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US-12618077-B1 - Genetic regulatory elements

US12618077B1US 12618077 B1US12618077 B1US 12618077B1US-12618077-B1

Abstract

Polynucleotides comprising genetic regulatory elements, as well as constructs, host cells, and transgenic organisms comprising the same are described. The polynucleotides can control the expression of an operably linked gene in a host cell or organism, such as a plant cell or a plant. Methods of using the polynucleotide to control the expression of an operably linked gene of interest in a plant or plant cell are further provided.

Inventors

  • Ian W. Davis
  • Tedd D. Elich

Assignees

  • MONSANTO TECHNOLOGY LLC

Dates

Publication Date
20260505
Application Date
20240215

Claims (20)

  1. 1 . A polynucleotide comprising a nucleotide sequence selected from the following, or a single stranded complement thereof: (a) the nucleotide sequence set forth in SEQ ID NO: 10; and (b) a nucleotide sequence having at least 99% sequence identity to the nucleotide sequence set forth in SEQ ID NO: 10, wherein the nucleotide sequence is capable of enhancing the expression of an operably linked gene of interest in a plant.
  2. 2 . The polynucleotide of claim 1 , further comprising a nucleotide sequence that has at least 90% sequence identity to SEQ ID NO: 1, wherein the polynucleotide comprises SEQ ID NO: 39, SEQ ID NO: 40, and SEQ ID NO: 41.
  3. 3 . A construct comprising at least one polynucleotide of claim 1 , and an operably linked transcribable sequence of interest.
  4. 4 . The construct of claim 3 , wherein the construct comprises in the 5′-3′ direction: (a) a promoter sequence, (b) a transcribable sequence of interest; and (c) a gene termination sequence.
  5. 5 . The construct of claim 3 , wherein the transcribable sequence of interest comprises an open reading frame encoding a polypeptide.
  6. 6 . A transgenic plant cell comprising a polynucleotide of claim 1 .
  7. 7 . A plant, plant part, or seed comprising the transgenic cell of claim 6 .
  8. 8 . The plant, plant part, or seed of claim 7 , wherein said polynucleotide is stably incorporated into the genome of the plant, plant part, or seed, and wherein the polynucleotide is operably linked to a transcribable sequence of interest.
  9. 9 . The plant, plant part, or seed of claim 7 , wherein said plant, plant part, or seed is a monocot.
  10. 10 . The plant, plant part, or seed of claim 7 , wherein said plant, plant part, or seed is selected from the group consisting of soybean, cotton, maize, sorghum, wheat, rice, switchgrass, sugarcane, millet, Brachypodium , and Arabidopsis.
  11. 11 . A progeny plant of the plant, plant part, or seed of claim 7 , wherein the progeny plant, plant part, or seed comprises said polynucleotide.
  12. 12 . A method of expressing a gene of interest in a plant or plant cell, said method comprising incorporating into a plant cell, a construct comprising the polynucleotide of claim 1 operably linked to a transcribable nucleotide sequence of interest, wherein the polynucleotide is capable of driving the expression of the operably linked sequence of interest in the plant cell.
  13. 13 . The method of claim 12 , further comprising regenerating a transformed plant from said plant cell.
  14. 14 . The method of claim 12 , wherein said plant cell is stably transformed with said construct.
  15. 15 . The plant, plant part, or seed of claim 7 , wherein said plant, plant part, or seed is dicot.
  16. 16 . The polynucleotide of claim 1 , further comprising a nucleotide sequence that has at least 90% sequence identity to SEQ ID NO: 2, wherein the polynucleotide comprises SEQ ID NO: 39, SEQ ID NO: 40, and SEQ ID NO: 41.
  17. 17 . The polynucleotide of claim 1 , further comprising a nucleotide sequence that has at least 90% sequence identity to SEQ ID NO: 3, wherein the polynucleotide comprises SEQ ID NO: 39, SEQ ID NO: 40, and SEQ ID NO: 41.
  18. 18 . The polynucleotide of claim 1 , further comprising a nucleotide sequence that has at least 90% sequence identity to SEQ ID NO: 4, wherein the polynucleotide comprises SEQ ID NO: 39, SEQ ID NO: 40, and SEQ ID NO: 41.
  19. 19 . The polynucleotide of claim 1 , further comprising a nucleotide sequence that has at least 90% sequence identity to SEQ ID NO: 5, wherein the polynucleotide comprises SEQ ID NO: 39, SEQ ID NO: 40, and SEQ ID NO: 41.
  20. 20 . The polynucleotide of claim 1 , further comprising a nucleotide sequence that has at least 90% sequence identity to SEQ ID NO: 42, wherein the polynucleotide comprises SEQ ID NO: 39, SEQ ID NO: 40, and SEQ ID NO: 41.

Description

CROSS-REFERENCE TO RELATED APPLICATIONS This application is a continuation of U.S. patent application Ser. No. 17/165,734, filed Feb. 2, 2021, which is a continuation of U.S. patent application Ser. No. 13/599,254, filed Aug. 30, 2012, which claims the benefit of U.S. Provisional Patent Application No. 61/529,001, filed Aug. 30, 2011, and U.S. Provisional Patent Application Ser. No. 61/535,109, filed Sep. 15, 2011. The entire disclosure of each of the above applications is incorporated herein by reference. SEQUENCE LISTING This application contains references to amino acid sequences and/or nucleic acid sequences which have been submitted concurrently herewith as the sequence listing .xml file entitled “000103uscob_SequenceListing.xml”, file size 95,522 bytes, created on Feb. 15, 2024. The aforementioned sequence listing is hereby incorporated by reference in its entirety pursuant to 37 C.F.R. § 1.52 (e) (5). TECHNICAL FIELD The invention generally relates to regulatory elements, such as promoters and expression-enhancing introns, and to polynucleotides, cells, and organisms comprising the same. BACKGROUND OF THE INVENTION The production of transgenic cells and organisms comprising a heterologous gene sequence is now routinely practiced by molecular biologists. Methods for incorporating an isolated gene sequence into an expression cassette, producing transformation vectors, and transforming many types of cells and organisms are well known. The regulation or control of expression of the heterologous gene and the protein encoded by the gene can often be critical in the development of a transgenic organism for commercial use. For example, in transgenic plants cells and whole plants comprising a heterologous gene that confers tolerance to herbicide that is normally toxic to the plant, it can be critical to have the heterologous gene expressed in a temporal and spatial manner that corresponds to when the plant is exposed to the herbicide and to what parts of the plant the herbicide normally exerts its phytotoxic effect. A number of genetic regulatory elements are known to play a role in regulating the expression of a gene in plants and other organisms including, for example, promoters, enhancers, 1h5′-untranslated regions (UTRs), 3′-untranslated regions, and expression-enhancing introns. To express a transgene in a plant or organism, one or more of these genetic regulatory elements is operably linked for expression to a nucleic acid sequence or gene of interest. Recently, it has become commonplace to introduce or “stack” multiple transgenes into a single transgenic crop plant. The stacking of multiple transgenes into a single transgenic plant has, however, proved to be problematic, particularly when the same genetic regulatory elements are used in more than one of the stacked transgenes. The use of multiple copies of the same regulatory sequence within two or more transgenes in a single plant is known to promote the activation of gene silencing mechanisms (Halpin (2005) Plant Biotech. J 3:141-155). Silencing of transgenes previously showing stable expression can also be triggered ‘de novo’ when a new transgene is added by crossing or re-transformation if, for example, the same promoter has been used in both transgenes in an effort to promote coordinated expression (Halpin (2005) Plant Biotech. J 3:141-155). Often, the use of the same promoter in multiple transgenes in a single plant is due to the lack of more than one promoter that gives the desired pattern and level of expression. For example, the Cauliflower Mosaic Virus (CaMV) 35S promoter is frequently used as the promoter in plant transgenes because it provides for high-level constitutive expression of an operably linked gene of interest. Because of a lack of suitable alternative promoters, the CaMV 35 promoter is often used to drive the high-level constitutive expression of two or more transgenes in the same plant. Thus, additional promoters and other genetic regulatory elements are needed to avoid gene silencing that might be caused by the use of a particular genetic regulatory element more than once when two, three, four, or more transgenes are stacked in a single crop plant. SUMMARY OF THE INVENTION The present invention provides polynucleotides comprising one or more genetic regulatory elements that can control the expression of an operably linked gene in a host cell or organism, such as a plant cell or a plant. The invention further provides transgenic cells and organisms comprising one or more of these polynucleotides. In particular, the invention provides polynucleotides comprising genetic regulatory elements which may act as promoters and/or expression enhancing introns. For example, polynucleotides having promoter activity may comprise a nucleotide sequence selected from SEQ ID NO: 1 to 5, 15 to 20, and 42 to 50, including fragments, and variants of these sequences. Polynucleotides acting at least in part as expression-enhancing introns include, bu