Sorghum Centromere Sequences and Minichromosomes

Abstract

The invention is generally related to MCs containing sorghum centromere sequences. In addition, the invention provides for methods of generating plants transformed with these MCs. MCs with novel compositions and structures are used to transform plants cells which are in turn used to generate the plant. Methods for generating the plant include methods for delivering the MC into plant cell to transform the cell, methods for selecting the transformed cell, and methods for isolating plants transformed with the MC.

1 . A  sorghum  plant comprising a  sorghum  mini-chromosome comprising a  sorghum  centromere, wherein the  sorghum  centromere comprises at least two copies of a first repeated nucleotide sequence that is at least 80% identical to the nucleotide sequence of any one of SEQ ID NOs:22-176 or hybridizes to the nucleotide sequence of any one of SEQ ID NOs:22-176 under stringent conditions comprising hybridization at 65° C. and washing three times for 15 minutes with 0.25×SSC, 0.1% SDS at 65° C. 
     
     
         2 . A  sorghum  plant cell comprising a  sorghum  mini-chromosome comprising at least two copies of a repeated nucleotide sequence that is at least 80% identical any one of SEQ ID NOs:22-176 or hybridizes to any one of SEQ ID NOs:22-176 under stringent conditions comprising hybridization at 65° C. and washing three times for 15 minutes with 0.25×SSC, 0.1% SDS at 65° C., and a Transgene Expression Cassette 
     
     
         3 . The  sorghum  plant cell of  claim 2 , further comprising
 at least two copies of a second repeated nucleotide sequence that is at least 80% identical over its length to a fragment of the  sorghum  retrotransposon sequence of SEQ ID NO:21 or hybridizes to a fragment of the  sorghum  retrotransposon sequence of SEQ ID NO:21 under stringent conditions comprising hybridization at 65° C. and washing three times for 15 minutes with 0.25×SSC, 0.1% SDS at 65° C.   
     
     
         4 . The  sorghum  plant cell of  claim 3 , comprising, wherein the  sorghum  centromere comprises
 (a) at least 5 copies of the first repeat within 1 kb of nucleotide sequence, and   (b) a transgene expression cassette comprising at least one exogenous nucleic acid.   
     
     
         5 - 8 . (canceled) 
     
     
         9 . The  sorghum  plant cell of  claim 4 , wherein the  sorghum  mini-chromosome exhibits a mitotic segregation efficiency in  sorghum  cells of at least 90%. 
     
     
         10 . The  sorghum  plant cell of  claim 4 , wherein at least one exogenous nucleic acid is operably linked to a heterologous regulatory sequence functional in  sorghum  cells. 
     
     
         11 . The  sorghum  plant cell of  claim 10 , wherein the exogenous nucleic acid is selected from the group consisting of a herbicide resistance gene, a nitrogen fixation gene, an insect resistance gene, a disease resistance gene, a plant stress-induced gene, a nutrient utilization gene, a gene that affects plant pigmentation, a gene that encodes an antisense or ribozyme molecule, a gene encoding a secretable antigen, a toxin gene, a receptor gene, a ligand gene, a seed storage gene, a hormone gene, an enzyme gene, an antibody gene, a growth factor gene, a drought resistance gene, a heat resistance gene, a chilling resistance gene, a freezing resistance gene, an excessive moisture resistance gene, or a salt stress resistance gene or a biofuel gene. 
     
     
         12 . (canceled) 
     
     
         13 . A  sorghum  plant cell comprising a transgene expression cassette comprising at least one exogenous nucleic acid not integrated into the plant cell genome, wherein the transgene expression cassette comprises
 (a) a polynucleotide sequence that is transcribed as a first RNA,   (b) a polynucleotide sequence that is transcribed as a second RNA, and   (c) a polynucleotide sequence that is transcribed as a third RNA   
       wherein transcription of the polynucleotide sequences results in increased biomass of a  sorghum  plant compared to the biomass of a wildtype  sorghum  plant. 
     
     
         14 - 15 . (canceled) 
     
     
         16 . The  sorghum  plant cell of  claim 4 , wherein the
 transgene expression cassette comprises at least three exogenous nucleic acids, and   wherein the first repeated nucleotide sequence and the transgene expression cassette are not integrated into the genome of the  sorghum  plant cell.   
     
     
         17 . A  sorghum  plant cell of  claim 4  that exhibits an altered phenotype associated with at least one exogenous nucleic acid within the  sorghum  [MC] mini-chromosome. 
     
     
         18 . The  sorghum  plant cell of  claim 17 , wherein the altered phenotype comprises increased altered expression of a native gene or the expression of an exogenous gene. 
     
     
         19 - 20 . (canceled). 
     
     
         21 . A  sorghum  plant, plant tissue or  sorghum  plant part comprising the plant cell of any one of  claim 2 . 
     
     
         22 - 24 . (canceled). 
     
     
         25 . A  sorghum  seed obtained from the plant of  claim 21 . 
     
     
         26 . A  sorghum  plant progeny comprising a  sorghum  mini-chromosome, wherein the plant progeny is the result of breeding a plant of  claim 21 . 
     
     
         27 . A method of using a  sorghum  plant of  claim 21 , the method comprising growing the plant to produce a recombinant protein encoded by an exogenous nucleic acid of the mini-chromosome, and alternatively, further comprising a step of harvesting or processing the  sorghum  plant.