Whisker-mediated transformation of plant cell aggregates and plant tissues and regeneration of plants thereof

Abstract

Plant cell aggregates and plant tissues can be transformed by elongated, needle-like structures called “whiskers”. The process comprises the agitation of plant cell aggregates and plant tissues of the plant to be transformed in the presence of DNA and whiskers, whereby DNA uptake and integration thereof is facilitated. The process may be applicable to other plant cell aggregates and plant tissues which have not proven easily transformable by other techniques.

1 . A method for transforming a plant cell which comprises: 
 (a) selecting at least one cell from a Type I, Type II, hypocotyl-derived, or cotyledon-derived callus culture or from a meristem, pollen, cotyledon, or germ cell tissue; and    (b) inserting DNA into said cell by whisker mediated transformation.    
     
     
         2 . A method for producing a fertile transgenic plant which comprises regenerating a cell produced by the method of  claim 1 .  
     
     
         3 . A method for producing a fertile transgenic plant comprising the steps of: (i) establishing a regenerable callus culture from a plant to be transformed wherein said callus culture is selected from the group consisting of Type I, Type II, hypocotyl-derived, and cotyledon-derived callus culture; (ii) selecting a plant cell aggregate therefrom for transformation; (iii) transforming said plant cell aggregate with DNA by whisker-mediated transformation; (iv) identifying transformed cell lines therefrom; and (iv) regenerating fertile transgenic plants therefrom.  
     
     
         4 . The method of  claim 3  wherein said Type I callus culture is established from  Zea mays  or  Oryza sativa.    
     
     
         5 . The method of  claim 3  wherein said Type II callus culture is established from  Zea mays.    
     
     
         6 . The method of  claim 3  wherein said hypocotyl-derived culture is established from  Gossypium hirsutum.    
     
     
         7 . The method of  claim 3  wherein said cotyledon-derived culture is established from  Gossypium hirsutum.    
     
     
         8 . The method of  claim 3  wherein said plant cell aggregate is initiated on solid medium.  
     
     
         9 . A fertile transgenic plant produced by the method of of  claim 3 .  
     
     
         10 . A DNA construct functional in a plant cell comprising in the 5′ to 3′ direction of transcription, a transcriptional regulatory region functional in said plant cell and having a DNA sequence according to SEQ ID NO:1, and a gene of interest, said gene being either in the sense or antisense orientation.  
     
     
         11 . The transcriptional regulatory region according to  claim 10  comprising the DNA sequence SEQ ID NO:1.