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Media type:
E-Article
Title:
Blasticidin-S deaminase, a new selection marker for genetic transformation of the diatom Phaeodactylum tricornutum
Contributor:
Buck, Jochen M.;
Río Bártulos, Carolina;
Gruber, Ansgar;
Kroth, Peter G.
imprint:
PeerJ, 2018
Published in:PeerJ
Language:
English
DOI:
10.7717/peerj.5884
ISSN:
2167-8359
Origination:
Footnote:
Description:
<jats:p>Most genetic transformation protocols for the model diatom <jats:italic>Phaeodactylum tricornutum</jats:italic> rely on one of two available antibiotics as selection markers: Zeocin (a formulation of phleomycin D1) or nourseothricin. This limits the number of possible consecutive genetic transformations that can be performed. In order to expand the biotechnological possibilities for <jats:italic>P. tricornutum</jats:italic>, we searched for additional antibiotics and corresponding resistance genes that might be suitable for use with this diatom. Among the three different antibiotics tested in this study, blasticidin-S and tunicamycin turned out to be lethal to wild-type cells at low concentrations, while voriconazole had no detectable effect on <jats:italic>P. tricornutum</jats:italic>. Testing the respective resistance genes, we found that the blasticidin-S deaminase gene (<jats:italic>bsr</jats:italic>) effectively conferred resistance against blasticidin-S to <jats:italic>P. tricornutum</jats:italic>. Furthermore, we could show that expression of <jats:italic>bsr</jats:italic> did not lead to cross-resistances against Zeocin or nourseothricin, and that genetically transformed cell lines with resistance against Zeocin or nourseothricin were not resistant against blasticidin-S. In a proof of concept, we also successfully generated double resistant (against blasticidin-S and nourseothricin) <jats:italic>P. tricornutum</jats:italic> cell lines by co-delivering the <jats:italic>bsr</jats:italic> vector with a vector conferring nourseothricin resistance to wild-type cells.</jats:p>