Northwest Plant Breeding Company

Washington State University

Kamterter Ii, L.l.c.

Enrique A Polle

Calvin Konzak

Weiguo Liu

Yuanming Zheng

Calvin F Konzak

The present invention provides methods for generating doubled haploid and/or haploid plants from microspores. In a presently preferred embodiment of the methods of the present invention, plant material is selected that bears reproductive organs containing microspores at a developmental stage that is amenable to androgenic induction. The microspores are treated by contacting the selected plant material with water and subjecting the selected plant material to temperature stress, and optionally to nutrient stress. Preferably the selected plant material is contacted with an effective amount of a sporophytic development inducer and an effective amount of an auxin and/or a cell spindle inhibiting agent. Optionally, the selected plant material is contacted with an effective amount of a cytokinin and/or an effective amount of a gibberellin. The treated microspores are isolated, preferably by density centrifugation utilizing a solution of 0.3 M mannitol layered over a higher density solution of a sugar, preferably maltose. The isolated, treated microspores are then cultured in a liquid nutrient suspension medium supplemented with at least one plant ovary or with an aliquot of plant ovary conditioned medium, until the microspores develop into embryoids. The embryoids are transferred to a regeneration medium and incubated therein until the embryoids develop into plants. The resulting plants may be haploid or doubled haploid and may also be genetically transformed.

Methods for generating doubled haploid plants

Foliar fertilizers which include a phosphate ester in concentrations greater than approximately &frac12; molar with water. The phosphate esters are preferably polyhydroxy alcohol orthophosphate esters, such as ethylene glycol phosphate esters. The fertilizer solutions or mixtures are provided with a water miscible hygroscopic agent which retains the applied fertilizers on the leaf in liquid form by extracting moisture from the atmosphere. The hygroscopic agent is preferably provided by an appropriate hygroscopic alcohol such as used in the production of the esters, or substitutes thereof. The esters can be formed into substituted salts using metals or other elements which are preferably of nutritional value to plants. Methods for the production of the esters and fertilizers are described. Also described are novel methods of use and fertilization using the foliar fertilizer compositions.

Foliar phosphate fertilizers

Processes for the production of ethylene glycol phosphate esters using ethylene glycol and phosphoric acid. These reactants mixed and heated with an esterification promoter which can be an oxide or hydroxide, such as an alkali, alkaline earth or other metal oxide or hydroxide. The mixture can be reacted at atmospheric pressures and temperatures in excess of 150.degree. C., thus speeding the esterification. Boiling is preferred during the reaction to improve water removal. Excess ethylene glycol can advantageously be used to aid in efficient conversion. Excess unreacted ethylene glycol is also significant when the reaction products are used as foliar fertilizers. The cations of some promoters substitute into the esters and can be used to provide secondary or trace elements of nutritional value when the reaction products are used as foliar fertilizers.

Growing community of inventors

Pullman, WA, United States of America

Enrique A Polle