Effectiveness of microalgae and cyanobacteria cryopreservation (for the strains from the All-Russian Collection of Microorganisms)

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Abstract

The efficiency of cryopreservation of cyanobacteria and eukaryotic microalgae of different taxonomic groups, including charophyte, chlorophyte and ochrophyte microalgae, was assessed using 24 strains of the algological part of the All-Russian Collection of Microorganisms (VKM) as an example. Microalgae cultures differing in cell size, morphological organization of the thallus, reproduction method, presence and size of mucous membranes, and ability to form dormant cells were selected for comparative study. Two types of cryoprotectors (methanol and dimethyl sulfoxide), two types of nutrient media (slant agar and liquid medium), and three methods for determining the survival of microalgae after cryopreservation were tested. It was shown that a two-step cryopreservation protocol using both cryoprotectants (methanol and dimethyl sulfoxide) was successfully applied to all 24 studied strains, regardless of their taxonomic affiliation and morphological features. Based on the results of the experiment, a standard operating procedure for cryopreservation of microalgae was developed, including a liquid culture medium with dimethyl sulfoxide, as well as two methods for determining the survival of microalgae after cryopreservation – growth in a liquid medium and streaks on agar. The proposed protocol not only ensures the preservation of cell viability and the possibility of further use of the strain as a morphologically and genetically representative sample, but also minimizes time and resource costs, as well as the risk of contamination of cultures.

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About the authors

V. V. Redkina

FRC PSCBI RAS

Author for correspondence.
Email: kalmykova_v_v@mail.ru

All-Russian Collection of Microorganisms (VKM), G.K. Skryabin Institute of Biochemistry and Physiology of Microorganisms of the Russian Academy of Sciences

Russian Federation, Pushchino, 142290

A. D. Temraleeva

FRC PSCBI RAS

Email: kalmykova_v_v@mail.ru

All-Russian Collection of Microorganisms (VKM), G.K. Skryabin Institute of Biochemistry and Physiology of Microorganisms of the Russian Academy of Sciences

Russian Federation, Pushchino, 142290

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Supplementary files

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2. Fig. 1. Microphotographs of strains of microalgae and cyanobacteria: a – Chlamydomonas applanata VKM Al-18; b – Spongiosarcinopsis terrestris VKM Al-23; c – Bracteacoccus bullatus VKM Al-97; d – Chlorosarcinopsis eremi VKM Al-132; e – Chlorococcum sphacosum VKM Al-188; f – Tetracystis pampae VKM Al-230; g – Deasonia granata VKM Al-250; h – Coelastrella striolata VKM Al-355; and – Edaphochloris andreevii VKM Al-104; j – Pseudostichococcus sp. VKM Al-293; l – Micractinium thermotolerans VKM Al-332; m – Neochlorella semenenkoi VKM Al-342; n – Chloroidium sacharophilum VKM Al-400; o – Pseudochlorella sp. VKM Al-474; n – Vischeria magna VKM Al-26; p – Vischeria sp. VKM Al-463; c – Botrydiopsis eriensis VKM Al-98; t – Xanthonema bristolianum VKM Al-290; y – Klebsormidium flaccidum VKM Al-9; f – Interfilum terricola VKM Al-274; x – Nostoc sp. VKM Al-391; c – Microcoleus sp. VKM Al-312; h – Anabaena cf. pirinica VKM Al-153; w – Leptolyngbya sp. VKM Al-382. Scale mark – 10 µm.

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3. Fig. 2. Scheme of the experiment to evaluate the effectiveness of cryopreservation of microalgae and cyanobacteria strains.

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4. Fig. 3. Evaluation of the survival rate of microalgae and cyanobacteria during cryopreservation: a, c – Spongiosarcinopsis terrestris VKM Al-23, cultivation period after thawing is 8 weeks; b – Vischeria magna VKM Al-26, cultivation period is 21 days; d – Leptolyngbya sp. VKM Al-382, cultivation period is 4 weeks. Legend in the captions on the dishes and test tubes: M – MetOH, D – DMSO.

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