CRYOPRESERVATION OF WILLOW SPECIES USING WINTER VEGTATIVE BUDS

Authors: Towill, Leigh; Widrlechner, Mark

Submitted to: HortScience
Publication Type: Abstract Only
Publication Acceptance Date: 3/20/2003
Publication Date: 8/20/2003
Citation: Towill, L.E., Widrlechner, M.P. 2003. Cryopreservation of willow species using winter vegtative buds. HortScience. 38:735-736.

Interpretive Summary: Willow (Salix species) is maintained in the field at the North Central Plant Introduction Station, Ames Iowa. Since willow species are fairly cold hardy in mid winter, long-term preservation using cryogenic storage is possible with winter-harvested field scions. In Fort Collins, we examined several variables to develop a cryopreservation procedure for these species. Most studies used Salix triandra, PI 505949. Previous work had shown that -5C/day was useful as a cooling rate. Samples cooled at this rate were transferred from various temperatures directly to about -180C; best survival, defined by subsequent post thaw shoot elongation or bud expansion) occurred with a transfer temperature of -30C or -35C. Cooling rate from -5C to -35C had a major influence on survival: samples exposed to rates greater than 10C/hr showed no survival. Rates of transfer from -35C to ca. -180C or -196C did not influence survival. Warming rates did influence survival. Rapid warming in +40C water (rates ca. 450C/min) gave no survival, nor did very slow rates (ca. < 1 degree/hr). Rates obtained by transfer to either a +2C cold room (ca. 15C/min or to -3C methanol (ca. 300C/min gave similar, high levels of survival. Eight of 10 other willow species tested using the information shown above had significant levels of survival after cryogenic treatment. In most cases the cambium remained green after treatment, but did develop a patchy brown appearance over time (ca 3-4 weeks). A number of species also developed roots after cryopreservation and direct rooting of sections may be possible. Some species had predominantly catkin buds and percent survival was more difficult to quantify. Recovery conditions will need to be examined to determine whether a direct rooting recovery of scions is feasible or whether either budding or in vitro culture will be necessary to obtain better levels of recovery from cryogenic temperatures.

Technical Abstract: Willow (Salix species) is maintained in the field at the North Central Plant Introduction Station, Ames Iowa. Since willow species are fairly cold hardy in mid winter, long-term preservation using cryogenic storage is possible with winter-harvested field scions. In Fort Collins, we examined several variables to develop a cryopreservation procedure for these species. Most studies used Salix triandra, PI 505949. Previous work had shown that -5C/day was useful as a cooling rate. Samples cooled at this rate were transferred from various temperatures directly to about -180C; best survival, defined by subsequent post thaw shoot elongation or bud expansion) occurred with a transfer temperature of -30C or -35C. Cooling rate from -5C to -35C had a major influence on survival: samples exposed to rates greater than 10C/hr showed no survival. Rates of transfer from -35C to ca. -180C or -196C did not influence survival. Warming rates did influence survival. Rapid warming in +40C water (rates ca. 450C/min) gave no survival, nor did very slow rates (ca. < 1 degree/hr). Rates obtained by transfer to either a +2C cold room (ca. 15C/min or to -3C methanol (ca. 300C/min gave similar, high levels of survival. Eight of 10 other willow species tested using the information shown above had significant levels of survival after cryogenic treatment. In most cases the cambium remained green after treatment, but did develop a patchy brown appearance over time (ca 3-4 weeks). A number of species also developed roots after cryopreservation and direct rooting of sections may be possible. Some species had predominantly catkin buds and percent survival was more difficult to quantify. Recovery conditions will need to be examined to determine whether a direct rooting recovery of scions is feasible or whether either budding or in vitro culture will be necessary to obtain better levels of recovery from cryogenic temperatures.