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Blasticidin S Deaminase: A New Efficient Selectable Marker for Chlamydomonas reinhardtii.

Frontiers in plant science (2020-03-27)
Félix de Carpentier, Jeanne Le Peillet, Nicolas D Boisset, Pierre Crozet, Stéphane D Lemaire, Antoine Danon
ABSTRACT

Chlamydomonas reinhardtii is a model unicellular organism for basic or biotechnological research, such as the production of high-value molecules or biofuels thanks to its photosynthetic ability. To enable rapid construction and optimization of multiple designs and strains, our team and collaborators have developed a versatile Chlamydomonas Modular Cloning toolkit comprising 119 biobricks. Having the ability to use a wide range of selectable markers is an important benefit for forward and reverse genetics in Chlamydomonas. We report here the development of a new selectable marker based on the resistance to the antibiotic blasticidin S, using the Bacillus cereus blasticidin S deaminase (BSR) gene. The optimal concentration of blasticidin S for effective selection was determined in both liquid and solid media and tested for multiple laboratory strains. In addition, we have shown that our new selectable marker does not interfere with other common antibiotic resistances: zeocin, hygromycin, kanamycin, paromomycin, and spectinomycin. The blasticidin resistance biobrick has been added to the Chlamydomonas Modular Cloning toolkit and is now available to the entire scientific community.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Hygromycin B from Streptomyces hygroscopicus, suitable for plant cell culture, BioReagent, ≥60% (HPLC), lyophilized powder
Sigma-Aldrich
Kanamycin sulfate from Streptomyces kanamyceticus, powder, BioReagent, suitable for cell culture, suitable for plant cell culture
Sigma-Aldrich
Blasticidin S Ready Made Solution, 10 mg/mL (20 mM HEPES), 0.22 μm filtered
Sigma-Aldrich
Paromomycin sulfate salt, suitable for plant cell culture, BioReagent
Sigma-Aldrich
Evans Blue, Dye content ≥75 %
Sigma-Aldrich
Blasticidine S hydrochloride
Sigma-Aldrich
Spectinomycin dihydrochloride pentahydrate, powder, BioReagent, suitable for cell culture