Biopolym. Cell. 2022; 38(1):58-68.
Transformation of the moss (Ceratodon purpureus) with plasmid DNA delivered by novel block-copolymers of the dimethylaminoethyl methacrylate
- Institute of Cell Biology, NAS of Ukraine
14/16, Drahomanov Str., Lviv, Ukraine, 79005
- Ivan Franko National University of Lviv
4, Hrushevskoho Str., Lviv, Ukraine, 79005
- Lviv Polytechnic National University
12, S. Bandery Str., Lviv, Ukraine, 79013
- Institute of Ecology of the Carpathians of National Academy of Sciences of Ukraine
4, Kozelnytska Str., Lviv, Ukraine, 79026
Aim. To investigate the potential of poly(2-dimethylamino)ethyl methacrylate (DMAEMA)-based block-like polymers to serve as gene delivery carriers in moss Ceratodon purpureus (Hedw.) Brid. protoplasts, and to evaluate the level of their phytotoxicity. Methods. Organic synthesis; DNA gel retardation assay; adapted PEG-mediated transformation protocol; PCR; light microscopy. Results. The formation of pDNA complex with DMAEMA-based carriers took place at 0.01-0.1 % concentrations of the polymer. The poly-DMAEMA carriers F8-DM1, F8-DM2 (fluorine-containing), LAcr-DM1, LAcr-DM2 (lauryl acrylate-containing), BAcr-DM1, and BAcr-DM2 (butyl acrylate-containing) were effective as carriers of plasmid DNA pSF3 at C. purpureus transformation. PCR analysis confirmed that the transformants of C. purpureus moss contain GFP as a gene of interest after the protoplast transformation by polymers LAcr-DM2, LAcr-DM1, BAcr-DM2, BAcr-DM1 and F8-DM2. The poly-DMAEMA carriers at working concentration (0.0025 %) were relatively non-toxic for protoplasts of C. purpureus moss. 83.1-93.9 % of viable protoplasts of C. purpureus moss were found after the treatment with studied carriers at that dose. However, at 0.25 % i.e. 100 times higher concentration than that used for moss transformation, the poly-DMAEMA carriers reached their IC50 level. Conclusion. The novel block-like poly-DMAEMA carriers were effective in transformation of C. purpureus moss protoplasts and demonstrated low toxicity.
Keywords: polymeric carrier, poly(2-dimethylamino)ethyl methacrylate, moss Ceratodon purpureus, protoplasts transformation, toxicity.