms21030718
DS Repositorio Institucional de la Universidad de Huelva
RD 1 jun 2026