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Title: CRYOPRESERVATION OF SALIX SPECIES USING SECTIONS FROM WINTER VEGETATIVE SCIONS.

Author
item Towill, Leigh
item Widrlechner, Mark

Submitted to: CryoLetters
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 12/1/2003
Publication Date: 6/1/2004
Citation: Towill, L.E., Widrlechner, M.P. 2004. Cryopreservation of salix species using sections from winter vegetative scions. CryoLetters 25:71-80.

Interpretive Summary: The USDA-ARS National Center for Genetic Resources Preservation needs to establish a method for long-term storage of a collection of Salix species from the North Central Regional Plant Introduction Station. Cryopreservation is currently the only method that provides long-term, low maintenance storage for vegetative propagules. We examined several variables in developing a two-step cryopreservation procedure using sections from scions collected in mid-winter. Samples cooled at 5C/day to -30C or -35C and then transferred to liquid nitrogen vapor gave the greatest survival, defined as shoot formation. Cooling rate to 35C had a major influence on survival. Cooling rate from -35C to ca. -180C or -209C did not influence survival. Warming procedures also affected survival and warming at +2C gave the highest survival. Eight of eleven Salix species tested using the established protocol had significant levels of survival after cryogenic treatment.

Technical Abstract: Buds of Salix species are candidates for cryopreservation procedures because they become tolerant of freezing temperatures during mid-winter. We examined several variables in developing a two-step cryopreservation procedure using sections from scions of these materials. Samples of Salix triandra cooled at 5C/day to -30C or -35C and then transferred to Liquid Nitrogen Vapor gave the greatest survival, defined as shoot formation. Cooling rate to 35C had a major influence on survival. Samples exposed to rates greater than 10C/hr showed no survival. The highest survival was achieved by cooling at 0.21C/hr. Cooling rate from -35C to ca. -180C or -209C did not influence survival. Warming procedures also affected survival, although the final temperature rather than the warming rate appeared to be more important. Transferring samples from -160C to either a +2C cold room or to -3C methanol gave similar levels of survival. Warming in +40C water or very slowly gave no survival. Eight of eleven Salix species tested using the established protocol had significant levels of survival after cryogenic treatment. In most cases the cambium remained green after treatment, but some species developed a patchy brown appearance after 5-6 weeks in peat.