Somatic Embryogenesis and Plant Regeneration from Suspension Cultures of Switchgrass

Abstract

Embryogenic cell suspension cultures represent an enhanced culture type for the selection of mutants, mass propagation, direct delivery of DNA in to intact regenerable cells and isolation of totipotent protoplasts. The objectives of this study with switchgrass (Panicum virgatum L.) were to establish embryogenic suspension cultures and to regenerate plants from these cultures. The cultures were initiated from embryogenic callus obtained from young inflorescences. They were maintained in Murashige and Skoog liquid medium containing 9.0 μM 2,4‐dichlorophenoxyacetic acid (2,4‐D), 4.4 μM 6‐benzylaminopurine (BAP) and 30 g ‐1 maltose. The s uspension was passed througha 210‐μm mesh and was comprised mainly of small, round densely cytoplasmic starch containing cells with distinct nuclei. Somatic embryos developed from these cells directly and were free floating in the medium. A sequence of developmental stages from embryogenic cells to fully developed embryos was observed in the cultures. The embryos had the typical morphology of a grass embryo. Embryogenic calluses were also obtained 4 wk after the suspension was plated onto solid medium containing various combinations of 2,4‐D and BAP. Embryos from suspension and suspension‐derived callus cultures germinated to produce plants that were successfully established in the greenhouse.

Switchgrass as a biofuels feedstock in the USA

“…Some of these advancements include identification of cpDNA polymorphisms between upland and lowland switchgrass phenotypes (Hultquist et al 1996), identification of genetic incompatibility systems and development of upland × lowland hybrids of switchgrass (Martinez-Reyna et al 2001;Martinez-Reyna and Vogel 2002), identification of the role of tiller dynamics and phytomer size on seedling development and biomass production of switchgrass (Smart et al 2004;Boe and Casler 2005), identification of the role of lignification in limiting cellulosic fermentation of switchgrass herbage and its agronomic implications (Casler et al 2002;Vogel et al 2002a ;Sarath et al 2005), and development of somatic embryogenesis for asexual propagation of switchgrass genotypes (Gupta and Conger 1999).…”

“…Increased efforts to identify heterotic combinations of upland and lowland genotypes, combined with intrapopulation improvement within both upland and lowland populations will improve the economic potential of switchgrass hybrids. Switchgrass plants with superior hybrid combining ability can be asexually propagated by somatic embryogenesis, providing a mechanism for largescale propagation of hybrid seed production fields for superior two-clone hybrids (Gupta and Conger 1999).…”

Molecular Breeding of Grasses by Transgenic Approaches for Biofuel Production

“…Embryogenic suspension cultures initiated from embryogenic callus were also established in switchgrass (Gupta & Conger, 1999), and osmotic pre-treatment with 0.3 M each of sorbitol and mannitol was effective for plant regeneration from suspension cultures (Odjakova & Conger, 1999). Another type of culture system that is highly attractive for gene transfer experiments was also established: multiple shoot clumps were induced from intact seedlings with various combinations of 2,4-D and 1-phenyl-3-(1,2,3-thidiazol-5-yl) urea (TDZ) (Gupta & Conger, 1998).…”

Bioenergy Feedstock Development Program Status Report

“…These tools include development of tissue culture techniques for clonal reproduction of parent plants, molecular fingerprinting, genetic mapping, and linkage of cultures. 68,69,70,71,72,73,74 These clonal propagation techniques are being used successfully at UT to provide tools to assist classical breeding (Fig. 2.10).…”