In vitro hardening of red raspberry through CO2 enrichment and relative humidity reduction on sugar-free medium

Abstract

Micropropagated shoots of red raspberry (Rubus idaeus L. ’Comet’) were rooted on modified Murashige-Skoog medium lacking sucrose, in specially constructed plexiglass chambers, under ambient (340 ± 20 ppm) or enriched (1500 ± 50 ppm) CO2 and ambient (ca. 100%) or reduced (90 ± 5%) relative humidity. Cultured plantlets were evaluated for their survival, rooting and relative vigor, leaf and root number, stem and root length, total leaf area, total fresh and dry weight, gas exchange rate, and stomatal features, prior to transplantation to soil and at intervals for 6 wk ex vitro. In vitro CO2 enrichment promoted plantlet growth, rooting and both the survival and early growth of transplants. CO2 enrichment increased stomatal aperture of plantlet leaves but did not apparently increase water stress at transplantation. Reduced in vitro RH did not affect plantlet growth but decreased stomatal apertures and stomatal index on leaves of cultured plantlets and promoted both the survival and early growth of transplants. In vitro CO2 and RH levels did not affect the photosynthetic rate of either plantlets or transplants. Only the stomata on leaves of plantlets from the ambient CO2 and reduced RH treatment were functional. Normal stomatal function was not observed in persistent leaves of transplants from the other treatments, even 2 wk after transplantation. In vitro CO2 enrichment acted synergistically with RH reduction in improving growth of plantlets both in vitro and ex vitro. Hardened red raspberry plantlets obtained through CO2 enrichment and RH reduction survived direct transfer to ambient greenhouse conditions without the necessity for specialized ex vitro acclimatization treatment. Key words: Acclimatization, growth analysis, photosynthesis, Rubus idaeus L., stomata, tissue culture