NURSING-MEDIA COMPOSITIONS AND METHODS FOR THEIR USE

PRIVATE

FIELD OF THE INVENTION

                    This invention relates to nursing media and to methods for promoting callus formation and increasing efficiency of plant regeneration utilizing nursing media.

BACKGROUND OF THE INVENTION

                    Callus culture is one of the most important plant science techniques for developing clonal populations, plant regeneration, and genetic manipulation.  This technique is essential for propagating large numbers of plants with desired characteristics and for producing plants transformed with new traits introduced through recombinant DNA technology.

                    Callus cultures can be initiated from a variety of explants including stem, petiole, leaf, rhizomes, roots, fruits, endosperm, seeds, hypocotyl, epicotyl, cotyledon, meristems, immature and mature embryos, immature inflorescence, microspores, and the like.  The explant is maintained on a nutrient medium in vitro in a process designed to produce dedifferentiated tissue, or callus tissue, a largely unorganized proliferating mass of parenchyma cells from which plants can be regenerated.

                    Callus tissue is useful in studying a number of physiological and developmental processes in plants, including dedifferentiation and differentiation, embryogenesis, regeneration, and organ formation.  In addition, callus tissue is important in genetic studies and in plant breeding.  For example, callus tissue is used to study mutagenesis followed by the isolation of new mutants, in studies of somaclonal variation, for interspecific hybridization and haploid plant production, and in various studies requiring a shorter breeding cycle.  Callus tissue is utilized in cryopreservation studies, plant pathology studies, and for protein extraction and isolation.  For a review of callus culture and its uses, see Indra, ed. (1984) Cell Culture and Somatic Cell Genetics of Plants, Vol. 1 (Academic Press, New York) and Green et al. (1987) Plant Tissue and Cell Culture (Alan R. Liss Inc., New York).

                    Callus culture is also important for plant transformation.  Both monocot and dicot callus tissue are used in plant transformation protocols.  Callus tissue serves as a target for transformation by ballistic particle acceleration, and the formation of callus tissue may serve as an intermediate step in the recovery of transformed plants following Agrobacterium-mediated transfer of DNA.  Transformation methods for important crops such as maize, wheat, canola, rice, soybean, tobacco, cotton, sorghum, sunflower, and barley may involve callus culture at some step in the overall transformation process for traits including, but not limited to, insect resistance, disease resistance, herbicide resistance, increased yield, increased tolerance to environmental stresses (such as drought, heat, etc.), enhanced seed quality (such as increased or modified starch, oil, and/or protein content), and the like.  Also, callus tissue is used in assays to assess transient gene expression and to evaluate prom...