Methods to identify evolutionarily significant changes in polynucleotide and polypeptide sequences in domesticated plants and animals
AssigneeEvolutionary Genomics, Inc.
InventorWalter Messier
Life of this patent
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Abstract
The present invention provides methods for identifying polynucleotide and polypeptide sequences which may be associated with commercially or aesthetically relevant traits in domesticated plants or animals. The methods employ comparison of homologous genes from the domesticated organism and its ancestor to identify evolutionarily significant changes and evolutionarily neutral changes. Sequences thus identified may be useful in enhancing commercially or aesthetically desirable traits in domesticated organisms or their wild ancestors.
Claims78
A method for identifying a polynucleotide sequence encoding a polypeptide of a wild ancestor or domesticated organism, wherein said polypeptide is or is suspected of being associated with a enhanced economic productivity in said wild ancestor or domesticated organism, comprising the steps of:
- a) comparing polypeptide-coding nucleotide sequences of said wild ancestor and said domesticated organism;
- and
- b) selecting a polynucleotide sequence in either the domesticated organism or the wild ancestor that contains a nucleotide change as compared to the corresponding sequence in the wild ancestor or domesticated organism, respectively, wherein said change is evolutionarily neutral or positively evolutionarily significant, whereby the polynucleotide which encodes a polypeptide associated with enhanced economic productivity is identified.
A method for identifying a polynucleotide sequence encoding a polypeptide of a wild ancestor of a domesticated organism, wherein said polypeptide is or is suspected of being associated with a stress-resistance trait that is unique, enhanced or altered in the wild ancestor of the domesticated organism as compared to the domesticated organism, comprising the steps of:
- a) comparing polypeptide-coding nucleotide sequences of said domesticated organism to polypeptide-coding nucleotide sequences of said wild ancestor;
- and
- b) selecting a polynucleotide sequence in the wild ancestor that contains a nucleotide change as compared to a corresponding sequence in the domesticated organism, wherein said change is evolutionarily neutral, whereby the wild ancestor's polynucleotide sequence is identified.
A method for identifying an evolutionarily neutral change in a polypeptide-coding polynucleotide sequence of a wild ancestor of a domesticated organism comprising the steps of:
- a) comparing polypeptide-coding polynucleotide sequences of said wild ancestor to corresponding sequences of said domesticated organism;
- and
- b) selecting a polynucleotide sequence in said domesticated organism that contains a nucleotide change as compared to the corresponding sequence of the wild ancestor, wherein the change is evolutionarily neutral and the polynucleotide is associated with a stress-resistance trait in the wild ancestor of the domesticated organism, whereby the evolutionarily neutral change in the polynucleotide is identified.
A method for large scale sequence comparison between polypeptide-coding nucleotide sequences of a wild ancestor of a domesticated organism and polypeptide-coding sequences from said domesticated organism, wherein the wild ancestor organism polypeptide confers or is suspected of conferring a stress-resistance trait that is unique, enhanced or altered in the wild ancestor as compared to the domesticated organism, comprising:
- a) aligning the wild ancestor sequences with corresponding sequences from the domesticated organism according to sequence homology;
- and
- b) identifying any nucleotide changes within the domesticated organism sequences or wild ancestor sequences as compared to the homologous sequences from the wild ancestor or domesticated organism, respectively.
A method for identifying a polynucleotide sequence encoding a polypeptide of a domesticated organism, wherein said polypeptide is or is suspected of being associated with improved yield in said domesticated organism as compared to a wild ancestor of said domesticated organism, comprising the steps of:
- a) comparing polypeptide-coding nucleotide sequences of said domesticated organism to polypeptide-coding nucleotide sequences of said wild ancestor;
- and
- b) selecting a polynucleotide sequence in the domesticated organism that contains a nucleotide change as compared to a corresponding sequence in the wild ancestor, wherein said change is evolutionarily significant, whereby the domesticated organism's polynucleotide sequence is identified.
A method for producing a transfected plant cell or transgenic plant comprising the steps of:
- a) transfecting a plant cell to contain a heterologous DNA segment encoding a polypeptide and derived from an EG307 polynucleotide not native to said cell;
- wherein said polynucleotide is operably linked to a promoter that can be used effectively for expression of transgenic proteins;
- b) optionally growing and maintaining said cell under conditions whereby a transgenic plant is regenerated therefrom;
- c) optionally growing said transgenic plant under conditions whereby said DNA is expressed, whereby the total amount of EG307 polypeptide in said plant is increased.
Plant cells, comprising heterologous DNA encoding an EG307 polypeptide.
A transgenic plant containing heterologous DNA which encodes an EG307 polypeptide that is expressed in plant tissue.
An isolated polynucleotide which includes a promoter operably linked to a polynucleotide that encodes the EG307 gene in plant tissue.
A method of making a transfected cell comprising:
- a) identifying an evolutionarily significant EG307 polynucleotide in an ancestor of a domesticated plant or a corresponding polynucleotide in a domesticated plant;
- b) using said EG307 polynucleotide to identify a non-polypeptide coding sequence that may be a transcription or translation regulatory element, enhancer, intron or other 5′ or 3′ flanking sequence;
- c) assembling a construct comprising said non-polypeptide coding sequence operably linked to a polynucleotide encoding a reporter protein;
- and
- d) transfecting said construct into a host cell.
A transfected host cell comprising a host cell transfected with a construct comprising a promoter, enhancer or intron polynucleotide from an evolutionarily significant EG307 polynucleotide or any combination thereof, operably linked to a polynucleotide encoding a reporter protein.
A method of identifying an agent which may modulate yield, said method comprising contacting at least one candidate agent with a plant or cell comprising an EG307 gene, wherein the agent is identified by its ability to modulate yield.
A method of providing increased yield in a plant comprising:
- a) producing a transfected plant cell having heterologous DNA encoding an EG307 polypeptide whereby EG307 is expressed in said plant cell;
- and
- b) growing a transgenic plant from the transfected plant cell wherein the EG307 transgene is expressed in the transgenic plant.
A method of producing an EG307 polypeptide comprising:
- a) providing a cell transfected with a polynucleotide encoding an EG307 polypeptide positioned for expression in the cell;
- b) culturing the transfected cell under conditions for expressing the polynucleotide;
- and
- c) isolating the EG307 polypeptide.
A method of increasing yield in a plant comprising:
- a) producing a transgenic plant cell having a transgene containing the EG307 gene under the control of a promoter providing constitutive expression of the EG307 gene;
- and
- b) growing a transgenic plant from the transgenic plant cell wherein the EG307 transgene is expressed constitutively in the transgenic plant.
A method of providing controllable yield in a transgenic plant comprising:
- a) producing a transgenic plant cell having a transgene containing the EG307 gene under the control of a promoter providing controllable expression of the EG307 gene;
- and
- b) growing a transgenic plant from the transgenic plant cell wherein the EG307 transgene is controllably expressed in the transgenic plant.
A method of isolating a yield-related gene or fragment thereof from a plant cell, comprising:
- a) providing a sample of plant cell polynucleotides;
- b) providing a pair of oligonucleotides having sequence homology to a conserved region of an EG307 gene;
- c) combining the pair of oligonucleotides with the plant cell polynucleotides sample under conditions suitable for polymerase chain reaction-mediated polynucleotide amplification;
- and
- d) isolating the amplified yield-related polynucleotide or fragment thereof.
A method of isolating a yield-related polynucleotide comprising:
- a) providing a preparation of polynucleotides selected from the group consisting of genomic plant cell DNA and recombinant plant cell library polynucleotides;
- b) contacting the preparation with an EG307 oligonucleotide under hybridization conditions providing detection of polynucleotides having 50% or greater sequence identity;
- and
- c) isolating a yield-related polynucleotide by its association with the EG307 oligonucleotide.
A method of isolating a yield-related gene from plant cell polynucleotides comprising:
- a) providing a sample of plant cell polynucleotides;
- b) providing a pair of oligonucleotides having sequence homology to a conserved region of an EG307 gene;
- c) combining the pair of oligonucleotides with the plant cell polynucleotides under conditions suitable for polymerase chain reaction-mediated polynucleotides amplification;
- and
- d) isolating the amplified yield-related polynucleotide or fragment thereof.
A method of identifying a plant yield-related gene comprising:
- a) providing a plant tissue sample;
- b) introducing into the plant tissue sample a candidate plant yield-related gene;
- c) expressing the candidate plant yield-related gene within the plant tissue sample;
- and
- d) determining whether the plant tissue sample exhibits change in yield response, whereby a change in response identifies a plant yield-related gene.
An isolated polynucleotide selected from the group consisting of:
- a) a polynucleotide selected from the group consisting of SEQ ID NO:
- 1, SEQ ID NO:
- 91, SEQ ID. NO:
- 2, SEQ ID NO:
- 4, SEQ ID NO:
- 5, SEQ ID NO:
- 7, SEQ ID NO:
- 10, SEQ ID NO:
- 11, SEQ ID NO:
- 12, SEQ ID NO:
- 14, SEQ ID NO:
- 15, and SEQ ID NO:
- 17, and SEQ ID NO:
- 18;
- and
- b) a polynucleotide having at least 85% homology to a polynucleotide of a), and which confers substantially the same yield as the polynucleotide of a).
An isolated polynucleotide selected from the group consisting of:
- a) a polynucleotide selected from the group consisting of SEQ ID NO:
- 20, SEQ ID NO:
- 21, SEQ ID. NO:
- 23, SEQ ID NO:
- 24, SEQ ID NO:
- 25, SEQ ID NO:
- 27, SEQ ID NO:
- 28, SEQ ID NO:
- 29, and SEQ ID NO:
- 90;
- and
- b) a polynucleotide having at least 85% homology to a polynucleotide of a), and which confers substantially the same yield as the polynucleotide of a).
An isolated polynucleotide selected from the group consisting of:
- a) a polynucleotide selected from the group consisting of SEQ ID NO:
- 33, SEQ ID NO:
- 34, SEQ ID. NO:
- 35, SEQ ID NO:
- 37, SEQ ID NO:
- 38, SEQ ID NO:
- 40, SEQ ID NO:
- 41, SEQ ID NO:
- 42, SEQ ID NO:
- 44, SEQ ID NO:
- 45, SEQ ID NO:
- 46, SEQ ID NO:
- 47, SEQ ID NO:
- 49, SEQ ID. NO:
- 50, SEQ ID NO:
- 51, SEQ ID NO:
- 53, SEQ ID NO:
- 54, SEQ ID NO:
- 55, SEQ ID NO:
- 57, SEQ ID NO:
- 58, SEQ ID NO:
- 60, SEQ ID NO:
- 62, SEQ ID NO:
- 63, and SEQ ID NO:
- 64;
- and
- b) a polynucleotide having at least 85% homology to a polynucleotide of a), and which confers substantially the same yield as the polynucleotide of a).
An isolated polynucleotide selected from the group consisting of:
- a) a polynucleotide selected from the group consisting of SEQ ID NO:
- 66, SEQ ID NO:
- 67, SEQ ID NO:
- 69, SEQ ID. NO:
- 70, SEQ ID NO:
- 71, SEQ ID NO:
- 73, SEQ ID NO:
- 74, SEQ ID NO:
- 75, SEQ ID NO:
- 77, SEQ ID NO:
- 59, and SEQ ID NO:
- 78;
- and
- b) a polynucleotide having at least 85% homology to a polynucleotide of a), and which confers substantially the same yield as the polynucleotide of a).
An isolated polynucleotide selected from the group consisting of:
- a) a polynucleotide selected from the group consisting of SEQ ID NO:
- 80, SEQ ID NO:
- 81, and SEQ ID NO:
- 82;
- and
- b) a polynucleotide having at least 85% homology to a polynucleotide of a), and which confers substantially the same yield as the polynucleotide of a).
An isolated polynucleotide selected from the group consisting of:
- a) a polynucleotide selected from the group consisting of SEQ ID NO:
- 84 and SEQ ID NO:
- 85;
- and
- b) a polynucleotide having at least 85% homology to a polynucleotide of a), and which confers substantially the same yield as the polynucleotide of a).
An isolated polypeptide selected from the group consisting of:
- a) a polypeptide encoded by a polynucleotide selected from the group consisting of SEQ ID NO:
- 1, SEQ ID NO:
- 91, SEQ ID. NO:
- 2, SEQ ID NO:
- 4, SEQ ID NO:
- 5, SEQ ID NO:
- 7, SEQ ID NO:
- 10, SEQ ID NO:
- 11, SEQ ID NO:
- 12, SEQ ID NO:
- 14, SEQ ID NO:
- 15, and SEQ ID NO:
- 17, and SEQ ID NO:
- 18;
- and
- b) a polypeptide encoded by a polynucleotide having at least 85% homology to a polynucleotide of a), and which confers substantially the same yield as the polynucleotide of a).
An isolated polypeptide selected from the group consisting of:
- a) a polypeptide encoded by a polynucleotide selected from the group consisting of SEQ ID NO:
- 20, SEQ ID NO:
- 21, SEQ ID. NO:
- 23, SEQ ID NO:
- 24, SEQ ID NO:
- 25, SEQ ID NO:
- 27, SEQ ID NO:
- 28, SEQ ID NO:
- 29, and SEQ ID NO:
- 90;
- and
- b) a polypeptide encoded by a polynucleotide having at least 85% homology to a polynucleotide of a), and which confers substantially the same yield as the polynucleotide of a).
An isolated polypeptide selected from the group consisting of:
- a) a polypeptide encoded by a polynucleotide selected from the group consisting of SEQ ID NO:
- 33, SEQ ID NO:
- 34, SEQ ID. NO:
- 35, SEQ ID NO:
- 37, SEQ ID NO:
- 38, SEQ ID NO:
- 40, SEQ ID NO:
- 41, SEQ ID NO:
- 42, SEQ ID NO:
- 44, SEQ ID NO:
- 45, SEQ ID NO:
- 46, SEQ ID NO:
- 47, SEQ ID NO:
- 49, SEQ ID. NO:
- 50, SEQ ID NO:
- 51, SEQ ID NO:
- 53, SEQ ID NO:
- 54, SEQ ID NO:
- 55, SEQ ID NO:
- 57, SEQ ID NO:
- 58, SEQ ID NO:
- 60, SEQ ID NO:
- 62, SEQ ID NO:
- 63, and SEQ ID NO:
- 64;
- and
- b) a polypeptide encoded by a polynucleotide having at least 85% homology to a polynucleotide of a), and which confers substantially the same yield as the polynucleotide of a).
An isolated polypeptide selected from the group consisting of:
- a) a polypeptide encoded by a polynucleotide selected from the group consisting of SEQ ID NO:
- 80, SEQ ID NO:
- 81, and SEQ ID NO:
- 82;
- and
- b) a polypeptide encoded by a polynucleotide having at least 85% homology to a polynucleotide of a), and which confers substantially the same yield as the polynucleotide of a).
An isolated polypeptide selected from the group consisting of:
- a) a polypeptide encoded by a polynucleotide selected from the group consisting of SEQ ID NO:
- 84, SEQ ID NO:
- 85;
- and
- b) a polypeptide encoded by a polynucleotide having at least 85% homology to a polynucleotide of a), and which confers substantially the same yield as the polynucleotide of a).
An isolated polypeptide selected from the group consisting of:
- a) a polypeptide encoded by a polynucleotide selected from the group consisting of SEQ ID NO:
- 66, SEQ ID NO:
- 67, SEQ ID NO:
- 69, SEQ ID. NO:
- 70, SEQ ID NO:
- 71, SEQ ID NO:
- 73, SEQ ID NO:
- 74, SEQ ID NO:
- 75, SEQ ID NO:
- 77, SEQ ID NO:
- 59, and SEQ ID NO:
- 78;
- and
- b) a polypeptide encoded by a polynucleotide having at least 85% homology to a polynucleotide of a).
An isolated polypeptide selected from the group consisting of:
- a) a polypeptide encoded by a polynucleotide selected from the group consisting of SEQ ID NO:
- 84 and SEQ ID NO:
- 85;
- and
- b) a polypeptide encoded by a polynucleotide having at least 85% homology to a polynucleotide of a).
Dependencies are inferred from the claim text.