Regulatory sequences for transgenic plants

Abstract

Regulatory seqeunces derived from the maize per5 gene have utility in plant biotechnology.

We claim:  
     
         1 . An isolated DNA molecule selected from the following per5 promoter sequences 
 bp 4086-4148 of SEQ ID NO 1,    bp 4086 to 4200 of SEQ ID NO 1,    bp 4086 to 4215 of SEQ ID NO 1,    bp 3187-4148 of SEQ ID NO 1,    bp 3187-4200 of SEQ ID NO 1,    bp 3187-4215 of SEQ ID NO 1,    bp 2532-4148 of SEQ ID NO 1,    bp 2532-4200 of SEQ ID NO 1,    bp 2532-4215 of SEQ ID NO 1,    bp 1-4148 of SEQ ID NO 1,    bp 14200 of SEQ ID NO 1, and    bp 1-4215 of SEQ ID NO 1,    or a fragment, genetic variant or deletion of such a sequence which retains the ability of functioning as a promoter in plant cells.    
     
     
         2 . An isolated DNA molecule selected from the following per5 intron sequences 
 bp 4426-5058 of SEQ ID NO 1,    bp 4420-5064 of SEQ ID NO 1,    bp 5251-5382 of SEQ ID NO 1,    bp 5245-5388 of SEQ ID NO 1,    bp 5549-5649 of SEQ ID NO 1, and    bp 5542-5654 of SEQ ID NO 1.    
     
     
         3 . An isolated DNA molecule corresponding to the per5 transcription termination sequence and having the sequence of bp 6068-6431 of SEQ ID NO 1.  
     
     
         4 . An isolated DNA molecule having a 20 base pair nucleotide portion identical in sequence to a consecutive 20 base pair portion of the sequence set forth in SEQ ID NO 1.  
     
     
         5 . A recombinant gene cassette competent for effecting preferential expression of a gene of interest in a selected tissue of transformed maize, said gene cassette comprising: 
 a) a promoter operable in maize;    b) an untranslated leader sequence;    c) the gene of interest;    d) a 3′UTR;    said promoter, untranslated leader sequence, gene of interest, and 3′UTR being operably linked from 5′ to 3′; and    e) an intron sequence that is incorporated in said untranslated leader sequence,in said gene of interest, or in said 3′UTR, said intron sequence being from an intron of a maize gene that is preferentially expressed in said selected tissue, and said intron sequence being from a gene other than the gene of interest.    
     
     
         6 . A recombinant gene cassette of  claim 5 , wherein the promoter is from a first maize gene, said first maize gene being one that is naturally expressed preferentially in the selected tissue.  
     
     
         7 . A recombinant gene cassette of  claim 5  wherein said intron sequence is incorporated in said untranslated leader.  
     
     
         8 . A recombinant gene cassette of  claim 5  wherein said selected tissue is root tissue.  
     
     
         9 . A recombinant gene cassette of  claim 8  wherein said intron sequence is comprised of bp 4420 to bp 5064 of SEQ ID NO 1.  
     
     
         10 . A recombinant gene cassette of  claim 5  wherein said promoter is a per5 promoter comprised of bp 2532-4148 of SEQ ID NO 1.  
     
     
         11 . A recombinant gene cassette of  claim 10  wherein said promoter is a per5 promoter comprised of bp 1-4148 of SEQ ID NO 1.  
     
     
         12 . A recombinant gene cassette of  claim 5  wherein the 3′UTR is a per5 3′UTR comprised of bp 6068 to bp 6431 of SEQ ID NO 1.  
     
     
         13 . A recombinant gene cassette competent for effecting constitutive expression of a gene of interest in transformed maize comprising: 
 a) a promoter from a first maize gene, said first maize gene being one that is naturally expressed preferentially in a specific tissue;    b) an untranslated leader sequence;    c) the gene of interest, said gene being one other than said first maize gene;    d) a 3′UTR;    said promoter, untranslated sequence, gene of interest, and 3′UTR being operably linked from 5′ to 3′; and    e) an intron sequence that is incorporated in said untranslated leader or in said gene of interest, said intron sequence being from an intron of a maize gene that is naturally expressed constitutively.    
     
     
         14 . A recombinant gene cassette of  claim 13  wherein said intron is the Adh1 intron 1 or an operative portion thereof.  
     
     
         15 . A recombinant gene cassette of  claim 14  wherein said promoter is a per5 promoter comprised of bp 2532 to 4148 of SEQ ID NO 1, or an operative portion thereof.  
     
     
         16 . In a recombinant gene cassette for effecting expression of a gene of interest in a transformed plant cell wherein said gene cassette is comprised of: 
 a promoter;    an untranslated leader sequence;    the gene of interest, said gene of interest being a gene other than per5; and    a 3′UTR;    the improvement wherein said 3′UTR is a per5 3′UTR comprised of bp 6068 to 6431 of SEQ ID NO 1.    
     
     
         17 . A recombinant gene cassette of  claim 16  wherein said promoter is selected from the group consisting of the 3 5T promoter, the ubiquitin promoter, and the per5 promoter comprising bp 2532 to 4148 of SEQ ID NO 1.  
     
     
         18 . A DNA construct comprising, operatively linked in the 5′ to 3′ direction, 
 a) a promoter comprising bp 4086-4148 bp of SEQ ID NO 1;  
 b) an untranslated leader sequence,  
 c) a gene of interest not naturally associated with said promoter;  
 d) a 3′UTR.  
 
     
     
         19 . A DNA construct of  claim 18  wherein the promoter and untranslated leader sequence together comprise bp 4086-4200 of SEQ ID NO 1.  
     
     
         20 . A DNA construct of  claim 18  wherein the promoter is comprised of bp 3187-4148 of SEQ ID NO 1.  
     
     
         21 . A DNA construct of  claim 18  wherein the promoter is comprised of bp 2532-4148 of SEQ ID NO 1.  
     
     
         22 . A DNA construct of  claim 18  wherein the promoter is comprised of bp 1-4148 of SEQ ID NO 1.  
     
     
         23 . A DNA construct of  claim 18  wherein said 3′UTR is the nos 3′UTR.  
     
     
         24 . A DNA construct of  claim 18  wherein said 3′UTR has the sequence of bp 6066-6550 of SEQ ID NO 1.  
     
     
         25 . A DNA construct comprising, operatively linked in the 5′ to 3′ direction, 
 a) a promoter comprised of bp 4086-4148 bp of SEQ ID NO 1;  
 b) an intron selected from the group consisting of Adh1 intron 1 and bp 4426-5058 of SEQ ID NO 1;  
 c) a gene of interest not normally associated with said promoter;  
 d) a 3′UTR.  
 
     
     
         26 . A DNA construct of  claim 25  wherein said 3′UTR is selected from the group consisting of nos and bp 6067-6340 of SEQ ID NO 1.  
     
     
         27 . A DNA construct of  claim 25  wherein said 3′UTR is selected from the group consisting of nos and bp 6067-6439 of SEQ ID NO 1.  
     
     
         28 . A DNA construct comprising, in the 5′ to 3′ direction, 
 a) a promoter having as at least part of its sequence bp 4086-4148 bp of SEQ ID NO 1;  
 b) an intron selected from the group consisting of Adh1 intron 1 and bp 4426-5058 of SEQ ID NO 1;  
 c) a cloning site;  
 d) a 3′UTR.  
 
     
     
         29 . A DNA construct of  claim 28  wherein said 3′UTR is selected from the group consisting of nos and bp 6067-6340 of SEQ ID NO 1.  
     
     
         30 . A plasmid including a promoter that is comprised of bp 4086-4148 of SEQ ID NO 1.  
     
     
         31 . A plasmid of  claim 30  wherein the promoter is comprised of bp 3187-4148 of SEQ ID NO 1.  
     
     
         32 . A plasmid of  claim 30  wherein the promoter is comprised of bp 2532-4148 of SEQ ID NO 1.  
     
     
         33 . A plasmid of  claim 30  wherein the promoter is comprised of bp 1-4148 of SEQ ID NO 1.  
     
     
         34 . A plasmid comprising a recombinant gene cassette of  claim 5 .  
     
     
         35 . A plasmid comprising a DNA construct of  claim 18 .  
     
     
         36 . A transformed plant comprising at least one plant cell that contains a recombinant gene cassette according to  claim 5 .  
     
     
         37 . A transformed plant comprising at least one plant cell that contains a DNA construct according to  claim 18 .  
     
     
         38 . Seed or grain that contains a recombinant gene cassette of  claim 5 .  
     
     
         39 . Seed or grain that contains a DNA construct of  claim 18 .  
     
     
         40 . A method for expressing a gene of interest preferentially in a selected tissue which comprises transforming maize with a gene cassette of  claim 5 .  
     
     
         41 . A method for expressing a gene of interest in maize preferentially in root tissue which comprises transforming maize with a gene cassette of  claim 5  wherein the selected tissue is root tissue.  
     
     
         42 . A method of  claim 41  wherein the intron sequence in the gene cassette is comprised of bp 4420 to 5064 of SEQ ID NO 1.  
     
     
         43 . A method of  claim 40 , wherein the promoter in the gene cassette is a per 5 promoter comprised of bp 2532 to 4148 of SEQ ID NO 1, or an operative portion thereof.