Plants were regenerated from 4 of 19 species of Trillium tested: T. decipiens Freeman, T. grandiflorum Salisb., T. flexipes Raf., and T. pusillum v. pusillum Michx. In 3 species, immature leaf tissues from buds formed hard, spherical outgrowths that produced shoots on a Murashige and Skoog basal medium supplemented with several combinations of auxin (2,4-D or NAA), or cytokinin (BA or 2iP), and coconut water (CW). Tissue from expanding leaves generally did not respond, but one species produced embryo-like structures from which plants developed. Ovular explants showed little or no growth in culture, while rhizome explants consistently showed contamination in vitro. Once initiated, shoot-forming tissue was maintained on a medium lacking growth regulators and CW and continued to produce shoots. Shoots transferred to fresh media produced roots in the presence or absence of the auxin IAA. Rooted plants continued to grow when transferred to soil in the greenhouse. Plants with 3 leaves, comparable to 5- to 7-year-old plants grown from seed, have been produced in <2 years. Chemical names used: (2,4-dichlorophenoxy)acetic acid (2,4-D); l-naphthal-eneacetic acid (NAA); N-(phenylmethyl)-1H-purin-6-amine (BA); N-(3-methyl-2-butenyl)-1H-purin-6-amine (2iP); and 1H-indoIe-3-acetic acid (IAA).
Tissue culture using mature-phase plant material is a useful tool for species conservation, but can be a challenge with oak (Quercus) species, often resulting in low growth and survival. Two different tissue culture media were compared and used to determine whether there was a survival, growth, or contamination response pattern in species representing three North American oak taxonomic sections: red oaks (section Lobatae), white oaks (section Quercus), and golden oaks (section Protobalanus). Mature phase cuttings were harvested in springtime from 12 oak species: arkansas oak (Q. arkansana), canby oak (Q. canbyi), slender oak (Q. graciliformis), nuttall oak (Q. texana), boynton sand post oak (Q. boyntonii), california scrub oak (Q. dumosa), engelmann oak (Q. engelmannii), gambel oak (Q. gambelii), canyon live oak (Q. chrysolepis), palmer oak (Q. palmeri), island oak (Q. tomentella), and huckleberry oak (Q. vacciniifolia). Excised shoot-tip explants were placed onto either Lloyd and McCown woody plant (WP) medium or Gresshoff and Doy (GD) medium. More growth responses and longer survival times were seen on explants grown on the WP medium than GD medium. Explants originating from species native to xeric habitats or those with smooth, glabrous young leaves had significantly higher contamination rates. Although no significant differences were found when grouped by taxonomic section, survival, growth, and contamination varied significantly by species. These findings contribute to the process of establishing tissue culture methods using mature oak material, particularly in relation to medium selection and sterilization protocols, which is critical to the conservation of this iconic group of species.