The first evidence for genotypic stability in a cryopreserved transgenic diatom
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Future algal biotechnology will need enhanced production strains, capable of more rapid growth, more efficient solar-energy conversion and/or higher levels of metabolite production. Almost certainly transgenic organisms will be used to ensure the cost-effective, economically viable production of a range of metabolites. As with all biotechnological processes, the functional stability, reliability and security of the production strains will be of paramount importance in algal biotechnology, as without this no biotechnological process is sustainable. In this study, the transgenic model strain Thalassiosira pseudonana CCAP 1085/23 was cryopreserved using a conventional, low-tech, colligative cryopreservation protocol. This employed dimethyl sulphoxide [5 % (v/v)] as a cryoprotectant, using a two-step cooling approach: initial controlled-rate cooling, followed by plunging into liquid nitrogen. High levels of post-thaw viability (70–85 %) were obtained, and on recovery of cryopreserved material no reduction in expression of the protein of the inserted gene (big1-GFP) was observed. Additionally, cryopreservation does not affect the localisation of the BIG1-GFP protein as demonstrated by microscopy of stained samples, nor its functionality as demonstrated by Western blotting.
KeywordsAlgal biotechnology Cryopreservation Genotypic stability Passive freezer Transgenic diatom
Rachel Hipkin was supported by a Norwich Research Park PhD studentship. John Day and Cecilia Rad-Menéndez acknowledge National Capability funding for the CCAP from NERC. Thomas Mock acknowledges funding from NERC (NE/J013730/1), the University of East Anglia for a proof-of-principle grant and Nicole Poulsen for providing vectors and advice regarding nuclear transformation of Thalassiosira pseudonana.
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