Is tissue culture a viable system with which to examine environmental and hormonal regulation of cold acclimation in woody plants?

Abstract

In vitro‐grown saskatoon berry (Amelanchier alnifolia Nutt.) plantlets were exposed to various hormonal treatments, dormancy‐inducing and cold acclimation conditions to determine if this in vitro system would be viable for dormancy/hardiness studies in woody plants. Low temperature induced significant hardiness levels in plantlets to −27°C after 6 weeks at 4°C but did not approach liquid nitrogen levels of fully hardened, field‐grown buds. Control plantlets were consistently killed at −5°C throughout this period. Significant hardiness was attained under both short and long day/low temperature conditions; however, hardiness was reduced under continuous light or dark treatments. A pre‐exposure to the typical short photoperiod regime of woody plants did not significantly increase the rate of acclimation in these plantlets. The presence/absence of phytohormones in the media have a pronounced influence on the ability of plantlets to cold acclimate. Hormone‐free media increased hardiness to −10.5°C after 2 weeks in treatment. Addition of abscisic acid (ABA) increased cold hardiness levels (−12°C) while addition of benzylaminopurine (BAP) to this hormone‐free media decreased hardiness to −5.3°C. A combination of BAP and ABA treatments produced LT50 values intermediate between individual applications of either hormone. Conversely, α‐naphthaleneacetic acid (NAA) could not counteract the ABA‐induced hardiness. ABA treatments alone were not able to harden plantlets to the extent attained under low temperature acclimation conditions. Further, ABA could not maintain the hardiness levels of cold‐acclimating treatments and plantlets de‐acclimated to −9°C in BAP + ABA media. Subculturing in itself significantly elevated cold hardiness in plantlets to −9°C on BAP + NAA media within 3 days after subculture and thereafter plantlets dehardened to −5°C. While tissue culture has value in specific cases, caution should be taken when using tissue‐cultured plantlets as a system to evaluate environmental regulation of cold acclimation in woody plants, in part, due to the influence of phytohormones in the media.