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US 12,593,767 B2Granted
Protoplast isolation and regeneration of plants
AssigneeThe Regents of the University of California
InventorDavid Tricoli
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Abstract
The present disclosure provides methods for inducing callus formation and plant regeneration from isolated protoplasts of grape. This technology allows for the production of non-chimeric gene edits in grape plants by allowing the delivery of DNA through the plant cell membrane with the recovery of whole plants from a single edited cell.
Claims19
A method of regenerating a grape plant from a grape protoplast, the method comprising:
- (i) encapsulating an isolated grape protoplast in a gel matrix;
- (ii) culturing the encapsulated protoplast in the presence of an osmotically conditioned grape cell suspension culture, wherein the osmotically conditioned grape cell suspension culture is supplemented with picloram and thidiazuron and wherein the protoplast undergoes cell division;
- (iii) culturing the encapsulated protoplast in the presence of activated charcoal and one or more antioxidants and polyamines, wherein the protoplast undergoes cell division and forms a callus colony;
- (iv) transferring the callus colony to an agar-solidified feeder medium, wherein the callus colony develops into a somatic embryo;
- and
- (v) transferring the somatic embryo to an agar-solidified plant medium, wherein the somatic embryo undergoes germination.
3 more elements
Dependencies are inferred from the claim text.