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Title: Changes in transcript expression patterns as a result of cryoprotectant treatment and liquid nitrogen exposure in Arabidopsis shoot tips

Author
item GROSS, BRIANA - University Of Minnesota
item Henk, Adam
item Bonnart, Remi
item Volk, Gayle

Submitted to: Plant Cell Reports
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 12/8/2016
Publication Date: 12/20/2016
Citation: Gross, B.L., Henk, A.D., Bonnart, R.M., Volk, G.M. 2016. Changes in transcript expression patterns as a result of cryoprotectant treatment and liquid nitrogen exposure in Arabidopsis shoot tips. Plant Cell Reports. 36:459-470. doi:10.1007/s00299-016-2095-7.

Interpretive Summary: Cryopreservation methods have been implemented in genebanks worldwide as a strategy to ensure long-term, secure back-ups of critical collections of plant genetic resources. Effective and efficient cryopreservation methods are particularly necessary for collections of cultivars that are propagated vegetatively, and are thus not amenable to long-term seed storage. Vitrification is a commonly implemented cryopreservation method that uses shoot tips excised from field, greenhouse, or in vitro plants as the source materials. Shoot tips (1 to 2 mm) are then treated with cryoprotectant solutions that remove or replace freezable water. They are then plunged into liquid nitrogen for long term preservation. They can be retrieved and regrown into plants after cryoexposure. We used the model system Arabidopsis thaliana to identify suites of genes that are up- or downregulated in response to cryoprotectant treatments and liquid nitrogen exposure. Of these 180 genes that exhibited significant changes in expression, 67 were related to stress, defense, wounding, lipid, carbohydrate, abscisic acid, oxidation, temperature (cold/heat) or osmoregulation. These gene expression experiments have allowed us to better understand which sets of genes are expressed in response to cryoprotectant treatment and LN exposure. This information can be used to develop improved, more robust cryopreservation methods for long-term conservation in genebanks.

Technical Abstract: Cryopreservation methods have been implemented in genebanks worldwide as a strategy to ensure long-term, secure back-ups of critical collections of plant genetic resources. Effective and efficient cryopreservation methods are particularly necessary for collections of cultivars that are propagated vegetatively, and are thus not amenable to long-term seed storage. Vitrification is a commonly implemented cryopreservation method that uses shoot tips excised from field, greenhouse, or in vitro plants as the source materials. Shoot tips are then treated with cryoprotectant solutions such as Plant Vitrification Solution 2 (PVS2) or Plant Vitrification Solution 3 (PVS3); these solutions remove or replace freezable water within the meristem cells. We used the model system Arabidopsis thaliana to identify suites of transcripts that are up- or downregulated in response to PVS2 and PVS3 treatment and liquid nitrogen (LN) exposure. In total, 180 transcripts showed significant, 2-fold changes in expression level after treatment with either the cryoprotectant or cryopreservation followed by recovery. Of these 180 transcripts, 67 were related to stress, defense, wounding, lipid, carbohydrate, abscisic acid, oxidation, temperature (cold/heat) or osmoregulation. The responses of five transcripts (At4g33030, At1g66180, At3g24500, At4g01250, and At1g75040), were confirmed using qPCR methods. These gene expression experiments have allowed us to better understand which sets of transcripts are expressed in response to cryoprotectant treatment and LN exposure, and the differential effects of treatment with PVS2 and PVS3.