Enhancing chestnut embryogenic culture productivity and somatic embryo quality by targeting steps in the embryogenesis protocol
Holtz, Christine Therese
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In vitro mass propagation systems have the potential to help re-establish the American chestnut (Castanea dentata) tree, which was devastated by the chestnut blight fungus in the 20th century. Somatic embryogenesis (SE) has been successfully applied for clonal propagation of American chestnuts. We tested the applicability of our published protocol for American chestnut SE induction from seeds of hybrid backcross trees produced by The American Chestnut Foundation (TACF), as well as pure Chinese chestnut. The protocol was successful in producing somatic embryos from the advanced generation hybrid backcross TACF material. In an attempt to improve proliferation of embryogenic tissue in suspension culture, air-lift bioreactors were compared with the standard method, continuously shaken flasks. Air-lift bioreactors showed great potential to enhance embryogenic tissue proliferation. Additionally, RITA® temporary immersion bioreactors were tested to assess their potential for improving somatic embryo germination frequency, but results were inconclusive. Finally, to improve embryo quality, different treatments in which the embryo development medium was supplemented with DLbuthionine–[S,R]-sulfoximine (BSO) were tested. A histological analysis was performed on embryos with different types of morphology. No significant treatment effects were found in the BSO study.