Sergey M. Shishlyannikov, Yulia R. Zakharova, Nadezhda A. Volokitina, Ivan S. Mikhailov, Darya P. Petrova, and Yelena V. Likhoshway
Limnol. Oceanogr. Methods 9:478-484 (2011) | DOI: 10.4319/lom.2011.9.478
ABSTRACT: Production of axenic cultures of diatoms is a highly relevant task, as these widespread organisms are objects of research in different fields of biology, from ecology to whole-genome studies, and promising as producers of various organic molecules, including biologically active substances. However, culturing of diatoms isolated from natural communities is accompanied by the development of associated microorganisms. We propose a procedure for establishing an axenic culture of the freshwater diatom Synedra asus subsp. radians (Kütz.) Skabitsch. from Lake Baikal. Such a culture has been produced and maintained by subculturing in our laboratory. The procedure involves filtration (to remove free bacteria), detergent treatment (to remove microorganisms associated with diatom frustules), selection of effective antibiotics and their concentrations nontoxic for diatoms (by the disk diffusion method), cell treatment with the selected antibiotic (ciprofloxacin), repeated filtration, and monoclonal culturing of diatom cells. Axenity of the culture was verified microscopically (after DAPI staining) and by molecular biological methods.