Validation of a New Multicistronic Plasmid for the Efficient and Stable Expression of Transgenes in Microalgae.

摘要: Low stability of transgenes and high variability their expression levels among the obtained transformants are still pending challenges in nuclear genetic transformation microalgae. We have generated a new multicistronic microalgal plasmid, called Phyco69, to make easier large phenotypic screening usually necessary for selection high-expression stable clones. This plasmid contains polylinker region (PLK) where any gene interest (GOI) can be inserted get linked, through short viral self-cleaving peptide amino terminus aminoglycoside 3'-phosphotransferase (APHVIII) from Streptomyces rimosus, which confers resistance antibiotic paromomycin. The has been validated by expressing second marker, ShBLE gene, phleomycin. It shown, RT-PCR studies, that fusion GOI selective marker APHVIII provides simple method screen select with highest level both transformants. Immunodetection studies shown transcript Phyco69 is correctly processed, producing independent products common promoter.