Expression cassettes for regulation of expression in monocotyledonous plants

Abstract

The present invention relates to expression cassettes comprising at least one transcription regulating nucleotide sequence obtainable from the group of genes of monocotyledonous plants consisting of caffeoyl-CoA-O-methyltransferase genes, C8,7-sterol isomerase genes, hydroxyproline-rich glycoprotein (HRGP) genes, lactate dehydrogenase genes, and chloroplast protein 12 like genes. More preferably the transcription regulating sequences are obtainable from Zea mays or Oryza sativa . The transcription regulating sequences are especially useful for root/kernel-preferential, leaf/endosperm-preferential, root/silk/kernel-preferential, or constitutive expression.

1 . An expression cassette for regulating expression in a monocotyledonous plant, said expression cassette comprising:
 a) at least one transcription regulating nucleotide sequence functional in a monocotyledonous plant comprising a sequence selected from the group consisting of:
 i) the nucleotide sequence of SEQ ID NO: 2, 3, 6, 7, 8, 11, 12, 13, 14, 15, 16, 19, 20, 21, 22, 23, 24, 27, 28, 29, 56, 57, 58, 61, 62, 63, 66, 67, 68, 71, 72, or 73; 
 ii) a fragment of at least 50 consecutive bases of the nucleotide sequence of SEQ ID NO: 2, 3, 6, 7, 8, 11, 12, 13, 14, 15, 16, 19, 20, 21, 22, 23, 24, 27, 28, 29, 56, 57, 58, 61, 62, 63, 66, 67, 68, 71, 72, or 73; 
 iii) a nucleotide sequence having at least 90% sequence identity to the nucleotide sequence of SEQ ID NO: 2, 3, 6, 7, 8, 11, 12, 13, 14, 15, 16, 19, 20, 21, 22, 23, 24, 27, 28, 29, 56, 57, 58, 61, 62, 63, 66, 67, 68, 71, 72, or 73; and 
 iv) a nucleotide sequence capable of hybridizing to the nucleotide sequence of i) or the complement thereof in 7% sodium dodecyl sulfate (SDS), 0.5 M NaPO 4 , 1 mM EDTA at 50° C. with washing in 2×SSC, 0.1% SDS at 50° C.; 
   and   b) at least one nucleic acid sequence which is heterologous in relation to said transcription regulating nucleotide sequence.   
     
     
         2 . The expression cassette of  claim 1 , wherein expression of the nucleic acid sequence of b) results in expression of a protein, an antisense RNA, a sense RNA, or a double-stranded RNA. 
     
     
         3 . The expression cassette of  claim 1 , wherein expression of the nucleic acid sequence of b) in a plant confers an agronomically valuable trait to said plant. 
     
     
         4 . The expression cassette of  claim 1 , further comprising at least one element selected from the group consisting of:
 a) a 5′-untranslated region of a plant expressed gene;   b) an intron sequence from a plant expressed gene; and   c) a transcription termination sequence from a plant expressed gene.   
     
     
         5 . The expression cassette of  claim 4 , wherein the transcription termination sequence of c) comprises the nucleotide sequence of SEQ ID NO: 32, 34, or 35. 
     
     
         6 . The expression cassette of  claim 4 , wherein the intron sequence of b) has expression enhancing properties. 
     
     
         7 . The expression cassette of  claim 4 , wherein the intron sequence of b) is an intron from an ubiquitin, actin, or alcohol dehydrogenase gene. 
     
     
         8 . A vector comprising the expression cassette of  claim 1 . 
     
     
         9 . A transgenic host cell or non-human organism comprising:
 a) the expression cassette of  claim 1 ; or   b) a vector comprising the expression cassette of a).   
     
     
         10 . A cell culture, part, organ, tissue or transgenic propagation material derived from the non-human organism of  claim 9 , wherein said cell culture, part, organ, tissue or transgenic propagation material comprises the expression cassette. 
     
     
         11 . A method for the transgenic expression of a nucleic acid, said method comprising growing or culturing the non-human organism of  claim 9  or cell cultures, parts, organs, tissues or transgenic propagation material derived therefrom, wherein said cell cultures, parts, organs, tissues or transgenic propagation material comprise the expression cassette. 
     
     
         12 . A transgenic plant comprising:
 a) the expression cassette of  claim 1 ; or   b) a vector comprising the expression cassette of a).   
     
     
         13 . A method for providing a transgenic expression cassette for heterologous expression in a monocotyledonous plant, said method comprising:
 a) isolating a transcription regulating nucleotide sequence from a monocotyledonous plant, wherein the transcription regulating nucleotide sequence shares at least 90% sequence identity to the nucleotide sequence of SEQ ID NO: 2, 3, 6, 7, 8, 11, 12, 13, 14, 15, 16, 19, 20, 21, 22, 23, 24, 27, 28, 29, 56, 57, 58, 61, 62, 63, 66, 67, 68, 71, 72, or 73; and   b) functionally linking said transcription regulating nucleotide sequence to another nucleotide sequence of interest, which is heterologous in relation to said transcription regulating nucleotide sequence.   
     
     
         14 . The method of  claim 13 , further comprising functionally linking said another nucleotide sequence of interest to a transcription termination sequence. 
     
     
         15 . The method of  claim 14 , wherein said transcription termination sequence comprises the nucleotide sequence of SEQ ID NO: 32, 34, or 35. 
     
     
         16 . A method for identifying and/or isolating a sequence with transcription regulating activity, said method comprising:
 a) obtaining polynucleotide sequences sharing at least 90% sequence identity with the nucleotide sequence of SEQ ID NO: 2, 3, 6, 7, 8, 11, 12, 13, 14, 15, 16, 19, 20, 21, 22, 23, 24, 27, 28, 29, 56, 57, 58, 61, 62, 63, 66, 67, 68, 71, 72, or 73;   b) preparing expression cassettes comprising any of the polynucleotide sequences of a) operably linked to a reporter gene or marker and optionally a terminator sequence;   c) testing the expression cassettes for expression activity of the polynucleotide sequence comprised therein; and   d) identifying and/or isolating a polynucleotide sequence having transcription regulating activity.   
     
     
         17 . A method for directing expression in a monocotyledonous plant, said method comprising:
 a) introducing into a plant cell the expression cassette of  claim 1 ;   b) selecting a transgenic cell which comprises said expression cassette, and   c) regenerating a plant from the transgenic cell, wherein the transcription regulating nucleotide sequence directs expression of the nucleic acid sequence which is heterologous in relation to said transcription regulating nucleotide sequence in the plant.