Cryopreservation of the gorgonian endosymbiont Symbiodinium

Meng

et al. 2017

Reviews in Aquaculture

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Cryopreservation has been extensively used in various mega‐industries and has recently been applied in genetic banking for conservation purposes. Compared with conventional cell preservation methods, cryopreservation can maintain the viability of cryopreserved cells for an indefinite time at a relatively lower cost and lesser manpower, with lower probabilities of contamination and genetic changes. This study presents the crucial role of sugar, a cryoprotectant supplement in cryopreservation. Sugar molecules typically interact with the lipid bilayer during the freezing phase to maintain plasma membrane integrity when cells undergo dehydration. When combined with other permeable cryoprotectants such as glucose with methanol and trehalose with dimethyl sulphoxide, sugar prolongs and enhances cellular post‐thaw viability. Moreover, terrestrial and marine organisms have benefited from the inclusion of sugar in the cryopreservation protocol. For wide range of cells such as the sperm, oocytes, embryos and larvae of marine vertebrates and invertebrates, as well as marine algae, cryopreservation with sugar produced positive results compared with cryopreservation without sugar. Not all sugar is beneficial, and the type and concentration of sugar should be applied according to the species. Moreover, the freezing method may also affect the function of sugar. Nevertheless, understanding the role of sugar in cryobiology and conducting a preliminary trial of sugar for cryopreserving cells would benefit future research on cryopreservation.

Section: Marine Algaementioning

confidence: 99%

“…Two‐step freezing is one of the most common cryopreservation techniques for marine algae, and using this technique, Chong et al . () cryopreserved Symbiodinium clade G with the CPT of 1 m methanol (MeOH) and 0.4 m sucrose. Tanniou et al .…”

Section: Marine Algaementioning

confidence: 99%

Sugars as supplemental cryoprotectants for marine organisms

Tsai

Meng

et al. 2017

Reviews in Aquaculture

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“…isolated from coral. Chong et al reported high viability when clade G was cryopreserved with 1M (4.05%, w/v) MeOH. Moreover, Symbiodinium clade B isolated from Pseudopterogorgia elisabethae was successfully cryopreserved with 20% MeOH or with 20% ethanol (Santiago‐Vázquez et al ), and Symbiodinium sp.…”

Section: Introductionmentioning

confidence: 99%

“…(Hagedorn and Spindler 2014). Chong et al (2016), Hagedorn et al (2010) and Santiago-Vázquez et al (2007) are among the few that have carried out studies on the cryopreservation of Symbiodinium sp. isolated from coral.…”

Section: Introductionmentioning

confidence: 99%

Use of luminometry and flow cytometry for evaluating the effects of cryoprotectants in the gorgonian coral endosymbiont Symbiodinium

Lin

Wang

et al. 2019

Phycological Research

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SUMMARY Cryopreservation has consistently proven to be a viable method for the storage of a wide variety of biological material, and there has been a recent focus on the cryopreservation of Symbiodinium spp. given the role of these dinoflagellates in the biology of the coral hosts with which they regularly associate. The present study aimed to identify the effect of various cryoprotectants (CPAs) on clade G Symbiodinium isolated from the whip coral Junceella fragilis by analyzing (i) dinoflagellate cell membrane integrity (via SYTOX® staining) and (ii) metabolic function (via an ATP assay) after cryopreservation. At low concentrations (1 M), none of the CPAs tested were harmful to the dinoflagellates at up to 60 min of incubation, and methanol and DMSO were the least harmful; neither caused a significant effect on cell ATP content even after 30 min incubations at 4 M concentrations. Detrimental effects of the CPAs increased in the following order: MeOH (lowest) = DMSO < EG < PG < Gly. With respect to the different assays, the ATP bioassay was more sensitive to CPA exposure than SYTOX staining. Given these findings, MeOH and DMSO should see more widespread use in cryopreservation protocols for clade G Symbiodinium, as well as other dinoflagellates.

“…Dinoflagellates were cryopreserved using an optimised freezing protocol according to a previously described protocol on dinoflagellate extraction and washing59. Dinoflagellate samples were incubated with a CPA (1.0 M methanol and 0.4 M sucrose) in straws for 30 min and were placed 5-cm above liquid nitrogen (LN 2 ) for 30 min.…”

Section: Methodsmentioning

confidence: 99%

Validation of reference genes for cryopreservation studies with the gorgonian coral endosymbiont Symbiodinium

Kuo

Tsai

et al. 2017

Sci Rep

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Quantification by real-time RT-PCR requires a stable internal reference known as a housekeeping gene (HKG) for normalising the mRNA levels of target genes. The present study identified and validated stably expressed HKGs in post-thaw Symbiodinium clade G. Six potential HKGs, namely, pcna, gapdh, 18S rRNA, hsp90, rbcl, and ps1, were analysed using three different algorithms, namely, GeNorm, NormFinder, and BestKeeper. The GeNorm algorithm ranked the candidate genes as follows in the order of decreasing stability: pcna and gapdh > ps1 > 18S rRNA > hsp90 > rbcl. Results obtained using the NormFinder algorithm also showed that pcna was the most stable HKG and ps1 was the second most stable HKG. We found that the candidate HKGs examined in this study showed variable stability with respect to the three algorithms. These results indicated that both pcna and ps1 were suitable for normalising target gene expression determined by performing real-time RT-PCR in cryopreservation studies on Symbiodinium clade G. The results of the present study would help future studies to elucidate the effect of cryopreservation on gene expression in dinoflagellates.