Search Results
Plants of Ilex argentina L., I. brasiliensis (S.) L., I. brevicuspis R., I. dumosa R., I. integerrima (V.C.) L., I. microdonta R., I. pseudoboxus R., and I. theezans C.M. were obtained by immature embryo culture. Heart-stage zygotic embryos were removed from immature fruits and cultured aseptically on quarter-strength Murashige and Skoog medium with 3% sucrose, 0.65% agar, and 0.1 mg·L-1 zeatin. Cultures were incubated at 27±2°C for 4 weeks, in darkness and subsequently transferred to a culture room with a 14-hour photoperiod (116 μmol·m-2·s-1) for another 4 weeks. Seedlings with two leaves, derived from germinated embryos, were successfully transplanted to pots containing 1 peat: 1 perlite: 1 sand (v/v) and were maintained in greenhouse conditions. From 95% to 100% of transplanted seedlings survived. Chemical name used: 6-(4-hydroxy-3-methylbut-2-enylamino) purine (zeatin).
An in vitro culture protocol was developed that increased the germination percentage and decreased the lag time to germination for Ilex dumosa R. pyrenes as a tool for replacing the laborious task of embryo rescue technique. This method involves transversely cutting surface-sterilized pyrenes with a scalpel blade, then placing the micropylar one-third end with the rudimentary embryo (≈0.25 mm long) on solidified (agar 0.65%) quarter-strength salts and vitamins of Murashige and Skoog, 1962 medium with 3% sucrose, and incubating in a growth room at 27 ± 2 °C with a 14-h photoperiod (116 μmol·m−2·s−1). Most of the cut pyrenes (greater than 50%) germinated within the first month after inoculation and achieved maximum germination (≈70%) in 2 months compared with whole pyrenes, which began to germinate 3 months after sowing and required more than 8 months for maximum germination (37%). Moreover, the germination percentage of cut pyrenes was significantly higher than the germination of isolated embryos (34%). Thus, the cut pyrenes culture is a simpler and more effective technique than embryo rescue. Easily, on average, a trained operator is able to culture ≈1000 cut pyrenes per day instead of ≈100 isolated embryos.
Pollen storage is of great importance for plant breeding and production besides an efficient means for preservation of haploid gene pool of plant genetic resources and rare or endangered species. Pollinia of Cohniella cepula were stored over 1 year at 4, −20, −70, and −196 °C. Fertilizing ability of fresh and stored (30 to 360 days) pollinia was determined by the fruit and seed formation for each treatment, as well as by the seed viability, in vitro seed germination, and seedling growth. Pollinia stored at −70 and −196 °C showed high fertilizing ability (94.4% to 100.0%) even 1 year after collection, revealing no significant differences with fresh pollinia. Seeds from all treatments showed high viability (91.2% to 94.3%) through the 2,3,5-triphenyltetrazolium chloride (TTC) reduction assay and high in vitro germination (91.7% to 97.3%). Thus, successful ultracold storage of C. cepula pollinia was feasible without any desiccation, cryoprotection, or precooling treatment before placing into an ultra freezer (−70 °C) or immersing in liquid nitrogen (LN) (−196 °C).