Factors Influencing Somatic Embryo Induction from Orchardgrass Anther Cultures

Somatic embryos from orchardgrass ( Ductylis glomerata L.) anther explants have been initiated in our lab but further research is needed to optimize this response. The objectives of this research were to: (i) determine the role of anther density/orientation and various medium supplements on embryogenesis; and (ii) examine chromosomes and nucleoli from regenerated plants to determine a gametic or somatic cell origin. Anthers were pretreated for 0, 3, or 6 wk at 4°C before planting on Schenk and Hildebrandt (SH) medium containing various levels of dicamba, sucrose, or the cytokinins, 6‐ furfurylaminopurine (kinetin) or benzylaminopurine (BAP). Incubation was in the dark at 25 °C. No differences in response were obtained from a flat vs. an on edge orientation but a density of 10 anthers/mL resulted in a higher frequency of embryogenesis than 100 anthers/mL. Optimum responses were obtained with 30 pmol L −1 dicamba and 263 μmol L −1 sucrose. Both cytokinins had a significant inhibitory effect at all concentrations tested. Somatic embryos were formed directly from somatic tissue of anthers receiving a 0‐ or 3‐wk cold pretreatment whereas those receiving a 6‐wk cold pretreatment initiated embryogenic calli. One of six plants regenerated from anthers exposed to the 6‐wk cold pretreatment was a mixoploid containing cells with 14, 28,56, and more than 100 chromosomes and 1 to 8 nucleoli per nucleus. This work suggests that cytokinins hinder whereas sucrose, dicamba, and a cold pretreatment promote embryogenesis from orchardgrass anthers.