Embryogenesis factors for cellular reprogramming of a plant cell

Abstract

Plant cell fate and development is altered by treating cells with cellular reprogramming factors. Embryogenesis inducing embryogenesis factor genes and/or morphogenic developmental genes are used as cellular reprogramming factors, specifically comprising polypeptides or polynucleotides encoding gene products for generating doubled haploids or haploid plants from gametes. Maize microspores treated by contacting the isolated cells with an exogenous purified, recombinant embryogenesis inducing embryogenesis factor gene products and/or morphogenic developmental gene polypeptide results in embryogenesis. The gametes of a maize plant develop into embryoids when transformed with a genetic construct including regulatory elements and structural genes capable of acting in a cascading fashion to alter cellular fate of plant cells. Embryogenesis factor proteins and/or developmental morphogenic proteins expressed from a genetic construct are used for ex situ treatment methods and for in planta cellular reprogramming.

1 . A method for promoting haploid embryo formation in a plant cell, the method comprising providing the plant cell with a developmental gene polypeptide and a cyclin delta-2 polypeptide under conditions suitable for embryogenesis. 
     
     
         2 . The method of  claim 1 , wherein the cyclin delta-2 polypeptide comprises an amino acid sequence having at least 90% identity to SEQ ID NO: 21. 
     
     
         3 . The method of  claim 1 , further comprising providing the plant cell with a cyclin-dependent kinase. 
     
     
         4 . The method of  claim 3 , wherein the cyclin-dependent kinase is a CDKA1 polypeptide or a CDKA3 polypeptide. 
     
     
         5 . The method of  claim 1 , wherein providing the plant cell with the developmental gene polypeptide and the cyclin delta-2 polypeptide comprises transforming the plant cell with one or more expression cassettes comprising a first polynucleotide sequence encoding the developmental gene polypeptide and a second polynucleotide sequence encoding the cyclin delta-2 polypeptide. 
     
     
         6 . The method of  claim 1 , wherein the developmental gene polypeptide is a WUS/WOX homeobox polypeptide, a BBM polypeptide, a LEC1 polypeptide, or a LEC2 polypeptide. 
     
     
         7 . The method of  claim 6 , wherein the WUS/WOX homeobox polypeptide comprises an amino acid sequence having at least 90% identity to SEQ ID NO: 247, 249, 251, 253, 255, 257, 259, 261, 263, 265, 267, 269, 271, 273, 275, 277, 279, 281, 283, 285, 287, 289, 291, 293, 295, 297, 299, 301, 303, 305, 307, 309, 311, 313, or 315. 
     
     
         8 . The method of  claim 6 , wherein the BBM polypeptide comprises an amino acid sequence having identity to SEQ ID NO: 121. 
     
     
         9 . The method of  claim 6 , wherein the LEC1 polypeptide comprises an amino acid sequence having at least 90% identity to SEQ ID NO: 125. 
     
     
         10 . The method of  claim 3 , wherein providing the plant cell with the cyclin-dependent kinase comprises transforming the plant cell with an expression cassette comprising a third polynucleotide sequence encoding the cyclin-dependent kinase. 
     
     
         11 . The method of  claim 4 , wherein the CDKA1 polypeptide comprises an amino acid sequence having at least 90% identity to SEQ ID NO: 114. 
     
     
         12 . The method of  claim 4 , wherein the CDKA3 polypeptide comprises an amino acid sequence having at least 90% identity to SEQ ID NO: 115. 
     
     
         13 . The method of  claim 5 , wherein the second polynucleotide sequence encoding the cyclin delta-2 polypeptide comprises a polynucleotide sequence having at least 90% identity to SEQ ID NO: 5. 
     
     
         14 . The method of  claim 1 , wherein promoting haploid embryo formation in the plant cell comprises promoting somatic embryogenesis, parthenogenesis, or microspore embryogenesis in the plant cell. 
     
     
         15 . A method for obtaining a doubled haploid plant, the method comprising:
 (a) providing a haploid plant cell with a developmental gene polypeptide and a cyclin delta-2 polypeptide   (b) obtaining a doubled haploid plant cell; and   (c) regenerating the doubled haploid plant from the doubled haploid plant cell.   
     
     
         16 . The method of  claim 15 , wherein the cyclin delta-2 polypeptide comprises an amino acid sequence having at least 90% identity to SEQ ID NO: 21. 
     
     
         17 . The method of  claim 15 , wherein the developmental gene polypeptide is a WUS/WOX homeobox polypeptide, a BBM polypeptide, a LEC1 polypeptide, or a LEC2 polypeptide. 
     
     
         18 . The method of  claim 17 , wherein the BBM polypeptide comprises an amino acid sequence having identity to SEQ ID NO: 121. 
     
     
         19 . The method of  claim 15 , wherein providing the haploid plant cell with the developmental gene polypeptide and the cyclin delta-2 polypeptide comprises transforming the plant cell with one or more expression cassettes comprising a first polynucleotide sequence encoding the developmental gene polypeptide and a second polynucleotide sequence encoding the cyclin delta-2 polypeptide. 
     
     
         20 . The method of  claim 19 , wherein the second polynucleotide sequence encoding the cyclin delta-2 polypeptide comprises a polynucleotide sequence having at least 90% identity to SEQ ID NO: 5. 
     
     
         21 . The method of  claim 15 , wherein obtaining the doubled haploid plant cell occurs without a chemical chromosome doubling agent.