Nucleic acid molecules that confer resistance to coleopteran pests

Abstract

This disclosure concerns nucleic acid molecules and methods of use thereof for control of coleopteran pests through RNA interference-mediated inhibition of target coding and transcribed non-coding sequences in coleopteran pests. The disclosure also concerns methods for making transgenic plants that express nucleic acid molecules useful for the control of coleopteran pests, and the plant cells and plants obtained thereby.

1 . A double-stranded ribonucleic acid (dsRNA) molecule containing a duplex region comprising:
 a first polyribonucleotide comprising at least 21 contiguous nucleotides of a first nucleotide sequence selected from the group consisting of SEQ ID NO:89, SEQ ID NO:90, SEQ ID NO:92, and the complements and reverse complements thereof; and   a second polyribonucleotide hybridized to the first polyribonucleotide.   
     
     
         2 . The dsRNA molecule of  claim 1 , wherein the first polyribonucleotide comprises at least 21 contiguous nucleotides of SEQ ID NO:91. 
     
     
         3 . The dsRNA molecule of  claim 1 , wherein the second polyribonucleotide comprises a second nucleotide sequence that is the complement or the reverse complement of the first nucleotide sequence. 
     
     
         4 . The dsRNA molecule of  claim 1 , wherein the molecule is a hairpin RNA (hpRNA) molecule further containing a loop region comprising a spacer polyribonucleotide, wherein the spacer polyribonucleotide is located between the first and second polyribonucleotides in the molecule. 
     
     
         5 . A nucleic acid molecule comprising a polynucleotide operably linked to a heterologous promoter, wherein the polynucleotide encodes a hairpin ribonucleic acid (hpRNA) molecule, the polynucleotide comprising:
 a first nucleotide sequence comprising at least 21 contiguous nucleotides of a nucleotide sequence selected from the group consisting of SEQ ID NO:73, SEQ ID NO:75, SEQ ID NO:81, and the reverse complements thereof;   a second nucleotide sequence; and   a third nucleotide sequence that is at least 90% complementary to the reverse complement of the first nucleotide sequence,   wherein the second nucleotide sequence is located between the first and third nucleotide sequences in the polynucleotide.   
     
     
         6 . The nucleic acid molecule of  claim 5 , wherein the first nucleotide sequence comprises at least 21 contiguous nucleotides of SEQ ID NO:77 or the reverse complement thereof. 
     
     
         7 . The nucleic acid molecule of  claim 5 , wherein the heterologous promoter is functional in a plant cell. 
     
     
         8 . A plant material comprising the dsRNA molecule of  claim 4 . 
     
     
         9 . The plant material of  claim 8 , wherein the plant material is a transgenic plant material that expresses the dsRNA molecule. 
     
     
         10 . The transgenic plant material of  claim 9 , wherein the plant material is a plant cell. 
     
     
         11 . The transgenic plant material of  claim 9 , wherein the plant material is canola. 
     
     
         12 . A transgenic plant or seed thereof comprising the nucleic acid molecule of  claim 7 , wherein the heterologous promoter is functional in the plant. 
     
     
         13 . The transgenic plant or seed thereof of  claim 12 , wherein the plant is canola. 
     
     
         14 . A transgenic plant commodity product comprising the dsRNA molecule of  claim 1 . 
     
     
         15 . A sprayable formulation or artificial pollen beetle diet comprising the dsRNA molecule of  claim 1 . 
     
     
         16 . A method for controlling a pollen beetle insect population, the method comprising feeding insects of the population with an agent comprising the dsRNA molecule of  claim 1 . 
     
     
         17 . The method according to  claim 16 , wherein the agent is a sprayable formulation or artificial diet. 
     
     
         18 . A method for controlling a pollen beetle insect population, the method comprising feeding insects of the population with an agent comprising the dsRNA molecule of  claim 4 . 
     
     
         19 . The method according to  claim 18 , wherein the agent is a sprayable formulation or artificial diet. 
     
     
         20 . The method according to  claim 18 , wherein the agent is a transgenic plant material expressing the dsRNA molecule. 
     
     
         21 . A method for improving the yield of a plant crop, the method comprising:
 cultivating in the crop the transgenic plant or seed of  claim 12 , such that the dsRNA molecule is expressed.   
     
     
         22 . The method according to  claim 21 , wherein the crop is canola. 
     
     
         23 . The method according to  claim 21 , wherein the transgenic plant or seed is canola. 
     
     
         24 . The method according to  claim 22 , wherein the transgenic plant or seed is canola. 
     
     
         25 . A method for producing a transgenic plant cell, the method comprising:
 transforming a plant cell with the nucleic acid molecule of  claim 7 ;   culturing the transformed plant cell under conditions sufficient to allow for development of a plant cell culture; and   selecting a transgenic plant cell that has integrated the polynucleotide in its genome and expresses the dsRNA molecule.   
     
     
         26 . A method for producing a pollen beetle-resistant transgenic plant, the method comprising regenerating a transgenic plant from the transgenic plant cell of  claim 10 .