constructs

Abstract

The disclosure relates to promoters, elements associated with promoters, promoter-reporter pairs, and constructs comprising the aforementioned elements suitable for plant transformation.

Description

CONSTRUCTSCROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims priority under 35 U.S.C. § 119(e) to U.S. Provisional Application No. 63 / 600,188, filed November 17, 2023. The contents of this application are incorporated herein by reference in its entirety.SEQUENCE LISTING

[0002] The instant application contains a Sequence Listing which has been submitted in XML format and is hereby incorporated by reference in its entirety. The Sequence Listing, created on November 11 , 2024, is named 16195_0016- 00304_SL.xml and is 157,582 bytes in size.BACKGROUND

[0003] In genetically engineered plants, foreign transgenes are introduced into the host plants by a variety of constructs for expression of the transgenes in those host plant cells. However, due to the interplay between different construct elements and a host plant cell’s expression machinery, a combination of specific regulatory elements with a foreign transgene may not result in the desired expression level of the transgene in the host plant cell. As genetic engineering of plant cells continues to increase in popularity, there is a need for constructs that can achieve high levels of expression of transgenes in the transformed plant cell. The present disclosure addresses this need.SUMMARY

[0004] In some embodiments, the present disclosure provides a nucleic acid molecule, comprising: a) a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to a nucleic acid sequence selected from SEQ ID NOs: 1 -42; and b) a reporter gene, wherein the promoter is operably linked to the reporter gene, such that in response to exposure to a stressor, the promoter induces expression of the reporter gene in a plant. In some embodiments, the nucleic acid molecule further comprises a 5' UTR comprising a nucleic acid sequence selected from SEQ ID NOs: 43-79, wherein the nucleic acidsequence encoding the 5' UTR is operably linked to the nucleic acid sequence encoding the promoter. In some embodiments, the reporter gene is selected from a gene encoding a fluorescent protein, a gene encoding a bioluminescent protein, and a gene encoding a pigment protein. In some embodiments, the reporter gene encodes a fluorescent protein. In some embodiments, the reporter gene encodes bFLO_GFP. In some embodiments, the nucleic acid molecule exhibits gene regulatory activity in a soybean (Glycine max) plant.

[0005] In some embodiments, the nucleic acid molecule comprises a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 4. In some embodiments, the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 46. In some embodiments, the promoter comprises SEQ ID NO: 4. In some embodiments, the promoter comprises SEQ ID NO: 4 and the 5' UTR comprises SEQ ID NO: 46. In some embodiments, the nucleic acid molecule comprises a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 14. In some embodiments, the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 56. In some embodiments, the promoter comprises SEQ ID NO: 14. In some embodiments, the promoter comprises SEQ ID NO: 14 and the 5' UTR comprises SEQ ID NO: 56. In some embodiments, the nucleic acid molecule comprises a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 15. In some embodiments, wherein the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 57. In some embodiments, the promoter comprises SEQ ID NO: 15. In some embodiments, the promoter comprises SEQ ID NO: 15 and the 5' UTR comprises SEQ ID NO: 57. In some embodiments, the nucleic acid molecule comprises a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 16. In some embodiments, the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 58. In some embodiments, the promoter comprises SEQ ID NO: 16. In some embodiments, the promoter comprises SEQ ID NO: 16 and the 5' UTR comprises SEQ ID NO: 58. In some embodiments, the nucleic acid molecule comprises a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 17. In some embodiments, the promoter is operably linked toa 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 59. In some embodiments, the promoter comprises SEQ ID NO: 17. In some embodiments, the promoter comprises SEQ ID NO: 17 and the 5’ UTR comprises SEQ ID NO: 59. In some embodiments, the nucleic acid molecule comprises a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 18. In some embodiments, the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 60. In some embodiments, the promoter comprises SEQ ID NO: 18. In some embodiments, the promoter comprises SEQ ID NO: 18 and the 5' UTR comprises SEQ ID NO: 60. In some embodiments, the nucleic acid molecule comprises a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 19. In some embodiments, the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 61. In some embodiments, the promoter comprises SEQ ID NO: 19. In some embodiments, the promoter comprises SEQ ID NO: 19 and the 5' UTR comprises SEQ ID NO: 61. In some embodiments, the nucleic acid molecule comprises a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 1. In some embodiments, the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 43. In some embodiments, the promoter comprises SEQ ID NO: 1. In some embodiments, the promoter comprises SEQ ID NO: 1 and the 5' UTR comprises SEQ ID NO: 43. In some embodiments, the nucleic acid molecule comprises a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 20. In some embodiments, the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 62. In some embodiments, the promoter comprises SEQ ID NO: 20. In some embodiments, the promoter comprises SEQ ID NO: 20 and the 5' UTR comprises SEQ ID NO: 62. In some embodiments, the nucleic acid molecule comprises a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 21 . In some embodiments, the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 63. In some embodiments, the promoter comprises SEQ ID NO: 21. In some embodiments, the promoter comprises SEQ ID NO: 21 and the 5' UTR comprises SEQ ID NO: 63. In some embodiments, the nucleic acid molecule comprises a promoter comprising a nucleic acid sequence having atleast 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 22. In some embodiments, the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 64. In some embodiments, the promoter comprises SEQ ID NO: 22. In some embodiments, the promoter comprises SEQ ID NO: 22 and the 5' UTR comprises SEQ ID NO: 64. In some embodiments, the nucleic acid molecule comprises a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 23. In some embodiments, the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 65. In some embodiments, the promoter comprises SEQ ID NO: 23. In some embodiments, the promoter comprises SEQ ID NO: 23 and the 5' UTR comprises SEQ ID NO: 65. In some embodiments, the nucleic acid molecule comprises a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 6. In some embodiments, the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 48. In some embodiments, the promoter comprises SEQ ID NO: 6. In some embodiments, the promoter comprises SEQ ID NO: 6 and the 5' UTR comprises SEQ ID NO: 48. In some embodiments, the nucleic acid molecule comprises a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 24. In some embodiments, the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 66. In some embodiments, the promoter comprises SEQ ID NO: 24. In some embodiments, the promoter comprises SEQ ID NO: 24 and the 5' UTR comprises SEQ ID NO: 66.

[0006] In some embodiments, the reporter gene is selected from a gene encoding a fluorescent protein, a gene encoding a bioluminescent protein, and a gene encoding a pigment protein. In some embodiments, the reporter gene encodes a fluorescent protein. In some embodiments, the reporter gene encodes bFLO_GFP. In some embodiments, the nucleic acid molecule exhibits gene regulatory activity in a soybean (Glycine max) plant.

[0007] In some embodiments, the present disclosure provides a binary vector, comprising: a) a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to a nucleic acid sequence selected from SEQ ID NOs: 1 -42; and b) a reporter gene, wherein the promoter is operably linked to the reporter gene, such that in response to exposure to a stressor, thepromoter induces expression of the reporter gene in a plant. In some embodiments, the binary vector further comprises a 5' UTR comprising a nucleic acid sequence selected from SEQ ID NOs: 43-79, wherein the nucleic acid sequence encoding the 5' UTR is operably linked to the nucleic acid sequence encoding the promoter. In some embodiments, the reporter gene is selected from a gene encoding a fluorescent protein, a gene encoding a bioluminescent protein, and a gene encoding a pigment protein. In some embodiments, the reporter gene encodes a fluorescent protein. In some embodiments, the reporter gene encodes bFLO_GFP. In some embodiments, the binary vector exhibits gene regulatory activity in a soybean (Glycine max) plant.

[0008] In some embodiments, the binary vector comprises a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 4. In some embodiments, the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 46. In some embodiments, the promoter comprises SEQ ID NO: 4. In some embodiments, the promoter comprises SEQ ID NO: 4 and the 5' UTR comprises SEQ ID NO: 46. In some embodiments, the binary vector comprises a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 14. In some embodiments, the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 56. In some embodiments, the promoter comprises SEQ ID NO: 14. In some embodiments, the promoter comprises SEQ ID NO: 14 and the 5' UTR comprises SEQ ID NO: 56. In some embodiments, the binary vector comprises a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 15. In some embodiments, the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 57. In some embodiments, the promoter comprises SEQ ID NO: 15. In some embodiments, the promoter comprises SEQ ID NO: 15 and the 5' UTR comprises SEQ ID NO: 57.

[0009] In some embodiments, the binary vector comprises a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 16. In some embodiments, the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 58. In some embodiments, the promoter comprises SEQ ID NO: 16. In some embodiments, the promoter comprises SEQ ID NO: 16 and the 5' UTR comprises SEQ ID NO: 58. In some embodiments, the binary vector comprises a promoter comprisinga nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 17. In some embodiments, the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 59. In some embodiments, the promoter comprises SEQ ID NO: 17. In some embodiments, the promoter comprises SEQ ID NO: 17 and the 5' UTR comprises SEQ ID NO: 59. In some embodiments, the binary vector comprises a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 18. In some embodiments, the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 60. In some embodiments, the promoter comprises SEQ ID NO: 18. In some embodiments, the promoter comprises SEQ ID NO: 18 and the 5' UTR comprises SEQ ID NO: 60. In some embodiments, the binary vector comprises a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 19. In some embodiments, the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 61. In some embodiments, the promoter comprises SEQ ID NO: 19. In some embodiments, the promoter comprises SEQ ID NO: 19 and the 5' UTR comprises SEQ ID NO: 61. In some embodiments, the binary vector comprises a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 1. In some embodiments, the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 43. In some embodiments, the promoter comprises SEQ ID NO: 1. In some embodiments, the promoter comprises SEQ ID NO: 1 and the 5' UTR comprises SEQ ID NO: 43. In some embodiments, the binary vector comprises a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 20. In some embodiments, the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 62. In some embodiments, the promoter comprises SEQ ID NO: 20. In some embodiments, the promoter comprises SEQ ID NO: 20 and the 5' UTR comprises SEQ ID NO: 62. In some embodiments, the binary vector comprises a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 21. In some embodiments, the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 63. In some embodiments, the promoter comprises SEQ ID NO: 21. In some embodiments, the promoter comprises SEQ ID NO: 21 and the 5' UTR comprises SEQ ID NO: 63. Insome embodiments, the binary vector comprises a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 22. In some embodiments, the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 64. In some embodiments, the promoter comprises SEQ ID NO: 22. In some embodiments, the promoter comprises SEQ ID NO: 22 and the 5' UTR comprises SEQ ID NO: 64. In some embodiments, the binary vector comprises a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 23. In some embodiments, the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 65. In some embodiments, the promoter comprises SEQ ID NO: 23. In some embodiments, the promoter comprises SEQ ID NO: 23 and the 5' UTR comprises SEQ ID NO: 65. In some embodiments, the binary vector comprises a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 6. In some embodiments, the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 48. In some embodiments, the promoter comprises SEQ ID NO: 6. In some embodiments, the promoter comprises SEQ ID NO: 6 and the 5' UTR comprises SEQ ID NO: 48. In some embodiments, the binary vector comprises a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 24. In some embodiments, the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 66. In some embodiments, the promoter comprises SEQ ID NO: 24. In some embodiments, the promoter comprises SEQ ID NO: 24 and the 5' UTR comprises SEQ ID NO: 66

[0010] In some embodiments, the reporter gene is selected from a gene encoding a fluorescent protein, a gene encoding a bioluminescent protein, and a gene encoding a pigment protein. In some embodiments, the reporter gene encodes a fluorescent protein. In some embodiments, the reporter gene encodes bFLO_GFP. In some embodiments, the binary vector exhibits gene regulatory activity in a soybean (Glycine max) plant.

[0011] In some embodiments, the present disclosure provides a plant cell comprising a nucleic acid molecule, wherein the nucleic acid molecule comprises: a) a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to a nucleic acid sequence selected from SEQ IDNOs: 1 -42; and b) a reporter gene, wherein the promoter is operably linked to the reporter gene, such that in response to exposure to a stressor, the promoter induces expression of the reporter gene in a plant. In some embodiments, the nucleic acid molecule further comprises a 5' UTR comprising a nucleic acid sequence selected from SEQ ID NOs: 43-79, wherein the nucleic acid sequence encoding the 5' UTR is operably linked to the nucleic acid sequence encoding the promoter. In some embodiments, the reporter gene is selected from a gene encoding a fluorescent protein, a gene encoding a bioluminescent protein, and a gene encoding a pigment protein. In some embodiments, the reporter gene encodes a fluorescent protein. In some embodiments, the reporter gene encodes bFLO_GFP. In some embodiments, the nucleic acid molecule exhibits gene regulatory activity in a soybean (Glycine max) plant.

[0012] In some embodiments, the plant cell comprises a nucleic acid molecule, wherein the nucleic acid molecule comprises a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 4. In some embodiments, the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 46. In some embodiments, the promoter comprises SEQ ID NO: 4. In some embodiments, the promoter comprises SEQ ID NO: 4 and the 5' UTR comprises SEQ ID NO: 46. In some embodiments, the plant cell comprises a nucleic acid molecule, wherein the nucleic acid molecule comprising a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 14. In some embodiments, the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 56. In some embodiments, the promoter comprises SEQ ID NO: 14. In some embodiments, the promoter comprises SEQ ID NO: 14 and the 5' UTR comprises SEQ ID NO: 56. In some embodiments, the plant cell comprises a nucleic acid molecule, wherein the nucleic acid molecule comprises a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 15. In some embodiments, wherein the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 57. In some embodiments, the promoter comprises SEQ ID NO: 15. In some embodiments, the promoter comprises SEQ ID NO: 15 and the 5' UTR comprises SEQ ID NO: 57. In some embodiments, the plant cell comprises a nucleic acid molecule, wherein the nucleic acid molecule comprises a promoter comprising a nucleic acidsequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 16. In some embodiments, the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 58. In some embodiments, the promoter comprises SEQ ID NO: 16. In some embodiments, the promoter comprises SEQ ID NO: 4 and the 5' UTR comprises SEQ ID NO: 58. In some embodiments, the plant cell comprises a nucleic acid molecule, wherein the nucleic acid molecule comprises a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 17. In some embodiments, the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 59. In some embodiments, the promoter comprises SEQ ID NO: 17. In some embodiments, the promoter comprises SEQ ID NO: 17 and the 5' UTR comprises SEQ ID NO: 59. In some embodiments, the plant cell comprises a nucleic acid molecule, wherein the nucleic acid molecule comprises a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 18. In some embodiments, the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 60. In some embodiments, the promoter comprises SEQ ID NO: 18. In some embodiments, the promoter comprises SEQ ID NO: 18 and the 5' UTR comprises SEQ ID NO: 60. In some embodiments, the plant cell comprises a nucleic acid molecule, wherein the nucleic acid molecule comprises a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 19. In some embodiments, the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 61. In some embodiments, the promoter comprises SEQ ID NO: 19. In some embodiments, the promoter comprises SEQ ID NO: 19 and the 5' UTR comprises SEQ ID NO: 61 . In some embodiments, the plant cell comprises a nucleic acid molecule, wherein the nucleic acid molecule comprises a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 1. In some embodiments, the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 43. In some embodiments, the promoter comprises SEQ ID NO: 1. In some embodiments, the promoter comprises SEQ ID NO: 1 and the 5' UTR comprises SEQ ID NO: 43. In some embodiments, the plant cell comprises a nucleic acid molecule, wherein the nucleic acid molecule comprises a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100%identity to SEQ ID NO: 20. In some embodiments, the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 62. In some embodiments, the promoter comprises SEQ ID NO: 20. In some embodiments, the promoter comprises SEQ ID NO: 20 and the 5' UTR comprises SEQ ID NO: 62. In some embodiments, the plant cell comprises a nucleic acid molecule, wherein the nucleic acid molecule comprises a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 21 . In some embodiments, the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 63. In some embodiments, the promoter comprises SEQ ID NO: 21. In some embodiments, the promoter comprises SEQ ID NO: 21 and the 5' UTR comprises SEQ ID NO: 63. In some embodiments, the plant cell comprises a nucleic acid molecule, wherein the nucleic acid molecule comprises a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 22. In some embodiments, the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 64. In some embodiments, the promoter comprises SEQ ID NO: 22. In some embodiments, the promoter comprises SEQ ID NO: 22 and the 5' UTR comprises SEQ ID NO: 64. In some embodiments, the plant cell comprises a nucleic acid molecule, wherein the nucleic acid molecule comprises a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 23. In some embodiments, the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 65. In some embodiments, the promoter comprises SEQ ID NO: 23. In some embodiments, the promoter comprises SEQ ID NO: 23 and the 5' UTR comprises SEQ ID NO: 65. In some embodiments, the plant cell comprises a nucleic acid molecule, wherein the nucleic acid molecule comprises a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 6. In some embodiments, the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 48. In some embodiments, the promoter comprises SEQ ID NO: 6. In some embodiments, the promoter comprises SEQ ID NO: 6 and the 5' UTR comprises SEQ ID NO: 48. In some embodiments, the plant cell comprises a nucleic acid molecule, wherein the nucleic acid molecule comprises a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 24. In some embodiments, the promoter is operably linked toa 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 66. In some embodiments, the promoter comprises SEQ ID NO: 24. In some embodiments, the promoter comprises SEQ ID NO: 24 and the 5' UTR comprises SEQ ID NO: 66. In some embodiments, the reporter gene is selected from a gene encoding a fluorescent protein, a gene encoding a bioluminescent protein, and a gene encoding a pigment protein. In some embodiments, the reporter gene encodes a fluorescent protein. In some embodiments, the reporter gene encodes bFLO_GFP. In some embodiments, the nucleic acid molecule exhibits gene regulatory activity in a soybean (Glycine max) plant.

[0013] In some embodiments, the present disclosure provides a plant, comprising a nucleic acid molecule as described herein, a binary vector as described herein, and / or a plant cell as described herein. In some embodiments, the plant is a soybean (Glycine max) plant.

[0014] In some embodiments, the present disclosure provides a grain, comprising a nucleic acid molecule as described herein, a binary vector as described herein, and / or a plant cell as described herein.BRIEF DESCRIPTION OF THE DRAWING(S)

[0015] FIGS. 1A-1 N depict representative images of plants transformed with promoter-reporter pairs and testing of the expression efficiency of these promoterreporter pairs in response to a fungal stressor at 0 hours and 72 hours after exposure to a fungal stressor.

[0016] FIGS. 2A-2H depict representative images of plants transformed with a promoter-reporter pair comprising GmChitinase and a GmChitinase 5’ UTR and testing of the expression efficiency in response to a fungal stressor at 0 hours and 72 hours after exposure to a fungal stressor.DETAILED DESCRIPTION

[0017] As used herein, “nucleic acid” refers to a polymeric compound including covalently linked nucleotides comprising natural subunits (e.g., purine or pyrimidine bases). In some embodiments, a nucleic acid molecule is a polynucleotide molecule. In some embodiments, a nucleic acid molecule comprises a transgene. Purine bases include adenine and guanine, and pyrimidine bases include uracil, thymine, andcytosine. Nucleic acid molecules include ribonucleic acid (RNA) and deoxyribonucleic acid (DNA), which includes cDNA, genomic DNA, and synthetic DNA, either of which are single- or double-stranded.

[0018] As used herein, the term “DNA” or “DNA molecule” refers to a doublestranded DNA molecule of genomic or synthetic origin, i.e., a polymer of deoxyribonucleotide bases or a polynucleotide molecule, read from the 5' (upstream) end to the 3' (downstream) end. As used herein, the term “DNA sequence” refers to the nucleotide sequence of a DNA molecule.

[0019] As used herein, the term “sequence identity” refers to the extent to which two optimally aligned nucleic acid sequences are identical. An optimal sequence alignment is created by manually aligning two sequences, e.g., a reference sequence and another sequence, to maximize the number of nucleotide-matches in the sequence alignment with appropriate internal nucleotide insertions, deletions, or gaps.

[0020] As used herein, the term “percent sequence identity” or “percent identity” or “% identity” is the identity fraction times 100. The “identity fraction” for a sequence optimally aligned with a reference sequence is the number of nucleotide- matches in the optimal alignment, divided by the total number of nucleotides in the reference sequence, e.g., the total number of nucleotides in the full length of the entire reference sequence.

[0021] The term “transgene” refers to a nucleic acid sequence that has been transferred to a cell (e.g., a plant cell). A transgene comprises nucleic acids, and is, in some embodiments, incorporated into a cell through any of the methods disclosed herein or known to those skilled in the art, including but not limited to, agrobacterium - mediated transformation and transformation through particle bombardment (e.g., using a particle / gene gun).

[0022] As used herein, the term “expression cassette” refers to a distinct component or section of vector DNA consisting of a gene and one or more regulatory elements to be expressed by a transformed cell.

[0023] As used herein, the term “operably linked” refers to the association of two or more nucleic acid molecules on a single nucleic acid fragment so that the function of one is affected by the other.

[0024] As used herein, a “regulatory element” is a DNA molecule having gene regulatory activity, i.e., one that has the ability to affect the transcription and / or translation of an operably linked transcribable nucleic acid molecule. The term “generegulatory activity” thus refers to the ability to affect the expression pattern of a nucleic acid molecule by affecting the transcription and / or translation of that nucleic acid molecule. Isolated regulatory elements, such as promoters and leaders that function in plants are therefore useful for modifying plant phenotypes through the methods of genetic engineering.

[0025] As used herein, the term “promoter” refers generally to a DNA molecule that is involved in recognition and binding of RNA polymerase II and other proteins (trans-acting transcription factors) to initiate transcription.

[0026] As used herein, the term “5' UTR” refers to a DNA molecule that is used during transcription to produce the 5' untranslated region (5' UTR) of an mRNA molecule. A 5' UTR may be operably linked to and located upstream of a nucleic acid molecule and may include polynucleotides that provide regulatory signals, including, but not limited to, signals that are capable of affecting transcription and translation. A 5' UTR may also be referred to as a “leader.”

[0027] As used herein, the term “3' UTR” refers to a DNA molecule that is used during transcription to produce the 3' untranslated region (3' UTR) of an mRNA molecule. A 3' UTR may be operably linked to and located downstream of a nucleic acid molecule and may include polynucleotides that provide a polyadenylation signal and other regulatory signals including, but not limited to, signals that are capable of affecting transcription, mRNA processing, or gene expression.

[0028] As used herein, the term “enhancer” or “enhancer element” refers to a cis-acting transcriptional regulatory element, i.e. , cis-element, which confers an aspect of the overall expression pattern, but is usually insufficient alone to drive transcription, of an operably linked nucleic acid sequence. Unlike promoters, enhancer elements do not usually include a transcription start site (TSS) or TATA box or an equivalent sequence. A promoter may naturally comprise one or more enhancer elements that affect the transcription of an operably linked nucleic acid sequence. An isolated enhancer element may also be fused to a promoter. A promoter may comprise one or more enhancer elements that affect the transcription of operably linked nucleic acid molecules. Enhancers may be positioned upstream or downstream of an operably linked nucleic acid molecule.

[0029] As used herein, the term “operably linked” refers to a first molecule joined to a second molecule, wherein the molecules are so arranged that the first molecule affects the function of the second molecule. The two molecules may or maynot be part of a single contiguous molecule and may or may not be adjacent. For example, in some embodiments, a promoter is operably linked to a nucleic acid molecule such that the promoter modulates transcription of the nucleic acid molecule in a cell.

[0030] As used herein, the term “reporter gene” refers to a gene encoding a reporter protein. The term “reporter protein” refers to a protein that produces a phenotypic change, phenomena, or signal that is ascertainable on the visible light spectra. The reporter protein may effect this phenotypic change, phenomena, or signal through enzymatic activity or other mechanisms known to those skilled in the art.

[0031] As used herein, the term “pigment protein” refers to a protein that is a pigment molecule, as well as proteins that are involved, either directly or indirectly, in the synthesis of pigment molecules. Pigment proteins change the pigment composition within a plant cell.

[0032] As used herein, the term “grain” refers to a fruit or caryopsis that is harvested for human and / or animal consumption. A grain may comprise an attached hull layer, or the hull layer may be detached.

[0033] As used herein, the term “agronomic trait” refers to a characteristic of a plant and / or crop that impacts their yield, quality, and / or ability to resist biotic and abiotic stressors.

[0034] As used herein, the term “construct” means any recombinant nucleic acid molecule such as a plasmid, cosmid, virus, autonomously replicating nucleic acid molecule, phage, or linear or circular single-stranded or double-stranded DNA or RNA polynucleotide molecule, derived from any source, capable of genomic integration or autonomous replication, comprising a nucleic acid molecule where one or more nucleic acid molecule has been linked in a functionally operative manner, i.e., operably linked. As used herein, the term “vector” means any recombinant nucleic acid construct that may be used for the purpose of transformation, i.e., the introduction of heterologous DNA into a host cell. The term includes an expression cassette isolated from any of the aforementioned molecules.

[0035] The term “transformation” refers to the introduction of nucleic acid into a recipient host. As used herein, the term “host” refers to bacteria, fungi, or plant, including any cells, tissue, organs, or progeny of the bacteria, fungi, or plant.

[0036] All publications, patents, and patent applications cited in this specification are incorporated herein by reference in their respective entireties to thesame extent as if each individual publication, patent, or patent application were specifically and individually indicated to be incorporated by reference. Furthermore, each cited publication, patent, or patent application is incorporated herein by reference in its respective entirety to disclose and describe the subject matter in connection with which the publications are cited.

[0037] Before the technology is further described, it is to be understood that this technology is not limited to the particular embodiments described, and as such may, of course, vary. It is also to be understood that the terminology used herein is for the purpose of describing particular embodiments only, and is not intended to be limiting, since the scope of the present disclosure will be limited only by the appended claims. It should also be understood that the headers used herein are not limiting and are merely intended to orient the reader, but the subject matter generally applies to the technology disclosed herein.Promoters

[0038] Transformation of a host cell with a transgene and / or construct does not always result in robust expression of the nucleic acid molecule, and the specific promoter that is operably linked to a nucleic acid molecule (e.g., transgene) can greatly affect the subsequent transcription (expression) of the nucleic acid molecule in a host cell (e.g., transgene). The present disclosure provides a first nucleic acid sequence encoding a promoter, that when operably linked to a second nucleic acid molecule, induces elevated expression of the second nucleic acid molecule in a host cell. In some embodiments, the promoters described herein induce transcription (expression) of a nucleic acid molecule in response to a plant stressor. In some embodiments, the plant stressor is an abiotic stressor. Abiotic stressors include, but are not limited to, environmental factors, such as water, temperature, wind, nutrient deficiency, and salinity. Abiotic stressors may further comprise chemical stressors that are associated with and arise from the application of various chemical treatments. In some embodiments, the plant stressor is a biotic stressor. Biotic stressors include, but are not limited to, insects, bacteria, viruses, fungi, and other plants.

[0039] Exemplary nucleic acid sequences encoding promoters suitable for driving expression of a transgene as described herein are provided in Table 1 .Table 1. Promoter Sequences.

[0040] In some embodiments, a nucleic acid molecule encoding a promoter as described herein comprises a promoter selected from SEQ ID NOs: 1 -42. In some embodiments, a nucleic acid molecule encoding a promoter as described hereincomprises a promoter that has the nucleic acid sequence of SEQ ID NO: 4. In some embodiments, a nucleic acid molecule encoding a promoter as described herein comprises a promoter that has the nucleic acid sequence of SEQ ID NO: 14. In some embodiments, a nucleic acid molecule encoding a promoter as described herein comprises a promoter that has the nucleic acid sequence of SEQ ID NO: 15. In some embodiments, a nucleic acid molecule encoding a promoter as described herein comprises a promoter that has the nucleic acid sequence of SEQ ID NO: 16. In some embodiments, a nucleic acid molecule encoding a promoter as described herein comprises a promoter that has the nucleic acid sequence of SEQ ID NO: 17. In some embodiments, a nucleic acid molecule encoding a promoter as described herein comprises a promoter that has the nucleic acid sequence of SEQ ID NO: 18. In some embodiments, a nucleic acid molecule encoding a promoter as described herein comprises a promoter that has the nucleic acid sequence of SEQ ID NO: 19. In some embodiments, a nucleic acid molecule encoding a promoter as described herein comprises a promoter that has the nucleic acid sequence of SEQ ID NO: 1. In some embodiments, a nucleic acid molecule encoding a promoter as described herein comprises a promoter that has the nucleic acid sequence of SEQ ID NO: 20. In some embodiments, a nucleic acid molecule encoding a promoter as described herein comprises a promoter that has the nucleic acid sequence of SEQ ID NO: 21 . In some embodiments, a nucleic acid molecule encoding a promoter as described herein comprises a promoter that has the nucleic acid sequence of SEQ ID NO: 22. In some embodiments, a nucleic acid molecule encoding a promoter as described herein comprises a promoter that has the nucleic acid sequence of SEQ ID NO: 23. In some embodiments, a nucleic acid molecule encoding a promoter as described herein comprises a promoter that has the nucleic acid sequence of SEQ ID NO: 6. In some embodiments, a nucleic acid molecule encoding a promoter as described herein comprises a promoter that has the nucleic acid sequence of SEQ ID NO: 24.

[0041] In some embodiments, a nucleic acid molecule encoding a promoter as described herein has at least about 85 percent identity, at least about 90 percent identity, at least about 95 percent identity, at least about 96 percent identity, at least about 97 percent identity, at least about 98 percent identity, at least about 99 percent identity, or 100 percent identity to a sequence selected from SEQ ID NOs: 1-42.

[0042] The promoters described herein may be operably linked to one or more additional regulatory elements described herein or known in the art. For example, insome embodiments, a promoter as described herein may be operably linked to an enhancer or a UTR (5' and / or 3'). In some embodiments, a promoter as described herein is operably linked to a 5' UTR. Exemplary nucleic acid sequences encoding 5' UTRs are provided below in Table 2. In some embodiments, a promoter as described herein is operably linked to a 5' UTR having a nucleic acid sequence selected from one of SEQ ID NOs: 43-79. Further, in some embodiments, a promoter as described herein may be operably linked to a transformation booster sequence (TBS) or other element that enhances promoter activity.Table 2. 5' UTR Sequences.

[0043] In some embodiments, a nucleic acid molecule encoding a promoter as described herein comprises the nucleic acid sequence of SEQ ID NO: 4 (GmPR4) operably linked to the nucleic acid sequence of SEQ ID NO:46 (GmPR4 5' UTR). In some embodiments, a nucleic acid molecule encoding a promoter as described herein comprises the nucleic acid sequence of SEQ ID NO: 14 (SolCHI4) operably linked to the nucleic acid sequence of SEQ ID NO:56 (SolCHI4 5' UTR). In some embodiments, a nucleic acid molecule encoding a promoter as described herein comprises the nucleic acid sequence of SEQ ID NO: 15 (GmPR2) operably linked to the nucleic acid sequence of SEQ ID NO: 57 (GmPR2 5' UTR). In some embodiments, a nucleic acid molecule encoding a promoter as described herein comprises the nucleic acid sequence of SEQ ID NO: 16 (GmMFP) operably linked to the nucleic acid sequence of SEQ ID NO: 58 (GmMFP 5' UTR). In some embodiments, a nucleic acid molecule encoding a promoter as described herein comprises the nucleic acid sequence of SEQ ID NO: 17 (GmChitinase) operably linked to the nucleic acid sequence of SEQ ID NO: 59 (GmChitinase 5’ UTR). In some embodiments, a nucleic acid molecule encoding a promoter as described herein comprises the nucleic acid sequence of SEQ ID NO: 18 (GmCHS7) operably linked to the nucleic acid sequence of SEQ ID NO: 60 (GmCHS7 5' UTR). In some embodiments, a nucleic acid molecule encoding a promoter as described herein comprises the nucleic acid sequence of SEQ ID NO: 19 (GmFDH) operably linked to the nucleic acid sequence of SEQ ID NO: 61 (GmFDH 5' UTR). In some embodiments, a nucleic acid molecule encoding a promoter as described herein comprises the nucleic acid sequence of SEQ ID NO: 1 (GmCHIT2) operably linked to the nucleic acid sequence of SEQ ID NO: 43 (GmCHIT2 5' UTR). In some embodiments, a nucleic acid molecule encoding a promoter as described herein comprises the nucleic acid sequence of SEQ ID NO: 20 (GmPI4) operably linked to the nucleic acid sequence of SEQ ID NO: 62 (GmPI4 5' UTR). In some embodiments, a nucleic acid molecule encoding a promoter as described herein comprises the nucleic acid sequence of SEQ ID NO: 21 (GmPR3V2) operably linked to the nucleic acid sequence of SEQ ID NO: 63 (GmPR3V2 5' UTR). In some embodiments, a nucleic acid molecule encoding a promoter as described herein comprises the nucleic acid sequence of SEQ ID NO: 22 (GmPI39) operably linked to the nucleic acid sequence of SEQ ID NO: 64 (GmPI39 5' UTR). In some embodiments, a nucleic acid molecule encoding a promoter as described herein comprises the nucleic acid sequence of SEQ ID NO: 23 (GmPR1 ) operably linked to the nucleic acid sequenceof SEQ ID NO: 65 (GmPR1 5' UTR). In some embodiments, a nucleic acid molecule encoding a promoter as described herein comprises the nucleic acid sequence of SEQ ID NO: 6 (GmPI09) operably linked to the nucleic acid sequence of SEQ ID NO: 48 (GmPI095' UTR). In some embodiments, a nucleic acid molecule encoding a promoter as described herein comprises the nucleic acid sequence of SEQ ID NO: 24 (GmPP012) operably linked to the nucleic acid sequence of SEQ ID NO: 66 (GmPPO12 5' UTR).Reporters

[0044] As described herein, the present disclosure provides a first nucleic acid molecule encoding a promoter that is operably linked to a second nucleic acid molecule. In some embodiments, the second nucleic acid molecule encodes a gene of interest. In some embodiments, the gene of interest is a gene that provides a useful agronomic trait. In some embodiments, the gene of interest providing a useful agronomic trait is a reporter. Reporters are genes that are not normally present in the cell before it is transformed, and include genes that encode for a protein that induces a phenotypic change within the transformed cell. Reporter genes may also, for example, include genes that encode a protein that provide for enzymatic activity within a cell. Reporter genes produce a signal that is detectable to observers of the cell.

[0045] In some embodiments the signal produced by the reporter gene is detectable on a macroscopic scale. In some embodiments, the signal produced by the reporter is detectable on a microscopic scale. Suitable reporter genes are generally known to those skilled in the art, and may include, but are not limited to, genes encoding fluorescent proteins, bioluminescent proteins, and pigment proteins that change the pigment composition of a plant cell such that phenotypic differences in the cell are recognizable. Fluorescent proteins compatible with the constructs described herein include, but are not limited to, green fluorescent protein (GFP), yellow fluorescent protein (YFP), cerulean fluorescent protein (CFP), red fluorescent protein (RFP), orange fluorescent protein (OFP), and variants thereof (e.g., tdTomato, mCherry, DsRed, and bFLO-GFP1 ). Bioluminescent proteins compatible with the constructs described herein include, but are not limited to, photoproteins and luciferases and variants thereof. Photoproteins and luciferases include, but are not limited to, aequorin, berovin, firefly luciferase, oplophorus luciferase, gaussian luciferase, dinoflagellate luciferase, renilla luciferase, and bacterial luciferase and variants thereof. Pigment proteins compatible with the constructs described hereininclude, but are not limited to, anthocyanins, betalains, carotenoids, and variants thereof.

[0046] In addition to the reporters described herein, it is understood that the embodiments described herein may comprise any reporter known in the art.

[0047] The present disclosure further provides for additional regulatory elements that are operably linked to the gene of interest (e.g., reporter gene) and / or a promoter. The additional regulatory elements include those regulatory elements described herein, including but not limited to, enhancers, 5' UTRs, and 3' UTRs.Binary Constructs

[0048] The constructs of the present invention may be provided, in some embodiments, as binary vector constructs comprising a first nucleic acid molecule encoding a promoter that is operably linked to a second nucleic acid molecule as described herein. In some embodiments, the binary construct comprises the right border (RB T-DNA repeat) and left border (LB T-DNA repeat) regions of the tumorinducing plasmid (Ti plasmid) isolated from Agrobacterium tumefaciens comprising a T-DNA, that along with transfer molecules provided by the A. tumefaciens cells, permits the integration of the T-DNA into the genome of a plant cell (see, for example, U.S. Pat. No. 6,603,061 ). The constructs may also comprise plasmid backbone DNA segments generally known in the art that provide replication function and antibiotic selection in bacterial cells, which for example, include an Escherichia coli origin of replication, a broad host range origin of replication such as oriV or oriRi, a coding region for a replication initiator (e.g., RepA), a coding region for a stability protein (e.g., StaA), and a coding region for a selectable marker gene.

[0049] The present disclosure further provides for constructs comprising additional regulatory elements, including but not limited to, promoters, leaders, UTRs, introns, and transcription termination regions.

[0050] Constructs and vectors described herein may further comprise a transit peptide coding sequence that expresses a linked peptide that is useful for transporting a protein product to a particular organelle, particularly to a chloroplast, leucoplast, or other plastid organelle; mitochondria; peroxisome; vacuole; or an extracellular location. Uses of chloroplast transit peptides are described in, for example, U.S. Pat. Nos. 5,188,642 and 5,728,925. Many chloroplast-localized proteins are expressed from nuclear genes as precursors and are transported to the chloroplast by a chloroplast transit peptide (CTP). CTPs and other transit peptides are generally knownin the art and can be used in the constructs described herein to improve transporting an expressed transgene to a specified location within a cell.

[0051] Methods for preparing and using constructs and host cells can be found in, for example, Molecular Cloning: A Laboratory Manual, 3. sup. rd edition Volumes 1 , 2, and 3 (2000) J. F. Sambrook, D. W. Russell, and N. Irwin, Cold Spring Harbor Laboratory Press. Methods for making recombinant vectors particularly suited to plant transformation may include, but are not limited to those described in U.S. Pat. Nos. 4,971 ,908; 4,940,835; 4,769,061 ; and 4,757,011. These types of vectors have also been reviewed in the scientific literature (see, for example, Rodriguez, et al., Vectors: A Survey of Molecular Cloning Vectors and Their Uses, Butterworths, Boston, (1988) and Glick, et al., Methods in Plant Molecular Biology and Biotechnology, CRC Press, Boca Raton, Fla. (1993)). Typical vectors useful for expression of nucleic acids in plants are well known in the art and include, but are not limited to, vectors derived from the Ti plasmid of Agrobacterium tumefaciens (Rogers, et al., Methods in Enzymology 153: 253-277 (1987)).Transformation and Use of Constructs

[0052] The present disclosure is also directed to methods of producing transformed cells and plants which comprise a promoter operably linked to a nucleic acid molecule (e.g., transgene) and / or the constructs described herein.

[0053] In some embodiments, the transformed cell is a soybean (Glycine max) cell. In some embodiments, the transformed plant is a soybean (Glycine max) plant.

[0054] Methods for introducing nucleic acid molecules into host cells generally comprise the steps of selecting a suitable host cell, transforming the host cell with a recombinant vector, and obtaining the transformed host cell. Suitable methods include those described herein, such as bacterial infection (e.g., Agrobacterium), generating binary bacterial artificial chromosome vectors, direct delivery of DNA (e.g., via PEG- mediated transformation, desiccation / inhibition-mediated DNA uptake, electroporation, agitation with silicon carbide fibers, and acceleration of DNA coated particles, etc. (reviewed in Potrykus, et al., Ann. Rev. Plant Physiol. Plant Mol. Biol. 42: 205 (1991 )). Any acceptable method for plant transformation may be utilized to transform a host cell with the transgenes and / or the constructs described herein.

[0055] The transformed plants may be analyzed for the presence of the transgenes and / or the constructs described herein and the expression level and / or profile can be conferred. Those skilled in the art are aware of the numerous methodsavailable for the analysis of transformed plants. For example, methods for plant analysis include, but are not limited to Southern blots or northern blots, PCR-based approaches, biochemical analyses, phenotypic screening methods, field evaluations, and immunodiagnostic assays.

[0056] The seeds of the plants transformed with the transgenes and / or the constructs described herein can be harvested from fertile transgenic plants and be used to grow progeny generations of transformed plants including hybrid plant lines comprising the transgenes and / or the constructs described herein and expressing a gene of agronomic interest.

[0057] Transgenic plants may pass along the transgenes and / or constructs to their progeny. Progeny includes any regenerable plant part or seed comprising the transgenes and / or constructs derived from an ancestor plant. The transgenic plant is preferably homozygous for the transgenes and / or constructs and transmits that nucleic acid molecule to all offspring as a result of sexual reproduction. Progeny may be grown from seeds produced by the transgenic plant. These additional plants may then be self-pollinated to generate a true breeding line of plants. The progeny from these plants are evaluated, among other things, for gene expression. The gene expression may be detected by several common methods such as western blotting, northern blotting, immuno-precipitation, and ELISA.

[0058] The following examples are provided by way of illustration, and are not intended to be limiting of the present disclosure. Those of skill in the art, in light of the present disclosure, should appreciate that many changes can be made in the specific embodiments that are disclosed and still obtain a like or similar result without departing from the spirit and scope of the present disclosure, therefore all matter set forth or shown in the examples and drawings is to be interpreted as illustrative and not in a limiting sense.EXAMPLESExample 1 : Identification of Genes of Interest.

[0059] Publicly available RNA-seq data sets were analyzed to identify genes induced in soy (Glycine max) plants under pathogen stress. Identified genes were ranked based on the fold-change in gene expression induction level in response to pathogen stress. In addition to genes induced in soy, a gene from potato (Solanum tuberosum) was also identified and included in this initial analysis. Genes identified in the analysis are provided in Table 3. Genes with the highest fold-change in responseto pathogen stress are at the top of the table, and genes with the smallest fold-change in response to pathogen stress are at the bottom of the table.Table 3. Genes of nterest.

[0060] Fourteen of the genes in Table 3 were selected, cloned, and transformed into soybean (Glycine max) plants for further testing in disease assays.Example 2: Promoter Testing in Plants

[0061] Testing of the promoters identified in Example 1 was carried out in soybean (Glycine max) plants. The identified promoters were operably linked to reporter genes and incorporated into Ti plasmids as described herein, and plants were transformed with the various constructs using Agrobacterium-mediated transformation as described herein. Soybean plants were grown in 9x4 flats (36 cells) filled with peatbased soil and watered with 0.4 electrical conductivity (EC) fertilizer mix (Jack’s 20- 20-20). Plants were grown under growth chamber conditions with 16-hour / 8-hour lightdark cycles using 400 pmols / m2LED lights, with temperatures of 24°C during days (light) and 22°C during nights (dark). While plants that had at least one trifoliate comprising two-inch long and one-inch-wide leaflets were selected for further testing, plants that appeared stressed or sick (e.g., yellow coloring or small leaves) were excluded from further testing. Six-well assay plates were prepared by inserting pre-cutfloral foam / rockwool pieces into wells and moistening the floral foam / rockwool with enough tap water to create a pool below / around, without covering, the floral foam / rockwool.

[0062] Alternaria alternata, a fungal stressor, was prepared by culturing the fungus on potato dextrose agar (PDA) plates at 27°C in ambient light for up to two weeks prior to transferring to fresh PDA plates or potato dextrose broth (PDB) for inoculum preparation. For liquid fungal growth, 6g of powered PDB was mixed in 250 mL of distilled water and the reconstituted broth was added to a 500 mL wide-mouth media bottle. A cheesecloth piece was folded three times and wedged into the mouth of the bottle to create a tight seal. A piece of aluminum foil was folded three times and formed over the mouth of the bottle to create a cap for the bottle. The bottle was then autoclaved for 20-35 minutes at 121 °C with the exhaust valve closed. The autoclaved media was then inoculated with three to four cm square pieces of plate grown Alternaria by aseptically transferring the cut fungal pieces into the liquid media bottles. The inoculated media bottles were then placed in a shaker at 28°C for five days.

[0063] To prepare the fungal solution for treatment administration, the mycelium was filtered from the liquid culture through four layers of cheesecloth using a Buchner funnel and vacuum pump. The filtered mycelium was rinsed at least five times with sterile distilled water, and, after rinsing, water was removed and depleted from the mycelium by squeezing the mycelium with gloved hands. The mycelium was then weighed and blended in a blender (using the puree setting) for two minutes in enough sterile distilled water to reach a final concentration of 1 % m / v mycelium. Tween was added to the mycelium solution in a quantity sufficient to create a 0.1 % tween:fungal inoculum solution (i.e., volume of tween (pL) = volume of mycelium solution (mL) x 0.001 ). The fungal inoculum solution was stored at 4°C until needed.

[0064] Ethephon treatment for the plants was prepared by adding 300 pL of Florel Brand Growth Regulator (3.9% Ethephon w / v) into a spray bottle with 500 mL DI water, providing a final solution of 0.002% Ethephon. To this solution, 500 pL of Tween-20 was added to give a final Tween-20 concentration of 0.1 %. The solution was stored at room temperature (approximately 22°C) until use. A mock ethephon treatment was prepared by mixing 500 mL DI water with 500 pL Tween-20 (final concentration 0.1 %), and this solution was also stored at room temperature until use.

[0065] To perform the assay, the middle leaflet of the trifoliate leaf in selected plants was cut in two places with a clean pair of scissors. The cut leaf piece was placedadaxial side up in a 6-well tray with the pre-wetted floral foam / rockwool. Each well was treated with the designated treatment by spraying and completely wetting the exposed leaf surface, and then a lid was place on each plate following the treatment. Initial mages were taken (timepoint 0), and plates were placed in a germination chamber set for a constant 23°C and a 16-hour / 8-hour light-dark photoperiod. Plants were subsequently imaged 72 hours after treatment. Images were captured on a custom- built fluorescent imager. Excitation light for green fluorescent protein was created by 4 Series Luxeon Z Blue LEDs and was filtered with a 472 / 36 band pass filter. Images were acquired using a Basler acA3088-57uc camera fitted with a MVL8M23 - 8 mm EFL, f / 1.4, for 2 / 3” C-Mount lens, housing a 520 / 36 band pass filter. Quantitative fluorescence measurements were determined using Imaged.

[0066] Table 4 provides the reporter gene induction following exposure to a fungal stressor as described above, using promoters identified in Example 1. The average final brightness, average raw induction, count of final brightness, max of final brightness, and max of raw induction are reported in Table 4. Raw induction was measured by dividing the brightness recorded in the 72-hour image by the brightness recording in the 0-hour image. Brightness was measured as counts per second (0- 255), and is a measure of the intensity of each of the three color channels (Red, Green, Blue) for each pixel in an image. To calculate the brightness of a leaf, a boundary was drawn around the leaf. The brightest pixels within the boundary, that comprised 10% of the overall pixels within the boundary, were identified and the average brightness of these pixels was calculated. The average was calculated only on the color channel that most closely corresponded to the color of the reporter (e.g., the green channel for bFLO GFP and the red channel for TdTomato RFP). Raw fold change was calculated by dividing the brightness of the final assay timepoint (typically 72 hours after inoculation) by the brightness of the initial timepoint (typically 0 hours after inoculation). Table 4. Efficacy of different promoters on reporter expression in response to

[0067] Surprisingly, several promoters that exhibited a lower fold change in gene expression induction level in Example 1 in response to plant stressors exhibited the highest average brightness measures when the efficacy of constructs comprising these promoters was assayed in plants (see, e.g., GmPPO, GmPR2, GmPR4, and GmCYP93C promoters), indicating that these promoters robustly induced expression of the operably linked reporter gene in soybeans in response to the fungal stressor. FIGs. 1A-1 N show representative images from plants at timepoint 0 and the 72-hour timepoint. FIGS. 1A and 1 B show reporter gene induction when the reporter gene is operably linked to the GmCHIT2 promoter at 0 and 72 hours, respectively. As evident in FIG. 1 B, the GmCHIT2 promoter drove expression of the reporter gene in plants 72 hours post-treatment at a level higher than in plants containing the other tested promoters operably linked to the same report gene. FIGS. 1 C and 1 D show reporter gene induction when the reporter gene is operably linked to the GmPR4 promoter at 0 and 72 hours, respectively. As shown in these figures, plants having the GmPR4 promoter exhibited robust expression of the reporter gene at 72-hours post treatment with the fungal stressor. FIGS. 1 E and 1 F show reporter gene induction when the reporter gene is operably linked to the GmCYP93C promoter at 0 and 72 hours, respectively. The GmCYP93C FIGS. 1 G and 1 H show reporter gene induction when the reporter gene is operably linked to the GmPPO promoter at 0 and 72 hours, respectively. Notably, plants comprising the GmPPO promoter operably linked to the reporter gene exhibited robust expression of the report gene almost immediately following treatment with the fungal stressor, and this expression continued through 72-hours post-treatment. FIGS. 11 and 1 J show reporter gene induction when the reporter gene is operably linked to the SolCHI4 promoter at 0 and 72 hours, respectively. The SolCHI4 promoter drove robust expression of the reporter gene at 72-hours posttreatment with the fungal stressor. FIGS. 1 K and 1 L show reporter gene induction when the reporter gene is operably linked to the GmPR2 promoter at 0 and 72 hours, respectively. The GmPR2 promoter quickly drove expression of the reporter gene, demonstrating marked expression of the reporter gene at the first timepoint, shortly after treatment with the fungal stressor. Further, expression of the reporter gene driven by the GmPR2 promoter increased in intensity by the 72-hour timepoint. FIGS. 1 M and 1 N show reporter gene induction when the reporter gene is operably linked to the GmPI09 promoter at 0 and 72 hours, respectively.

[0068] Without being bound by any particular theory, the promoters described herein appear to be effective drivers of reporter gene expression in soybeans, as demonstrated by elevated brightness and induction levels (see, e.g., GmPR2, GmPR2, SolCHI4). These promoters may therefore function as important regulators of reporter gene expression in response to plant stressors, such as fungal stressors, and may be useful in binary vectors described herein for plant transformation.Example 3: in-vivo Promoter Testing in Soil Grown Plants

[0069] Testing of various promoters in soil grown soybean (Glycine max) plants was conducted. After transformation with plasmids carrying any one of the various promoters described herein, TO plants (also referred to herein as “Event(s)”) were grown to maturity. Six seeds were collected from each event, and seeds were sown into two separate pots. Plants were then grown for 3 weeks to the V1 / V2 stage, in which one pot was treated with Cercospera sojina fungal spores and the other was mock treated according to the methods described herein. Pots were imaged for reporter gene fluorescence at treatment (Oh) and 3 days post-treatment (72h). Fluorescence was quantified from each image. Counts Per Second (CPS) represents pixel intensity values (also referred to herein as “Count(s)”) in an image scaled by the exposure time (in seconds) used for image acquisition. Images were collected in 8-bit images, with a pixel intensity range from 0 to 255 (inclusive of the endpoints). CPS quantifications were determined by dividing the pixel intensity values (counts) by the exposure time (in seconds). Images were acquired in linear mode with no postacquisition scaling. Events were counted as a putative hit if the quantification met the following three criteria: (i) fluorescence values were > CPS measured at 3 days post-treatment; (ii) CPS at 3 days post-fungal treatment were 1.5 times greater than the time of fungal treatment; and (iii) CPS at 3 days post-fungal treatment were 1 .5 times greater than the CPS of mock treated plants at 72 hours post-mock treatment.

[0070] Prior to treatment plants were grown to V1 / V2 in a growth chamber under long days (16 hours light / 6 hours dark, at 24°Celsius). Mock treated plants were treated with water and 0.05% surfactant Tween-20. Fungal treated plants were treated with water, surfactant and Cercospera sojina spores at a concentration of ~150,000 spores / mL.Table 5. Efficacy of different promoters in driving reporter gene fluorescence in soil grown plants in response to a fungal stressor.

[0071] The soil-grown plant data generated and detailed in Table 5 revealed that multiple promoters, both identified in examples 1 and 2 and newly identified in the present example, are particularly effective at meeting the putative hit criteria as described herein. Of note, the GmPR4, GmCHS7, and GmChitinase promoters were among the most effective at meeting the putative hit criteria as described herein. In line with the putative hit data in Table 5, and as evident in FIGs. 2A-2H, plants comprising the GmChitinase promoter (SEQ ID NO: 17) and GmChitinase 5’ UTR (SEQ ID NO: 59) exhibited increased reporting fluorescence between Oh (FIG 2F) and 72h (FIG. 2H) when treated with fungus compared to mock treated plants comprising the GmChitinase promoter (SEQ ID NO: 17) and GmChitinase 5’ UTR (SEQ ID NO: 59) between Oh (FIG. 2B) and 72h (FIG 2D). Thus, the GmChitinase promoter appears to be an effective driver of reporter gene expression and can be used to effectively report fungal presence in soil grown whole plants.

[0072] Without being bound by any particular theory, as indicated by these soil grown plant data, the promoters described herein appear to be effective drivers of reporter gene expression in soybeans, as demonstrated by their ability to meet putative hit criteria levels. These data thus provide further evidence that thesepromoters may therefore function as important regulators of reporter gene expression in response to plant stressors, such as fungal stressors, and may be useful in binary vectors described herein for plant transformation, including but not limited to, their ability to saliently act as reporters in soil grown whole plants.

Claims

CLAIMSWhat is claimed is:1 . A nucleic acid molecule, comprising: a. a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to a nucleic acid sequence selected from SEQ ID NOs: 1 -42; and b. a reporter gene, wherein the promoter is operably linked to the reporter gene, such that in response to exposure to a stressor, the promoter induces expression of the reporter gene in a plant.

2. The nucleic acid molecule of claim 1 , further comprising a 5' UTR comprising a nucleic acid sequence selected from SEQ ID NOs: 43-79, wherein the nucleic acid sequence encoding the 5' UTR is operably linked to the nucleic acid sequence encoding the promoter.

3. The nucleic acid molecule of claim 1 , wherein the reporter gene is selected from a gene encoding a fluorescent protein, a gene encoding a bioluminescent protein, and a gene encoding a pigment protein.

4. The nucleic acid molecule of claim 3, wherein the reporter gene encodes a fluorescent protein.

5. The nucleic acid molecule of claim 4, wherein the reporter gene encodes bFLO_GFP.

6. The nucleic acid molecule of claim 1 , wherein the nucleic acid molecule exhibits gene regulatory activity in a soybean (Glycine max) plant.

7. The nucleic acid molecule of claim 1 , wherein the promoter comprises a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 4.

8. The nucleic acid molecule of claim 7, wherein the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 46.

9. The nucleic acid molecule of claim 7, wherein the promoter comprises SEQ ID NO: 4.

10. The nucleic acid molecule of claim 7, wherein the promoter comprises SEQ ID NO: 4 and is operably linked to a 5' UTR comprising SEQ ID NO: 46.11 . The nucleic acid molecule of claim 7, wherein the reporter gene is selected from a gene encoding a fluorescent protein, a gene encoding a bioluminescent protein, and a gene encoding a pigment protein.

12. The nucleic acid molecule of claim 11 , wherein the reporter gene encodes a fluorescent protein.

13. The nucleic acid molecule of claim 12, wherein the reporter gene encodes bFLO_GFP.

14. The nucleic acid molecule of claim 7, wherein the nucleic acid molecule exhibits gene regulatory activity in a soybean (Glycine max) plant.

15. The nucleic acid molecule of claim 1 , wherein the promoter comprises a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 17.

16. The nucleic acid molecule of claim 15, wherein the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 59.

17. The nucleic acid molecule of claim 15, wherein the promoter comprises SEQID NO: 17.

18. The nucleic acid molecule of claim 15, wherein the promoter comprises SEQ ID NO: 17 and is operably linked to a 5' UTR comprising SEQ ID NO: 59.

19. The nucleic acid molecule of claim 15, wherein the reporter gene is selected from a gene encoding a fluorescent protein, a gene encoding a bioluminescent protein, and a gene encoding a pigment protein.

20. The nucleic acid molecule of claim 19, wherein the reporter gene encodes a fluorescent protein.21 . The nucleic acid molecule of claim 20, wherein the reporter gene encodes bFLO_GFP.

22. The nucleic acid molecule of claim 15, wherein the nucleic acid molecule exhibits gene regulatory activity in a soybean (Glycine max) plant.

23. The nucleic acid molecule of claim 1 , wherein the promoter comprises a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 18.

24. The nucleic acid molecule of claim 23, wherein the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 60.

25. The nucleic acid molecule of claim 23, wherein the promoter comprises SEQ ID NO: 18.

26. The nucleic acid molecule of claim 23, wherein the promoter comprises SEQ ID NO: 18 and is operably linked to a 5' UTR comprising SEQ ID NO: 60.

27. The nucleic acid molecule of claim 23, wherein the reporter gene is selected from a gene encoding a fluorescent protein, a gene encoding a bioluminescent protein, and a gene encoding a pigment protein.

28. The nucleic acid molecule of claim 27, wherein the reporter gene encodes a fluorescent protein.

29. The nucleic acid molecule of claim 28, wherein the reporter gene encodes bFLO_GFP.

30. The nucleic acid molecule of claim 23, wherein the nucleic acid molecule exhibits gene regulatory activity in a soybean (Glycine max) plant.31 . A binary vector for plant transformation, comprising: a nucleic acid sequence encoding i) a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to a nucleic acid sequence selected from SEQ ID NO: 1 -42, and ii) a reporter gene, wherein the promoter is operably linked to the reporter gene and induces expression of the reporter gene in a plant in response to exposure to a stressor.

32. The binary vector of claim 31 , further comprising a 5' UTR comprising a nucleic acid sequence selected from SEQ ID NOs: 43-79, wherein the nucleic acid sequence encoding the 5' UTR is operably linked to the nucleic acid sequence encoding the promoter.

33. The binary vector of claim 31 , wherein the reporter gene is selected from a gene encoding a fluorescent protein, a gene encoding a bioluminescent protein, and a gene encoding a pigment protein.

34. The binary vector of claim 33, wherein the reporter gene encodes a fluorescent protein.

35. The binary vector of claim 34, wherein the reporter gene encodes bFLO_GFP.

36. The binary vector of claim 31 , wherein the nucleic acid molecule exhibits gene regulatory activity in a soybean (Glycine max) plant.

37. The binary vector of claim 31 , wherein the promoter comprises a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 4.

38. The binary vector of claim 37, wherein the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 46.

39. The binary vector of claim 37, wherein the promoter comprises SEQ ID NO: 4.

40. The binary vector of claim 37, wherein the promoter comprises SEQ ID NO: 4 and is operably linked to a 5' UTR comprising SEQ ID NO: 46.41 . The binary vector of claim 37, wherein the reporter gene is selected from a gene encoding a fluorescent protein, a gene encoding a bioluminescent protein, and a gene encoding a pigment protein.

42. The binary vector of claim 41 , wherein the reporter gene encodes a fluorescent protein.

43. The binary vector of claim 42, wherein the reporter gene encodes bFLO_GFP.

44. The binary vector of claim 37, wherein the nucleic acid molecule exhibits gene regulatory activity in a soybean (Glycine max) plant.

45. The binary vector of claim 31 , wherein the promoter comprises a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 17.

46. The binary vector of claim 45, wherein the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 59.

47. The binary vector of claim 45, wherein the promoter comprises SEQ ID NO: 17.

48. The binary vector of claim 45, wherein the promoter comprises SEQ ID NO:17 and is operably linked to a 5' UTR comprising SEQ ID NO: 59.

49. The binary vector of claim 45, wherein the reporter gene is selected from a gene encoding a fluorescent protein, a gene encoding a bioluminescent protein, and a gene encoding a pigment protein.

50. The binary vector of claim 49, wherein the reporter gene encodes a fluorescent protein.51 . The binary vector of claim 50, wherein the reporter gene encodes bFLO_GFP.

52. The binary vector of claim 45, wherein the nucleic acid molecule exhibits gene regulatory activity in a soybean (Glycine max) plant.

53. The binary vector of claim 31 , wherein the promoter comprises a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 18.

54. The binary vector of claim 53, wherein the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 60.

55. The binary vector of claim 53, wherein the promoter comprises SEQ ID NO: 18.

56. The binary vector of claim 53, wherein the promoter comprises SEQ ID NO:18 and is operably linked to a 5' UTR comprising SEQ ID NO: 60.

57. The binary vector of claim 53, wherein the reporter gene is selected from a gene encoding a fluorescent protein, a gene encoding a bioluminescent protein, and a gene encoding a pigment protein.

58. The binary vector of claim 57, wherein the reporter gene encodes a fluorescent protein.

59. The binary vector of claim 58, wherein the reporter gene encodes bFLO_GFP.

60. The binary vector of claim 53, wherein the nucleic acid molecule exhibits gene regulatory activity in a soybean (Glycine max) plant.61 . A plant cell, comprising a nucleic acid molecule comprising: a. a promoter comprising a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to a nucleic acid sequence selected from SEQ ID NOs: 1-42; and b. a reporter gene, wherein the promoter is operably linked to the reporter gene, such that in response to exposure to a stressor, the promoter induces expression of the reporter gene in a plant.

62. The plant cell of claim 61 , further comprising a 5' UTR comprising a nucleic acid sequence selected from SEQ ID NOs: 43-79, wherein the nucleic acid sequence encoding the 5' UTR is operably linked to the nucleic acid sequence encoding the promoter.

63. The plant cell of claim 61 , wherein the reporter gene is selected from a gene encoding a fluorescent protein, a gene encoding a bioluminescent protein, and a gene encoding a pigment protein.

64. The plant cell of claim 63, wherein the reporter gene encodes a fluorescent protein.

65. The plant cell of claim 64, wherein the reporter gene encodes bFLO_GFP.

66. The plant cell of claim 61 , wherein the nucleic acid molecule exhibits gene regulatory activity in a soybean (Glycine max) plant.

67. The plant cell of claim 61 , wherein the promoter comprises a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 4.

68. The plant cell of claim 67, wherein the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 46.

69. The plant cell of claim 67, wherein the promoter comprises SEQ ID NO: 4.

70. The plant cell of claim 67, wherein the promoter comprises SEQ ID NO: 4 and is operably linked to a 5' UTR comprising SEQ ID NO: 46.71 . The plant cell of claim 67, wherein the reporter gene is selected from a gene encoding a fluorescent protein, a gene encoding a bioluminescent protein, and a gene encoding a pigment protein.

72. The plant cell of claim 71 , wherein the reporter gene encodes a fluorescent protein.

73. The plant cell of claim 72, wherein the reporter gene encodes bFLO_GFP.

74. The plant cell of claim 67, wherein the nucleic acid molecule exhibits gene regulatory activity in a soybean (Glycine max) plant.

75. The plant cell of claim 61 , wherein the promoter comprises a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 17.

76. The plant cell of claim 75, wherein the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 59.

77. The plant cell of claim 75, wherein the promoter comprises SEQ ID NO: 17.

78. The plant cell of claim 75, wherein the promoter comprises SEQ ID NO: 17 and is operably linked to a 5' UTR comprising SEQ ID NO: 59.

79. The plant cell of claim 75, wherein the reporter gene is selected from a gene encoding a fluorescent protein, a gene encoding a bioluminescent protein, and a gene encoding a pigment protein.

80. The plant cell of claim 79, wherein the reporter gene encodes a fluorescent protein.81 . The plant cell of claim 80, wherein the reporter gene encodes bFLO_GFP.

82. The plant cell of claim 75, wherein the nucleic acid molecule exhibits gene regulatory activity in a soybean (Glycine max) plant.

83. The plant cell of claim 61 , wherein the promoter comprises a nucleic acid sequence having at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 18.

84. The plant cell of claim 83, wherein the promoter is operably linked to a 5' UTR comprising the nucleic acid sequence of SEQ ID NO: 60.

85. The plant cell of claim 83, wherein the promoter comprises SEQ ID NO: 18.

86. The plant cell of claim 83, wherein the promoter comprises SEQ ID NO: 18 and is operably linked to a 5' UTR comprising SEQ ID NO: 60.

87. The plant cell of claim 83, wherein the reporter gene is selected from a gene encoding a fluorescent protein, a gene encoding a bioluminescent protein, and a gene encoding a pigment protein.

88. The plant cell of claim 87, wherein the reporter gene encodes a fluorescent protein.

89. The plant cell of claim 88, wherein the reporter gene encodes bFLO_GFP.

90. The plant cell of claim 83, wherein the nucleic acid molecule exhibits gene regulatory activity in a soybean (Glycine max) plant.91 . A plant, comprising the nucleic acid molecule of any one of claims 1 -30, the binary vector of any one of claims 31 -60, and / or the plant cell of any one of claims 61-90.

92. The plant of claim 91 , wherein the plant is a soybean (Glycine max) plant.

93. A grain, comprising the nucleic acid molecule of any one of claims 1 -30, the binary vector of any one of claims 31 -60, and / or the plant cell of any one of claims 61-90.

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