|KRECKEL, H - Colorado State University
|SAMUELS, F - Colorado State University
|STICH, D - University Of Colorado
|LEVINGER, N - Colorado State University
Submitted to: Plants
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 6/1/2023
Publication Date: 6/8/2023
Citation: Kreckel, H.D., Samuels, F.M., Bonnart, R.M., Volk, G.M., Stich, D., Levinger, N.E. 2023. Tracking permeation of dimethyl sulfoxide (DMSO) in Mentha x piperita shoot tips using coherent Raman microscopy. Plants. 12(12). Article e2247. https://doi.org/10.3390/plants12122247.
Interpretive Summary: It is difficult to ensure the long-term security of genebank collections that are maintained as actively growing plants in the field, greenhouse, or screenhouse. It is usually too expensive to create duplicated collections at more than one location. The USDA-ARS National Laboratory for Genetic Resources Preservation develops and uses cryopreservation technologies to place one millimeter shoot tips of these field, greenhouse, and screenhouse collections into liquid nitrogen storage. The shoot tips can be revived and grown into plants when they are needed. Most successful shoot tip cryopreservation methods rely on the use of cryoprotectant solutions that may contain sucrose, dimethyl sulfoxide, ethylene glycol and/or glycerol. The mechanism by which these cryoprotectant solution components function to protect cells during liquid nitrogen exposure is not known. This paper uses several microscopic techniques to visualize the changes that take place within mint shoot tips during dimethyl sulfoxide exposure. Dimethyl sulfoxide enters into the shoot tip cells appears to become concentrated in subcellular compartments. Shoot tips also swell in response to dimethyl sulfoxide treatment. These visualization techniques provide a first look at where dimethyl sulfoxide localizes in living shoot tips.
Technical Abstract: Cryopreservation has emerged as a low-maintenance, cost-effective solution for the long-term preservation of vegetatively propagated crops. Shoot tip cryopreservation often makes use of vitrification methods that employ highly concentrated mixtures of cryoprotecting agents; however, little is understood as to how these cryoprotecting agents protect cells and tissues from freezing. In this study, we use coherent anti-Stokes Raman scattering microscopy to directly visualize where dimethyl sulfoxide (DMSO) localizes within Mentha × piperita shoot tips. We find that DMSO fully penetrates the shoot tip tissue within 10 min of exposure, suggesting an interaction with cellular components that leads to DMSO accumulation in specific regions.