A tissue culture laboratory exercise illustrating regeneration of whole plants from leaf segments of chrysanthemum by organogenesis is described. Using simple, common media, shoots can be generated in 5 weeks and rooted after an additional 3 weeks. Acclimatization of plants can be accomplished in a simple mistbed in the greenhouse. The exercise is adaptable to depict genotype differences among cultivars, optimization of shoot induction, effects of growth regulators, and experimental design. Callus is typically not formed during shoot formation; however, co-cultivation of leaf segments with a virulent strain of Agrobacterium tumefaciens produces callus with a strain of disarmed A. tumefaciens harboring NPTII construct affects regeneration of plants resistant to kanamycin.
R.N. Trigiano and R.A. May
R.A. May and R.N. Trigiano
Somatic embryogenesis from leaf midrib explants of Dendranthema grandiflora Tzvelev. `Iridon' cultured on modified Murashige and Skoog basal medium (MSB) containing 1.0 mg 2,4-D and 0.2 mg BA/liter was influenced by light and sucrose concentration. Somatic embryos formed directly from explants when cultured on medium containing 9% to 18% sucrose and incubated first in the dark for 28 days, followed by 10 days in light, and then returned to the dark for 14 days. Embryogenesis did not occur in continuous darkness and was drastically reduced when explants were incubated in light only. The most embryos were formed on medium containing either 12% or 15% sucrose; lower concentrations stimulated shoot and root development. Light also mediated embryogenesis from leaf explants of 'other cultivars. White-opaque or occasionally light-green cotyledon-stage somatic embryos germinated on MSB medium without growth regulators but containing 3% sucrose. Twelve of the 23 cultivars evaluated produced somatic embryos, but plants were recovered from only five. Regenerated plants were phenotypically similar to parent plants in growth habit, leaf morphology, and flower color. Chemical names used: N- (phenylmethyl)-1 H- purine-6-amine (BA); (2,4-dichlorophenoxy) acetic acid (2,4-D).
May Elfar Altamimi, Rhonda R. Janke, Kimberly A. Williams, Nathan O. Nelson and Leigh W. Murray
Greenhouse experiments were conducted to determine the response of Brassica oleracea L., pac choi to fertilizer rates and sources and to establish optimal soluble nitrogen (N) application rates and nitrate meter sufficiency ranges. Conventional soluble fertilizer was formulated from inorganic salts with a 4:1 NO3-N:NH4-N ratio. Phosphorus (P) was held at 1.72 mm and potassium (K) at 0.83 mm for all treatment levels. The organic soluble fertilizer, fish hydrolyzate (2N–1.72P–0.83K), was diluted to provide the same N levels as with conventional treatments. Both fertilizers were applied at N rates of 0, 32, 75, 150, 225, 300, and 450 mg·L−1. Seedlings were transplanted and fertilizer application began at 18 days. Plants were harvested at 7 weeks (5 weeks post-transplanting) after receiving 15 fertilizer applications during production. Samples of the most recently matured leaves were harvested weekly and analyzed for petiole sap NO3-N and leaf blade total N concentration. Leaf count, leaf length, and chlorophyll content were also measured weekly. Fresh and dry weights were determined on whole shoots and roots. Optimum yield was achieved at the 150-mg·L−1 fertility rate with both conventional and organic fertilizers. Field and high tunnel experiments were conducted to validate the sufficiency ranges obtained from the greenhouse studies. Sufficiency levels of NO3-N for pac choi petiole sap during Weeks 2 to 3 of production were 800 to 1500 mg·L−1 and then dropped to 600 to 1000 mg·L−1 during Weeks 4 through harvest for both conventional and organic fertilizers sources. Total N in leaf tissue was less responsive to fertilizer rate effects than petiole sap NO3-N. Chlorophyll content was not useful in evaluating pac choi N status. These guidelines will provide farmers with information for leaf petiole sap NO3-N to guide in-season N applications.
Christian M. Baldwin, Eugene K. Blythe, A. Douglas Brede, Jami J. Mayer and R. Golembiewski
The use of glyphosate-tolerant perennial ryegrass (Lolium perenne L.) (PRG) cultivars JS501 and Replay provides turfgrass managers a unique option for annual bluegrass (Poa annua L.) (ABG) control. Both cultivars can tolerate a maximum glyphosate rate of 0.81 kg·ha−1 acid equivalent (a.e.) after establishment under optimal growing temperatures (16 to 24 °C). However, tolerance to applications made immediately after germination and during low air temperatures has received limited investigation. Therefore, objectives of this research were to determine the seedling tolerance and low-temperature response after a fall season glyphosate application to both cultivars. Field trials were conducted in Idaho and Oregon. For the fall application response trial in Idaho, glyphosate was applied at 0, 0.15, 0.29, 0.58, 1.16, 1.74, 2.32, and 3.48 kg·ha−1 a.e. In Oregon, glyphosate was applied at 0, 0.15, 0.29, 0.44, 0.58, 1.16, and 3.48 kg·ha−1 a.e. At both sites, applications were made between late September and early October. To determine seedling tolerance, both cultivars were sprayed with glyphosate at the one-leaf stage (LS), two LS, three LS, or four LS at rates of 0, 0.15, 0.29, and 0.58 kg·ha−1 a.e. Across all trials, ratings included PRG color, cover, and injury. At both trial locations, regression analysis revealed a rate of ≈0.27 kg·ha−1 a.e. was required to cause 20% leaf firing in the fall application response trial. In the seedling tolerance trial, glyphosate applied at 0.58 kg·ha−1 a.e. at the one LS, two LS, and three LS had color ratings 8.0 or greater; however, color ratings dropped to 4.6 when an application was made at the four LS. Based on the environmental conditions of each trial, results suggest glyphosate applications greater than 0.27 kg·ha−1 a.e. as minimum air temperatures approach 0 °C should be avoided. Also, applications should be avoided at the three to four LS if the application rate is greater than 0.29 kg·ha−1 a.e.