WO2012009551A1 - Promoteur, éléments de contrôle de promoteur et leurs combinaisons et utilisations - Google Patents

Promoteur, éléments de contrôle de promoteur et leurs combinaisons et utilisations Download PDF

Info

Publication number
WO2012009551A1
WO2012009551A1 PCT/US2011/044033 US2011044033W WO2012009551A1 WO 2012009551 A1 WO2012009551 A1 WO 2012009551A1 US 2011044033 W US2011044033 W US 2011044033W WO 2012009551 A1 WO2012009551 A1 WO 2012009551A1
Authority
WO
WIPO (PCT)
Prior art keywords
motif
seq
plant
nucleic acid
promoter
Prior art date
Application number
PCT/US2011/044033
Other languages
English (en)
Inventor
Michael F. Portereiko
Richard Schneeberger
Smita Sanbui
Tatiana Tatarinova
Yiwen Fang
Nickolai Alexandrov
Kenneth Feldmann
Nestor Apuya
Ke Zhang
Roger I. Pennell
Jack Okamuro
Original Assignee
Ceres, Inc.
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Ceres, Inc. filed Critical Ceres, Inc.
Publication of WO2012009551A1 publication Critical patent/WO2012009551A1/fr

Links

Classifications

    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/415—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from plants
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09—Recombinant DNA-technology
    • C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
    • C12N15/79—Vectors or expression systems specially adapted for eukaryotic hosts
    • C12N15/82—Vectors or expression systems specially adapted for eukaryotic hosts for plant cells, e.g. plant artificial chromosomes (PACs)
    • C12N15/8216—Methods for controlling, regulating or enhancing expression of transgenes in plant cells

Definitions

  • the present invention relates to promoters and promoter control elements that are useful for modulating transcription of a desired polynucleotide.
  • Such promoters and promoter control elements can be included in polynucleotide constructs, expression cassettes, vectors, or inserted into the chromosome or as an exogenous element, to modulate in vivo and in vitro transcription of a polynucleotide.
  • Host cells including plant cells, and organisms, such as regenerated plants therefrom, with desired traits or characteristics using polynucleotides comprising the promoters and promoter control elements described herein are also a part of the invention.
  • the accompanying file named 11696- 0274WOl_sequence_listing_7_2_10.txt was created on July 2, 2010, and is 85.6 KB.
  • the file can be accessed using Microsoft Word on a computer that uses Windows OS.
  • This document relates to promoter sequences and promoter control element sequences which are useful for the transcription of polynucleotides in a host cell or transformed host organism.
  • An introduced gene is generally a chimeric gene composed of the coding region that confers the desired trait and regulatory sequences.
  • One regulatory sequence is the promoter, which is located 5' to the coding region. This sequence is involved in regulating the pattern of expression of a coding region 3' thereof.
  • the promoter sequence binds R A polymerase complex as well as one or more transcription factors that are involved in producing the RNA transcript of the coding region.
  • the promoter region of a gene used in plant transformation is most often derived from a different source than is the coding region. It may be from a different gene of the same species of plant, from a different species of plant, from a plant virus, an algae species, a fungal species, or it may be a composite of different natural and/or synthetic sequences. Properties of the promoter sequence generally determine the pattern of expression for the coding region that is operably linked to the promoter. Promoters with different characteristics of expression have been described.
  • the promoter may confer broad expression as in the case of the widely-used cauliflower mosaic virus (CaMV) 35 S promoter.
  • the promoter may confer tissue-specific expression as in the case of the seed- specific phaseolin promoter.
  • the promoter may confer a pattern for developmental changes in expression.
  • the promoter may be induced by an applied chemical compound, or by an environmental condition applied to the plant.
  • the promoter that is used to regulate a particular coding region is determined by the desired expression pattern for that coding region, which itself is determined by the desired resulting phenotype in the plant. For example, herbicide resistance is desired throughout the plant so the 35 S promoter is appropriate for expression of an herbicide-resistance gene.
  • a seed-specific promoter is appropriate for changing the oil content of soybean seed.
  • An endosperm-specific promoter is appropriate for changing the starch composition of corn seed.
  • a root-specific promoter can be important for improving water or nutrient up-take in a plant. Control of expression of an introduced gene by the promoter is important because it is sometimes detrimental to have expression of an introduced gene in non-target tissues.
  • a gene which induces cell death can be expressed in male and/or female gamete cells in connection with bioconfmement.
  • organisms such as plants, mammals, yeast, and prokaryotes having particular desired characteristics or traits. Examples of these characteristics or traits abound and may include, for example, in plants, virus resistance, insect resistance, herbicide resistance, enhanced stability or additional nutritional value.
  • Recent advances in genetic engineering have enabled researchers in the field to incorporate polynucleotide sequences into host cells to obtain the desired qualities in the organism of choice. This technology permits one or more polynucleotides from a source different than the organism of choice to be transcribed by the organism of choice.
  • the transcription and/or translation of these new polynucleotides can be modulated in the organism to exhibit a desired characteristic or trait.
  • new patterns of transcription and/or translation of polynucleotides endogenous to the organism can be produced.
  • the present document is directed to isolated polynucleotide sequences that comprise promoters and promoter control elements from plants, especially Sorghum bicolor, Panicum virgatum, Oryza sativa, and Arabidopsis thaliana, and other promoters and promoter control elements functional in plants. It is an object of the present document to provide isolated polynucleotides that are promoter or promoter control sequences. These promoter sequences comprise, for example,
  • Promoter or promoter control element sequences of the present document are capable of modulating preferential transcription.
  • this document features an isolated nucleic acid that includes a regulatory region having 90 percent or greater sequence identity (e.g., 95 percent or greater, or 98 percent or greater) to the nucleotide sequence set forth in any one of SEQ ID NOs. 1 - 44 or a fragment thereof, wherein the regulatory region directs transcription of an operably linked heterologous polynucleotide.
  • the regulatory region can consist of any one of the nucleotide sequences set forth in SEQ ID NOs. 1 - 44 or a fragment thereof.
  • the nucleic acid can include one or more motifs selected from the group consisting of an ABRE motif, ABREATCONSENSUS motif, ABREL ATERD 1 motif, ACE motif,
  • ACGTABREMOTIFAOSOSEM motif ACIIPVPAL2 motif, 3-AFl ⁇ binding ⁇ site light responsive element motif, ATCT motif, ATHB2ATCONSENSUS motif,
  • ATHB6COREAT motif CAAT-box motif
  • CACGTGMOTIF motif CACGTGMOTIF motif
  • CACGCAATGMGH3 motif CARGCW8GAT motif, CARGNCAT motif,
  • SORLREP2AT motif SP8BFIBSP8AIB motif, TATABOX motif, TATABOX2 motif, TATABOX4 motif, TATABOX5 motif, TATC-box motif, TCA1 Motif motif, TCA- Element motif, TC richVepeats motif, UP1ATMSD motif, UP2ATMSD motif, and UPRMOTIFIIAT motif.
  • This document also features a vector construct that includes a first nucleic acid that includes a regulatory region having 90 percent or greater sequence identity (e.g., 95 percent or greater, or 98 percent or greater) to any one of SEQ ID NOs. 1 - 44 or a fragment thereof, wherein the regulatory region directs transcription of an operably linked heterologous polynucleotide; and a second nucleic acid to be transcribed, wherein the first and second nucleic acids are heterologous to each other and are operably linked.
  • the first nucleic acid consists of the nucleic acid set forth in any one of SEQ ID NOs: 1-44.
  • the second nucleic acid includes a nucleic acid sequence that encodes a polypeptide.
  • the second nucleic acid can be operably linked to the first nucleic acid in sense orientation.
  • the second nucleic acid can be transcribed into an RNA molecule that expresses the polypeptide encoded by the second nucleic acid.
  • the second nucleic acid can be operably linked to the first nucleic acid in antisense orientation.
  • the second nucleic acid can be transcribed into an antisense R A molecule.
  • the second nucleic acid can be transcribed into an interfering RNA against an endogenous gene.
  • this document features a transgenic plant or plant cell transformed with an isolated nucleic acid described herein that is operably linked to a heterologous polynucleotide, or a vector construct described herein.
  • This document also features seeds of such a plant.
  • the heterologous nucleic acid encodes a polypeptide of agronomic interest.
  • This document also features a method of directing transcription by combining, in an environment suitable for transcription: a first nucleic acid that includes a regulatory region having 90 percent or greater sequence identity (e.g., 95 percent or greater, or 98 percent or greater) to any one of SEQ ID NOs. 1 - 44 or a fragment thereof; and a second nucleic acid to be transcribed; wherein the first and second nucleic acids are heterologous to each other and operably linked.
  • the first nucleic acid molecule can consist of a sequence according to any one of SEQ ID NOs: 1 - 44.
  • the operably linked first and second nucleic acids can be inserted into a plant cell and the plant cell regenerated into a plant.
  • this document features a method of expressing an exogenous coding region in a plant.
  • the method includes transforming a plant cell with a vector described herein; regenerating a stably transformed plant from the transformed plant cell; and selecting plants containing a transformed plant cell, wherein expression of the vector results in production of a polypeptide encoded by the second nucleic acid.
  • This document also features a method of altering the expression of a gene in a plant.
  • the method includes transforming a plant cell with a nucleic acid described herein that is operably linked to a heterologous polynucleotide, and
  • Plants prepared according to such a method also are featured, as well as seeds obtained from such plants.
  • this document features a method of producing a transgenic plant.
  • the method introducing into a plant cell (i) an isolated polynucleotide described herein that is operably linked to a heterologous polynucleotide, or (ii) a vector described herein, and growing a plant from the plant cell.
  • the heterologous polynucleotide described herein that is operably linked to a heterologous polynucleotide, or (ii) a vector described herein
  • polynucleotide can include a nucleic acid sequence encoding a polypeptide.
  • the heterologous polynucleotide can be operably linked to the regulatory region in the antisense orientation.
  • the heterologous polynucleotide can be transcribed into an interfering RNA.
  • the present promoter control elements are capable of serving as or fulfilling the function, for example, as a core promoter, a TATA box, a polymerase binding site, an initiator site, a transcription binding site, an enhancer, an inverted repeat, a locus control region, and/or a scaffold/matrix attachment region.
  • polynucleotide that includes at least a first and a second promoter control element.
  • the first promoter control element is a promoter control element sequence as discussed above, and the second promoter control element is heterologous to the first control element; wherein, the first and second control elements are operably linked.
  • Such promoters may modulate transcript levels preferentially in a particular tissue or under particular conditions.
  • the present isolated polynucleotide comprises a promoter or a promoter control element as described above, wherein the promoter or promoter control element is operably linked to a polynucleotide to be transcribed.
  • the promoter and promoter control elements of the instant document are operably linked to a heterologous polynucleotide that is a regulatory sequence.
  • Host cells include, for instance, bacterial, yeast, insect, mammalian, fungus, algae, and plant.
  • the host cell can comprise a promoter or promoter control element exogenous to the genome. Such a promoter can modulate transcription in cis- and in trans-.
  • the host cell is a plant cell capable of regenerating into a plant.
  • This method comprises providing a polynucleotide or vector according to the present document as described above, and contacting the sample of the polynucleotide or vector with conditions that permit transcription.
  • the polynucleotide or vector preferentially modulates, depending upon the function of the particular promoter, constitutive transcription, stress induced transcription, light induced transcription, dark induced transcription, leaf transcription, root transcription, stem or shoot transcription, silique or fruit transcription, callus transcription, rhizome transcription, stem node transcription, gamete tissue transcription, flower transcription, immature bud or floret and inflorescence specific transcription, senescing induced transcription, germination transcription and/or drought transcription.
  • the Tables consist of the Expression Report(s) for each promoter described herein and provide the nucleotide sequence for each promoter and details for expression driven by each of the nucleic acid promoter sequences as observed in transgenic plants.
  • the results are presented as summaries of the spatial expression, which provides information as to gross and/or specific expression in various plant organs and tissues.
  • the observed expression pattern is also presented, which gives details of expression during different generations or different developmental stages within a generation. Additional information is provided regarding the source organism of the promoter, and the vector and marker genes used for the construct. The following symbols are used consistently throughout the Tables:
  • Figure 1 is a schematic representation of a vector that is useful to insert promoters described herein into a plant.
  • the definitions of the abbreviations used in the vector map are as follows: Ori - the origin of replication used by an E. coli host; RB - sequence for the right border of the T-DNA from pMOG800; Sfil - restriction enzyme cleavage site used for cloning; EGFP - an enhanced version of the green fluorescent protein gene; OCS - the terminator sequence from the octopine synthase gene; p28716 (a.k.a 28716 short) - promoter used to drive expression of the PAT (BAR) gene; PAT (BAR) - a marker gene conferring herbicide resistance; LB - sequence for the left border of the T-DNA from pMOG800; Spec - a marker gene conferring spectinomycin resistance; TrfA - transcription repression factor gene; RK2-OriV - origin of replication for
  • the document disclosed herein provides promoters capable of driving the expression of an operably linked transgene.
  • the design, construction, and use of these promoters is one object of this document.
  • the promoter sequences, SEQ ID NOs: 1 - 44 are capable of transcribing operably linked nucleic acid molecules in particular plant tissues/organs or during particular plant growth stages, and therefore can selectively regulate expression of transgenes in these tissues/organs or at these times of plant development.
  • nucleic acid and “polynucleotide” are used interchangeably herein, and refer to both RNA and DNA, including cDNA, genomic DNA, synthetic DNA, and DNA or RNA containing nucleic acid analogs. Polynucleotides can have any three-dimensional structure. A nucleic acid can be double-stranded or single-stranded, i.e., a sense strand or an antisense strand.
  • Non-limiting examples of polynucleotides include genes, gene fragments, exons, introns, messenger RNA (mRNA), transfer RNA, ribosomal RNA, siRNA, micro-RNA, ribozymes, cDNA, recombinant polynucleotides, branched polynucleotides, plasmids, vectors, isolated DNA of any sequence, isolated RNA of any sequence, nucleic acid probes, and primers, as well as nucleic acid analogs.
  • mRNA messenger RNA
  • transfer RNA transfer RNA
  • ribosomal RNA siRNA
  • micro-RNA micro-RNA
  • ribozymes cDNA
  • recombinant polynucleotides branched polynucleotides
  • plasmids vectors, isolated DNA of any sequence, isolated RNA of any sequence, nucleic acid probes, and primers, as well as nucleic acid analogs.
  • An isolated nucleic acid can be, for example, a naturally-occurring DNA molecule, provided one of the nucleic acid sequences normally found immediately flanking that DNA molecule in a naturally-occurring genome is removed or absent.
  • an isolated nucleic acid includes, without limitation, a DNA molecule that exists as a separate molecule, independent of other sequences, e.g., a chemically synthesized nucleic acid, or a cDNA or genomic DNA fragment produced by the polymerase chain reaction (PCR) or restriction endonuclease treatment.
  • An isolated nucleic acid also refers to a DNA molecule that is incorporated into a vector, an autonomously replicating plasmid, or a virus, or transformed into the genome of a prokaryote or eukaryote.
  • an isolated nucleic acid can include an engineered nucleic acid such as a DNA molecule that is part of a hybrid or fusion nucleic acid.
  • Chimeric is used to describe polynucleotides or genes, or constructs wherein at least two of the elements of the polynucleotide or gene or construct, such as the promoter and the polynucleotide to be transcribed and/or other regulatory sequences and/or filler sequences and/or complements thereof, are heterologous to each other.
  • “broadly expressing promoters” actively promote transcription under most, but not necessarily all, environmental conditions and states of development or cell differentiation.
  • Examples of broadly expressing promoters include the cauliflower mosaic virus (CaMV) 35S transcript initiation region and the 1 ' or 2' promoter derived from T-DNA of
  • Agrobacterium tumefaciens and other transcription initiation regions from various plant genes, such as the maize ubiquitin-1 promoter, known to those of skill.
  • Domains are fingerprints or signatures that can be used to characterize protein families and/or parts of proteins. Such fingerprints or signatures can comprise conserved (1) primary sequence, (2) secondary structure, and/or (3) three- dimensional conformation. A similar analysis can be applied to polynucleotides.
  • each domain has been associated with either a conserved primary sequence or a sequence motif. Generally these conserved primary sequence motifs have been correlated with specific in vitro and/or in vivo activities.
  • a domain can be any length, including the entirety of the polynucleotide to be transcribed. Examples of domains include, without limitation, AP2, helicase, homeobox, zinc finger, etc.
  • Endogenous refers to any polynucleotide, polypeptide or protein sequence which is a natural part of a cell or organism(s) regenerated from said cell.
  • endogenous coding region or “endogenous cDNA” refers to the coding region that is naturally operably linked to the promoter.
  • Enhancer/Suppressor An "enhancer” is a DNA regulatory element that can increase the steady state level of a transcript, usually by increasing the rate of transcription initiation. Enhancers usually exert their effect regardless of the distance, upstream or downstream location, or orientation of the enhancer relative to the start site of transcription.
  • a "suppressor” is a corresponding DNA regulatory element that decreases the steady state level of a transcript, again usually by affecting the rate of transcription initiation.
  • the essential activity of enhancer and suppressor elements is to bind a protein factor(s). Such binding can be assayed, for example, by methods described below. The binding is typically in a manner that influences the steady state level of a transcript in a cell or in an in vitro transcription extract.
  • Exogenous is any polynucleotide, polypeptide or protein sequence, whether chimeric or not, that is introduced into the genome of a host cell or organism regenerated from said host cell by any means other than by a sexual cross. Examples of means by which this can be accomplished are described below, and include Agrobacterium-mediatGd transformation (of dicots - e.g. Salomon et al. (1984) EMBO J. 3 : 141 ; Herrera-Estrella et al. (1983) EMBO J. 2:987; of monocots, representative papers are those by Escudero et al. (1996) Plant J.
  • exogenous nucleic acid is referred to here as a To for the primary transgenic plant and Ti for the first generation.
  • exogenous is also intended to encompass inserting a naturally found element into a non-naturally found location.
  • Heterologous sequences are those that are not operatively linked or are not contiguous to each other in nature.
  • a promoter from corn is considered heterologous to an Arabidopsis coding region sequence.
  • a promoter from a gene encoding a growth factor from corn is considered heterologous to a sequence encoding the corn receptor for the growth factor.
  • Regulatory element sequences such as UTRs or 3 ' end termination sequences that do not originate in nature from the same gene as the coding sequence, are considered heterologous to said coding sequence.
  • homologous polynucleotide refers to a polynucleotide that shares sequence similarity with the polynucleotide of interest. This similarity may be in only a fragment of the sequence and often represents a functional domain such as, examples including, without limitation, a DNA binding domain or a domain with tyrosine kinase activity.
  • inducible promoter in the context of the current document refers to a promoter, the activity of which is influenced by certain conditions, such as light, temperature, chemical concentration, protein concentration, conditions in an organism, cell, or organelle, etc.
  • environmental conditions that may affect transcription by inducible promoters include anaerobic conditions, elevated temperature, the presence or absence of a nutrient or other chemical compound or the presence of light.
  • misexpression refers to an increase or a decrease in the transcription of a coding region into a complementary R A sequence as compared to the wild-type. This term also encompasses expression and/or translation of a gene or coding region or inhibition of such transcription and/or translation for a different time period as compared to the wild-type and/or from a non-natural location within the plant genome, including a gene or coding region from a different plant species or from a non- plant organism.
  • Modulate Transcription Level describes the biological activity of a promoter sequence or promoter control element. Such modulation includes, without limitation, up- and down-regulation of initiation of transcription, rate of transcription, and/or transcription levels.
  • Operable Linkage is a linkage in which a promoter sequence or promoter control element is connected to a polynucleotide sequence (or sequences) in such a way as to place transcription of the polynucleotide sequence under the influence or control of the promoter or promoter control element.
  • Two DNA sequences are said to be operably linked if induction of promoter function results in the transcription of mRNA encoding the polynucleotide and if the nature of the linkage between the two DNA sequences does not (1) result in the introduction of a frame-shift mutation, (2) interfere with the ability of the promoter sequence to direct the expression of the protein, antisense R A, R Ai or ribozyme, or (3) interfere with the ability of the DNA template to be transcribed.
  • a promoter sequence would be operably linked to a polynucleotide sequence if the promoter was capable of effecting transcription of that polynucleotide sequence.
  • Percentage of sequence identity refers to the degree of identity between any given query sequence and a subject sequence.
  • a subject sequence typically has a length that is from about 80 percent to 250 percent of the length of the query sequence, e.g., 82, 85, 87, 89, 90, 93, 95, 97, 99, 100, 105, 110, 115, or 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 percent of the length of the query sequence.
  • a query nucleic acid or amino acid sequence is aligned to one or more subject nucleic acid or amino acid sequences using the computer program ClustalW (version 1.83, default parameters), which allows alignments of nucleic acid or protein sequences to be carried out across their entire length (global alignment). Chenna et al. (2003) Nucleic Acids Res. 31(13):3497-500.
  • ClustalW calculates the best match between a query and one or more subject sequences, and aligns them so that identities, similarities and differences can be determined. Gaps of one or more residues can be inserted into a query sequence, a subject sequence, or both, to maximize sequence alignments. For fast pairwise alignment of nucleic acid sequences, the following default parameters are used: word size: 2;
  • window size 4; scoring method: percentage; number of top diagonals: 4; and gap penalty: 5.
  • gap opening penalty 10.0; gap extension penalty: 5.0; and weight transitions: yes.
  • word size 1; window size: 5; scoring method: percentage; number of top diagonals: 5; gap penalty: 3.
  • weight matrix blosum; gap opening penalty: 10.0; gap extension penalty: 0.05; hydrophilic gaps: on; hydrophilic residues: Gly, Pro, Ser, Asn, Asp, Gin, Glu, Arg, and Lys; residue-specific gap penalties: on.
  • the output is a sequence alignment that reflects the relationship between sequences.
  • ClustalW can be run, for example, at the Baylor College of Medicine Search Launcher website and at the
  • the sequences are aligned using Clustal W and the number of identical matches in the alignment is divided by the query length, and the result is multiplied by 100.
  • the output is the percent identity of the subject sequence with respect to the query sequence. It is noted that the percent identity value can be rounded to the nearest tenth. For example, 78.11, 78.12, 78.13, and 78.14 are rounded down to 78.1, while 78.15, 78.16, 78.17, 78.18, and 78.19 are rounded up to 78.2.
  • Plant Promoter is a promoter capable of initiating transcription in plant cells and can modulate transcription of a polynucleotide. Such promoters need not be of plant origin.
  • promoters derived from plant viruses such as the CaMV35S promoter or from Agrobacterium tumefaciens such as the T-DNA promoters, can be plant promoters.
  • a typical example of a plant promoter of plant origin is the maize ubiquitin-1 (ubi-1) promoter known to those of skill in the art.
  • Plant Tissue includes differentiated and undifferentiated tissues or plants, including but not limited to roots, stems, shoots, rhizomes, cotyledons, epicotyl, hypocotyl, leaves, pollen, seeds, gall tissue and various forms of cells in culture such as single cells, protoplast, embryos, and callus tissue.
  • the plant tissue may be in plants or in organ, tissue or cell culture.
  • Preferential Transcription is defined as transcription that occurs in a particular pattern of cell types or developmental times or in response to specific stimuli or combination thereof.
  • Non-limitive examples of preferential transcription include: high transcript levels of a desired sequence in root tissues; detectable transcript levels of a desired sequence in certain cell types during embryogenesis; and low transcript levels of a desired sequence under drought conditions.
  • Such preferential transcription can be determined by measuring initiation, rate, and/or levels of transcription.
  • promoter is a DNA sequence that directs the transcription of a polynucleotide. Typically a promoter is located in the 5' region of a polynucleotide to be transcribed, proximal to the transcriptional start site of such polynucleotide.
  • promoters are defined as the region upstream of the first exon; more typically, as a region upstream of the first of multiple transcription start sites; more typically, as the region downstream of the preceding gene and upstream of the first of multiple transcription start sites; more typically, the region downstream of the polyA signal and upstream of the first of multiple transcription start sites; even more typically, about 3,000 nucleotides upstream of the ATG of the first exon; even more typically, 2,000 nucleotides upstream of the first of multiple transcription start sites.
  • the promoters of the document comprise at least a core promoter as defined above. Frequently promoters are capable of directing transcription of genes located on each of the complementary DNA strands that are 3' to the promoter.
  • promoters exhibit bidirectionality and can direct transcription of a downstream gene when present in either orientation (i.e. 5' to 3' or 3' to 5' relative to the coding region of the gene).
  • the promoter may also include at least one control element such as an upstream element.
  • control elements include UARs and optionally, other DNA sequences that affect transcription of a polynucleotide such as a synthetic upstream element.
  • Promoter Control Element describes elements that influence the activity of the promoter.
  • Promoter control elements include transcriptional regulatory sequence determinants such as, but not limited to, enhancers, scaffold/matrix attachment regions, TATA boxes, transcription start locus control regions, UARs, URRs, other transcription factor binding sites and inverted repeats.
  • Public sequence refers to any sequence that has been deposited in a publicly accessible database prior to the filing date of the present application. This term encompasses both amino acid and nucleotide sequences. Such sequences are publicly accessible, for example, on the BLAST databases on the NCBI FTP web site (accessible via the internet). The database at the NCBI FTP site utilizes "gi" numbers assigned by NCBI as a unique identifier for each sequence in the databases, thereby providing a non- redundant database for sequence from various databases, including GenBank, EMBL, DBBJ (DNA Database of Japan) and PDB (Brookhaven Protein Data Bank).
  • regulatory region refers to nucleotide sequences that, when operably linked to a sequence, influence transcription initiation or translation initiation or transcription termination of said sequence and the rate of said processes, and/or stability and/or mobility of a transcription or translation product.
  • operably linked refers to positioning of a regulatory region and said sequence to enable said influence.
  • Regulatory regions include, without limitation, promoter sequences, enhancer sequences, response elements, protein recognition sites, inducible elements, protein binding sequences, 5' and 3' untranslated regions (UTRs), transcriptional start sites, termination sequences, polyadenylation sequences, and introns. Regulatory regions can be classified in two categories, promoters and other regulatory regions.
  • regulatory sequence refers to any nucleotide sequence that influences transcription or translation initiation and rate, or stability and/or mobility of a transcript or polypeptide product. Regulatory sequences include, but are not limited to, promoters, promoter control elements, protein binding sequences, 5' and 3' UTRs, transcriptional start sites, termination sequences, polyadenylation sequences, introns, certain sequences within amino acid coding sequences such as secretory signals, protease cleavage sites, etc.
  • Specific promoters refers to a subset of promoters that have a high preference for modulating transcript levels in a specific tissue, or organ or cell and/or at a specific time during development of an organism.
  • high preference is meant at least 3-fold, preferably 5- fold, more preferably at least 10-fold still more preferably at least 20-fold, 50-fold or 100-fold increase in transcript levels under the specific condition over the transcription under any other reference condition considered.
  • Typical examples of temporal and/or tissue or organ specific promoters of plant origin that can be used with the polynucleotides of the present document, are: PTA29, a promoter which is capable of driving gene transcription specifically in tapetum and only during anther development (Koltonow et al. (1990) Plant Cell 2: 1201; RCc2 and RCc3, promoters that direct root-specific gene transcription in rice (Xu et al. (1995) Plant Mol. Biol. 2 ⁇ 2 ⁇ > ⁇ ; TobRB27, a root-specific promoter from tobacco (Yamamoto et al. (1991) Plant Cell 3:371).
  • tissue-specific promoters under developmental control include promoters that initiate transcription only in certain tissues or organs, such as root, ovule, fruit, seeds, or flowers.
  • Other specific promoters include those from genes encoding seed storage proteins or the lipid body membrane protein, oleosin. A few root-specific promoters are noted above. See also "Preferential transcription.”
  • Stringency is a function of nucleic acid molecule probe length, nucleic acid molecule probe composition (G + C content), salt concentration, organic solvent concentration and temperature of hybridization and/or wash conditions. Stringency is typically measured by the parameter T m , which is the temperature at which 50% of the complementary nucleic acid molecules in the hybridization assay are hybridized, in terms of a temperature differential from T m . High stringency conditions are those providing a condition of T m - 5°C to T m - 10°C. Medium or moderate stringency conditions are those providing T m - 20°C to T m - 29°C. Low stringency conditions are those providing a condition of T m - 40°C to T m - 48°C. The relationship between hybridization conditions and T m (in °C) is expressed in the mathematical equation:
  • N is the number of nucleotides of the nucleic acid molecule probe.
  • T m 81.5+16.6 log ⁇ [Na + ]/(l+0.7[Na + ]) ⁇ + 0.41(%G+C)-500/L 0.63(%formamide) (II)
  • L represents the number of nucleotides in the probe in the hybrid
  • T m of Equation II is affected by the nature of the hybrid: for DNA-R A hybrids, T m is 10-15 °C higher than calculated; for RNA-RNA hybrids, T m is 20-25 °C higher. Because the T m decreases about 1°C for each 1% decrease in homology when a long probe is used (Frischauf et al. (1983) J. Mol Biol, 170: 827-842), stringency conditions can be adjusted to favor detection of identical genes or related family members.
  • Equation II is derived assuming the reaction is at equilibrium.
  • hybridizations according to the present document are most preferably performed under conditions of probe excess and allowing sufficient time to achieve equilibrium.
  • the time required to reach equilibrium can be shortened by using a hybridization buffer that includes a hybridization accelerator such as dextran sulfate or another high volume polymer.
  • Stringency can be controlled during the hybridization reaction, or after hybridization has occurred, by altering the salt and temperature conditions of the wash solutions.
  • the formulas shown above are equally valid when used to compute the stringency of a wash solution.
  • Preferred wash solution stringencies lie within the ranges stated above; high stringency is 5-8°C below T m , medium or moderate stringency is 26- 29°C below T m and low stringency is 45-48°C below T m .
  • T 0 refers to the whole plant, explant or callus tissue, inoculated with the transformation medium.
  • Ti refers to either the progeny of the T 0 plant, in the case of whole-plant transformation, or the regenerated seedling in the case of explant or callous tissue transformation.
  • T 2 refers to the progeny of the Ti plant. T 2 progeny are the result of self-fertilization or cross-pollination of a Ti plant.
  • T3 refers to second generation progeny of the plant that is the direct result of a transformation experiment. T3 progeny are the result of self- fertilization or cross-pollination of a T 2 plant.
  • TATA to start shall mean the distance, in number of nucleotides, between the primary TATA motif and the start of transcription.
  • Transgenic plant is a plant having one or more plant cells that contain at least one exogenous polynucleotide introduced by recombinant nucleic acid methods.
  • translational start site is usually an ATG or AUG in a transcript, often the first ATG or AUG.
  • a single protein encoding transcript may have multiple translational start sites.
  • Transcription start site is used in the current document to describe the point at which transcription is initiated. This point is typically located about 25 nucleotides downstream from a TFIID binding site, such as a TATA box. Transcription can initiate at one or more sites within the gene, and a single polynucleotide to be transcribed may have multiple transcriptional start sites, some of which may be specific for transcription in a particular cell-type or tissue or organ. is stated relative to the transcription start site and indicates the first nucleotide in a transcript.
  • Upstream Activating Region (UAR): An "Upstream Activating
  • Region is a position or orientation dependent nucleic acid element that primarily directs tissue, organ, cell type, or environmental regulation of transcript level, usually by affecting the rate of transcription initiation.
  • Corresponding DNA elements that have a transcription inhibitory effect are called herein "Upstream Repressor
  • Regions or "URR”s.
  • the essential activity of these elements is to bind a protein factor. Such binding can be assayed by methods described below. The binding is typically in a manner that influences the steady state level of a transcript in a cell or in vitro transcription extract.
  • Untranslated region UTR
  • UTR Untranslated region
  • a "UTR” is any contiguous series of nucleotide bases that is transcribed, but is not translated.
  • a 5' UTR lies between the start site of the transcript and the translation initiation codon (ATG codon) and includes the +1 nucleotide of the messenger RNA or cDNA.
  • a "plant 5' UTR” can be a native or non-native 5 ' UTR that is functional in plant cells.
  • a 5 ' UTR can be used as a 5' regulatory element for modulating expression of an operably linked transcribable polynucleotide molecule.
  • 5' UTRs derived from heat shock protein genes have been demonstrated to enhance gene expression in plants (see for example, U.S. Pat. No. 5,659,122 and U.S. Pat. No. 5,362,865, all of which are incorporated herein by reference).
  • Examples of 5' UTRs include those shown in SEQ ID NOs: l-7, 9-12, 16-18.
  • a 3' UTR lies between the translation termination codon and the end of the transcript. UTRs can have particular functions such as increasing mRNA message stability or translation attenuation. Examples of 3' UTRs include, but are not limited to polyadenylation signals and transcription termination sequences.
  • the promoters and promoter control elements of this document are capable of modulating transcription. Such promoters and promoter control elements can be used in combination with native or heterologous promoter fragments, control elements or other regulatory sequences to modulate transcription and/or translation.
  • promoters and control elements of the document can be used to modulate transcription of a desired polynucleotide, which includes without limitation:
  • the promoter also can modulate transcription in a host genome in cis- or in trans-.
  • the promoters and promoter control elements of the instant document are useful to produce preferential transcription which results in a desired pattern of transcript levels in a particular cells, tissues, or organs, or under particular conditions.
  • the promoters and promoter control elements of the present document are presented in the Promoter Reports of the Tables and were identified from Sorghum bicolor, Panicum virgatum, Oryza sativa, and Arabidopsis thaliana.
  • Isolation from genomic libraries of polynucleotides comprising the sequences of the promoters and promoter control elements of the present document is possible using known techniques. For example, polymerase chain reaction (PCR) can amplify the desired polynucleotides utilizing primers designed from SEQ ID NOs: 1 - 44.
  • Polynucleotide libraries comprising genomic sequences can be constructed according to Sambrook et al.,
  • RACE amplification of cDNA ends
  • promoters and promoter control elements described in the Promoter Reports in the Tables can be chemically synthesized according to techniques in common use. See, for example, Beaucage et al. (1981) Tet. Lett. 22: 1859 and U.S. Pat. No. 4,668,777. Such chemical oligonucleotide synthesis can be carried out using commercially available devices, such as, Biosearch 4600 or 8600 DNA synthesizer, by Applied Biosystems, a division of Perkin-Elmer Corp., Foster City, California, USA; and Expedite by Perceptive Biosystems, Framingham, Massachusetts, USA.
  • promoters exhibiting nucleotide sequence identity to SEQ. ID. Nos. 1 - 44.
  • promoters of this document can exhibit at least 80% sequence identity (e.g., at least 85%>, at least 90%>, at least 95%>, 96%>, 97%), 98%o or 99%> sequence identity) compared to the nucleotide sequence set forth in any one of SEQ. ID. Nos. 1 - 44.
  • Sequence identity can be calculated by the algorithms and computers programs described above.
  • promoters described herein also can be a fragment of any one of SEQ ID NO: 1 - 44 as long as the fragment retains the ability to direct transcription of a polynucleotide.
  • Suitable fragments can be, for example at least 80% (e.g., at least 85, 90, 95, 96, 97, 98, or 99%) of the length of the nucleotide sequence set forth in any one of SEQ ID NOs: 1 - 44.
  • a regulatory region can be a fragment of anyone of SEQ ID NOs: 1-44 that is 100, 200, 300, 400, 500, 600, 700, 800, 900, 1000, 1100, 1200, 1300, 1400, 1500, 1600, 1700, 1800, 1900, 2000, 2100, 2200, or 2400 nucleotides in length that retains the ability to direct expression of an operably linked nucleic acid.
  • a regulatory region can contain conserved regulatory motifs. Such a regulatory region can be have a nucleotide sequence set forth in anyone of SEQ ID NOs: 1-44, or a regulatory region having a nucleotide sequence that deviates from that set forth in SEQ ID NOs: 1-44, while retaining the ability to direct expression of an operably linked nucleic acid.
  • a regulatory region can contain a CAAT box or a TATA box.
  • a CAAT box is a conserved nucleotide sequence involved in initiation of transcription.
  • a CAAT box functions as a recognition and binding site for regulatory proteins called transcription factors.
  • a TATA box is another conserved nucleotide sequence involved in transcription initiation. A TATA box seems to be important in determining accurately the position at which transcription is initiated.
  • a regulatory region can be analyzed using the PLACE (PLAnt Cis-acting regulatory DNA Elements) Web Signal Scan program on the world wide web at dna.affrc.go.jp/PLACE/signalscan.html. See, Higo et al., Nucleic Acids Research, 27(l):297-300 (1999); and Prestridge, CABIOS, 7:203-206 (1991). Examples of conserved regulatory motifs can be found in the PLACE database on the world wide web at dna.affrc.go.jp/PLACE/. See, Higo et al, supra.
  • SEQ ID NOs: 1-44 or a regulatory region having a nucleotide sequence that deviates from that set forth in SEQ ID NOs: 1-44, while retaining the ability to direct expression of an operably linked nucleic acid, can contain one or more conserved regulatory motifs, which can be found in the PLACE database.
  • a regulatory region can have an ABA-responsive element such as an ABREATCONSENSUS motif having the consensus sequence (C/T)ACGTGGC. See, Choi et al., J Biol Chem. 275: 1723-1730 (2000).
  • a regulatory region can contain an ABRE motif having the consensus sequence ACGTG or an ABRELATRDl motif having the consensus sequence CGMCACGTGB (SEQ ID NO: 45).
  • a regulatory region can contain an ABREATRD22 motif having the consensus sequence RYACGTGGYR (SEQ ID NO:46). See, Iwasaki et al., Mol Gen Genet 247:391-398 (1995); Bray, Trends Plant Sci. 2:48-54 (1997); Busk and Pages, Plant Mol Biol 37:425-435 (1998).
  • a regulatory region can contain an ABRERATCAL motif having the consensus sequence MACGYGB. See, Kaplan et al., Plant Cell. 18:2733-2748 (2006).
  • a regulatory region can contain an ABREZMRAB28 motif having the consensus sequence CCACGTGG. See, Suzuki et al., Plant
  • a regulatory region can contain an ACGTABREMOTIFA20SEM motif having the consensus sequence ACGTGKC. See, Hattori et al, Plant Cell Physiol 43: 136-140 (2002); and Narusaka et al. , Plant J. 34: 137-148 (2003).
  • a regulatory region can have an ACE motif having the consensus sequence GACACGTAGA. See, Hartmann et al, Plant Mol Biol 36: 741-754 (1998).
  • a regulatory region can contain an ACIIPVPAL2 motif having the consensus sequence CCACCAACCCCC (SEQ ID NO: 47). See, Patzlaff et al, Plant Mol Biol. 53:597-608 (2003); Hatton et al, Plant J 7:859-876 (1995); and Gomez-Maldonado et al, Plant J. 39:513-526 (2004).
  • a regulatory region can contain an 3-AFl ⁇ binding ⁇ site light responsive element having the consensus sequence AAATAGATAAATAAAAACATT.
  • a regulatory region can contain an AMYBOX1 motif having the consensus sequence TAACARA. See, Huang et al, Plant Mol Biol 14:655-668 (1990).
  • a regulatory region can contain an ARE1 motif having the consensus sequence
  • a regulatory region can contain an ATCT motif having the consensus sequence AATCT(A/G)ATCB (SEQ ID NO: 49).
  • a regulatory region can contain an ATHB1ATCONSENSUS motif having the consensus sequence CAATWATTG. See, Sessa et al., EMBO J 12:3507-3517 (1993).
  • a regulatory region can contain an
  • ATHB6COREAT motif having the consensus sequence CAATTATTA. See,
  • a regulatory region can contain an AUXRETGA2GMGH3 motif having the consensus sequence TGACGTAA. See, Liu et al, Plant Cell 6:645-657 (1994); Liu et al, Plant Physiol 115:397-407 (1997); and Guilfoyle et al, Plant Physiol 118: 341-347 (1998).
  • a regulatory region can contain a BOXllPCCHS motif having the consensus sequence ACGTGGC.
  • a regulatory region can contain a CAATBOX1 motif having the consensus sequence CCAAT. See, Shirsat et al, Mol Gen Genet 215:326-331 (1989). A regulatory region can contain a CACGCAATGMGH3 motif having the consensus sequence CACGCAAT. See, Ulmasov et al, Plant Cell 7: 1611-1623 (1995).
  • a regulatory region can contain a CACGTGMOTIF motif having the consensus sequence CACGTG. See, Hudson and Quail, Plant Physiol 133: 1605-1616 (2003).
  • a regulatory region can contain a CARGCW8GAT motif having the consensus sequence CWWWWWWWG (SEQ ID NO: 50). See, Tang and Perry, J Biol C/zem.278:28154-28159 (2003); Folter and Angenent, Trends Plant Sci. 11 :224-231 (2006).
  • a regulatory region can contain a CARGNCAT motif having the consensus sequence CCWWWWWWWWGG (SEQ ID NO: 51). See Wang et al, Plant Cell, 16: 1206-1219 (2004).
  • a regulatory region can contain a CACGTGMOTIF motif having the consensus sequence CACGTG. See, Hudson and Quail, Plant Physiol. 133: 1605-1616 (2003).
  • a regulatory region can contain a CCA1ATLHCB1 motif having the consensus sequence AAMAATCT. See, Wang et al, Plant Cell 9:491-507 (1997).
  • a regulatory region can contain a
  • a regulatory region can contain a CTRMCAMV35S motif having the consensus sequence TCTCTCTCT.
  • a regulatory region can contain an E2FAT motif having the consensus sequence TYTCCCGCC. See, Ramirez-Parra et al, Plant J. 33: 801-811 (2003).
  • a regulatory region can contain an E2FCONSENSUS motif having the consensus sequence WTTSSCSS. See, Vandepoele et al, Plant Physiol.139: 316-328 (2005).
  • a regulatory region can contain an ELI -box 3 motif having the consensus sequence AAACCAATT in any orientation. See, Lois et al.
  • a regulatory region can contain an ERELEE4 motif having the consensus sequence AWTTCAAA. See, Itzhaki et al, Proc Natl Acad Sci USA 91 :8925-8929 (1994); Montgomery et al, Proc Natl Acad Sci USA 90:5939-5943 (1993); Tapia et al, Plant Physiol. 138:2075-2086 (2005); and Rawat et al, Plant Mol Biol. 57: 629-643 (2005).
  • a regulatory region can contain a GADOWNAT motif having the consensus sequence ACGTGTC. See, Ogawa et al, Plant Cell 15: 1591-1604 (2003); and
  • a regulatory region can contain a GADOWNAT motif having the consensus sequence ACGTGTC. See Sangwan and O'Brian, Plant Physiol. 129: 1788-1794 (2002).
  • a regulatory region can contain a GAGAGMGSA1 motif having the consensus sequence GAGAGAGAGAGAGAGA (SEQ ID NO: 52). See, Sutoh and Yamauchi, Plant J. 34: 636-645 (2003).
  • a regulatory region can contain a GARE20SREP1 motif having the consensus sequence TAACGTA. Sutoh and Yamauchi, Plant J. 34: 636-645 (2003).
  • a regulatory region can contain a GAREAT motif having the consensus sequence TAACAAR.
  • a regulatory region can contain a GATABox motif having the consensus sequence GATAGGA. See, Mongkolsiriwatana et al, KasetsartJ. (Nat. Sci.) 43 : 164 - 177 (2009).
  • a regulatory region can contain a GCCYBox motif having the consensus sequence TAAGAGCCGCC. See, Yamamoto et al, Plant J 20:571-579 (1999).
  • a regulatory region can contain a GT1MOTIFPSRBCS motif having the consensus sequence KWGTGRWAAWRW. See, Villain et al, J Biol Chem 271 :32593- 32598 (1996).
  • a regulatory region can contain a HSE motif having the consensus sequence AGAANNTTCT. See, Nover et al, Cell Stress Chaperones. 6(3): 177-89 (2001).
  • a regulatory region can contain an IBOX motif having the consensus sequence GATAAG. See, Giuliano et al, Proc Natl Acad Sci USA 85:7089-7093 (1988).
  • a regulatory region can contain an IBOXCORENT motif having the consensus sequence GATAAGR. See, Martinez-Hernandez et al, Plant Physiol 128: 1223-1233 (2002).
  • a regulatory region can contain an INRNTPSADB motif having the consensus sequence YTCANTYY. See, Nakamura et a/., /antJ29: 1-10 (2002).
  • a regulatory region can contain a LEAFY ATAG motif having the consensus sequence CCAATGT.
  • a regulatory region can contain a LRENPCABE motif having the consensus sequence ACGTGGCA. See, Castresana et al, EMBO J 1:1929-1936 (1988).
  • a regulatory region can contain a MARTBOX motif having the consensus sequence TTWTWTTWTT (SEQ ID NO: 53). See, Gasser et al, Int Rev Cyto 119:57-96 (1989).
  • a regulatory region can contain a MBS motif having the consensus sequence TAACTG. See, Mongkolsiriwatana et al, Kasetsart J. (Nat. Sci.) 43: 164 - 177 (2009).
  • a regulatory region can contain a
  • a regulatory region can contain a MYBIAT motif having the consensus sequence AAACCA.
  • a regulatory region can contain a MYBPLANT motif having the consensus sequence MACCWAMC. See, Sablowski et al, EMBO J 13: 128-137 (1994); Tamagnone et al, Plant Cell 10: 135-154 (1998).
  • a regulatory region can contain a NR BNEXTA motif having the consensus sequence TAGTGGAT. See, Elliott and Shirsat, Plant Mol Biol 37:675-687 (1998).
  • a regulatory region can contain an
  • OSE2ROOTNODULE motif having the consensus sequence CTCTT See, Vieweg et ⁇ ., ⁇ , 17(l):62-69 (2004).
  • a regulatory region can contain a P1BS motif having the consensus sequence GNATATNC. See, Rubio et al, Genes Dev. 15: 2122- 2133.(2001); Shunmann et al., J Exp Bot. 55: 855-865. (2004); and Shunmann et al, Plant Physiol. 136: 4205-4214 (2004).
  • a regulatory region can contain a P-box gibberellin-responsive element having the consensus sequence TAACAAA. See, Gubler and Jacobsen, Plant Cell, Vol. 4, 1435-1441 (1992).
  • a regulatory region can contain a phosphate starvation induced binding motif. See, Rubio et al, Genes Dev 15:2122-2133 (2001).
  • a regulatory region can contain a PRECONSCRHSP70A motif having the consensus sequence SCGAYNRNNNNNNNNNNNNNHD (SEQ ID NO: 54). See, von Gromoff et al, Nucleic Acids Res. 34:4767-4779 (2006).
  • a regulatory region can contain an RGATAOS (R-GATA (GATA motif binding factor) binding site) motif having the consensus sequence CAGAAGATAA. See, Yin et al, Plant J., 12: 1179-1188 (1997).
  • a regulatory region can contain a ROOTMOTIFTAPOX1 motif having the consensus sequence ATATT.
  • a regulatory region can contain a RYREPEATGMGY2 motif having the consensus sequence CATGCAT. See, Lelievre et al, Plant Physiol 98:387-391 (1992).
  • a regulatory region can contain a RYREPEATVFLEB4 motif having the consensus sequence CATGCATG. See, Curaba et al, Plant Physiol. 136: 3660-3669 (2004); and Nag et al, Plant Mol Biol. 59: 821-838 (2005).
  • a regulatory region can contain a SBOXATRBCS motif having the consensus sequence CACCTCCA. See, Acevedo- Hernandez et al, Plant J. 43:506-519 (2005).
  • a regulatory region can contain a
  • a regulatory region can contain a SP8BFIBSP8BIB motif having the consensus sequence TACT ATT. See, Ishiguro and Nakamura, Plant Mol Biol 18:97-108 (1992); Ishiguro and Nakamura, Mol Gen Genet 244: 563-571 (1994).
  • a regulatory region can contain a TATAbox motif such as a TATABOX1, TATABOX2, TATABOX4, or TATABOX5 motif having the consensus sequence CTATAAATAC (SEQ ID NO: 55), TATAAAT, TATATAA, and TTATTT,
  • a regulatory region can contain a TATABOXOSPAL motif having the consensus sequence TATTTAA. See, Zhu et al, Plant Cell 14: 795-803 (2002).
  • a regulatory region can contain a TATC-box motif having the consensus sequence TATCCCA.
  • a regulatory region can contain a TCA1 motif having the sequence TATCCCA.
  • a regulatory region can contain a TC_rich ⁇ repeats motif having the consensus sequence ATTCTCTAAC (SEQ ID NO: 56).
  • a regulatory region can contain a TCA-element motif having the consensus sequence CCATCTTTTT (SEQ ID NO:57). See,
  • a regulatory region can contain a TATCCAYMOTIFOSRAMY3D motif having the consensus sequence TATCCAY. See, Toyofuku et al, FEBS Lett, 428:275-280 (1998); and Rubio- Somoza et al, Plant J. 47: 269-281 (2006).
  • a regulatory region can contain a
  • a regulatory region can contain a
  • TRANSINITMONOCOTS motif having the consensus sequence RMNAUGGC. See, Joshi et al, Plant Mol Biol 35:993-1001 (1997).
  • a regulatory region can contain a UP1ATMSD motif having the consensus sequence GGCCCA(A/T)(A/T)(A/T). See, Ma and Bohnert, Genome Biol. 8(4): R49 (2007).
  • a regulatory region can contain a
  • a regulatory region can contain an
  • UPRMOTIFIIAT motif having the consensus sequence CCNNNNNNNNNNCCACG (SEQ ID NO:58). See, Martinez and Chrispeels, Plant Cell. 15:561-576 (2003); and Oh et al, Biochem Biophys Res Commun. 301 :225-230 (2003).
  • a regulatory region has a nucleotide sequence with 90% or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO: l or a fragment thereof, wherein the nucleic acid contains a TATABOX4, GAREAT, and CARGCW8GAT motif.
  • the TATABOX4 motif can be the motif at nucleotides 1408 to 1414 of SEQ ID NO: 1 or a TATABOX4 motif heterologous to those in SEQ ID NO: 1.
  • the GAREAT motif can be the motif at nucleotides 338 to 344 of SEQ ID NO: 1 or a GAREAT motif heterologous to that in SEQ ID NO: l .
  • the CARGCW8GAT motif can be the motif at nucleotides 298 to 307 of SEQ ID NO:l or a CARGCW8GAT motif heterologous to that in SEQ ID NO: 1. In some cases, such a regulatory region can also include a 5' UTR.
  • a regulatory region has a nucleotide sequence with 90% or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO:2 or a fragment thereof, wherein the nucleic acid contains a CARGNCAT and a CARGCW8GAT motif.
  • the CARGNCAT motif can be the motif at nucleotides 844 to 855 of SEQ ID NO:2 or a CARGNCAT motif heterologous to that in SEQ ID NO:2.
  • the CARGCW8GAT motif can be the motif at nucleotides 291 to 300, nucleotides 706 to 717, or nucleotides 845 to 854 of SEQ ID NO:2 or a CARGCW8GAT motif heterologous to those in SEQ ID NO:2.
  • a regulatory region can also include a 5' UTR.
  • a regulatory region has a nucleotide sequence with 90% or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO: 3 or a fragment thereof, wherein the nucleic acid contains an UPRMOTIFIIAT, a CACGCAATGMGH3, and a SP8BFIBSP8AIB motif.
  • the UPRMOTIFIIAT motif can be the motif at nucleotides 769 to 787 of SEQ ID NO:3 or an UPRMOTIFIIAT motif heterologous to that in SEQ ID NO:3.
  • the CACGCAATGMGH3 motif can be the motif at nucleotides 715 to 722 of SEQ ID NO:3 or a CACGCAATGMGH3 motif heterologous to that in SEQ ID NO:3.
  • the SP8BFIBSP8AIB motif can be the motif at nucleotides 488 to 495 of SEQ ID NO:3 or a SP8BFIBSP8AIB motif heterologous to that in SEQ ID NO:3.
  • such a regulatory region can contain an intron (e.g., at nucleotides 1184 to 1199 of SEQ ID NO:3.
  • such a regulatory region can also include a 5' UTR.
  • a regulatory region has a nucleotide sequence with 90%) or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO:4 or a fragment thereof, wherein the nucleic acid contains a TATABOX2 motif and OSE1ROOTNODULE motif.
  • the TATABOX2 motif can be the motif at nucleotides 910 to 916 of SEQ ID NO:4 or a TATABOX2 motif heterologous to that in SEQ ID NO:4.
  • the OSEIROOTNODULE motif can be the motif at nucleotides 58 to 63, 359 to 363, 522 to 526, or 801 to 806 of SEQ ID NO:4 or a OSEIROOTNODULE motif heterologous to those in SEQ ID NO:4.
  • a regulatory region can also include a 5' UTR.
  • a regulatory region has a nucleotide sequence with 90% or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO: 5 or a fragment thereof, wherein the nucleic acid contains an
  • the ATHBIATCONSENSUS motif can be the motif at nucleotides 1569 to 1577 of SEQ ID NO:5 or a ATHBIATCONSENSUS motif heterologous to that in SEQ ID NO:5.
  • the GAREAT motif can be the motif at nucleotides 862 to 868 of SEQ ID NO:5 or a GAREAT motif heterologous to that in SEQ ID NO:5.
  • the P IBS motif can be the motif at nucleotides 119 to 126 of SEQ ID NO:5 or a P1BS motif heterologous to that in SEQ ID NO:5.
  • the GARE20SREP1 motif can be the motif at nucleotides 891 to 897 of SEQ ID NO:5 or a GARE20SREP1 motif heterologous to that in SEQ ID NO:5.
  • the ABRE motif can be the motif at nucleotides 1521 to 1527 of SEQ ID NO:5 or an ABRE motif heterologous to that in SEQ ID NO:5.
  • the TATC-box motif can be the motif at nucleotides 621 to 627 of SEQ ID NO: 5 or a TATC-box motif heterologous to that in SEQ ID NO:5.
  • such a regulatory region can also include a 5' UTR.
  • the 5' UTR can be the 5' UTR at nucleotides 1690 to 1760 of SEQ ID NO: 5 or can be a heterologous UTR.
  • a regulatory region has a nucleotide sequence with 90% or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO:6 or a fragment thereof (e.g., SEQ ID NO:7, which contains nucleotides 118 to 600 of SEQ ID NO: 6), wherein the nucleic acid contains a Phosphate-starvation induced binding site and a GAREAT motif.
  • the Phosphate-starvation induced binding site motif can be the motif at nucleotides 411 to 416 of SEQ ID NO:6 or a Phosphate-starvation induced binding site motif heterologous to that in SEQ ID NO:6.
  • the GAREAT motif can be the motif at nucleotides 55 to 60 or 239 to 244 of SEQ ID NO:6 or a GAREAT motif heterologous to those in SEQ ID NO:6.
  • a regulatory region can also include a 5' UTR.
  • the 5' UTR can be the 5' UTR at nucleotides 466 to600 of SEQ ID NO: 6 or can be a heterologous UTR.
  • a regulatory region has a nucleotide sequence with 90% or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO: 7 or a fragment thereof, wherein the nucleic acid contains a P1BS and GAREAT motif.
  • the P IBS motif can be the motif at nucleotides 295 to 300 of SEQ ID NO: 7 or a P1BS motif heterologous to that in SEQ ID NO:7.
  • the GAREAT can be the motif at nucleotides 123 to 128 of SEQ ID NO:7 or a GAREAT motif heterologous to that in SEQ ID NO:7.
  • such a regulatory region can also include a 5' UTR.
  • the 5' UTR can be the 5' UTR at nucleotides 350 to 484 of SEQ ID NO:7 or can be a heterologous UTR.
  • a regulatory region has a nucleotide sequence with 90% or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO: 8 or a fragment thereof, wherein the nucleic acid contains a TATABOX4 or TCA- element motif.
  • the TATABOX4 motif can be the motif at nucleotides 223 to 229 of SEQ ID NO: 8 or a TATABOX4 motif heterologous to that in SEQ ID NO:8.
  • the TCA- element motif can be the motif at nucleotides 174 to 183 of SEQ ID NO: 8 or a TCA- element motif heterologous to that in SEQ ID NO:8.
  • such a regulatory region can contain an intron (e.g., at nucleotides 323 to 1268 of SEQ ID NO: 8).
  • such a regulatory region can also include a 5' UTR.
  • the 5' UTR can be the 5' UTR at nucleotides 253 to 322 or 1269 to 1290 of SEQ ID NO:8 or can be a heterologous UTR.
  • a regulatory region has a nucleotide sequence with 90%) or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO: 9 or a fragment thereof, wherein the nucleic acid contains a TATABOX4 and an ATHB6COREAT motif.
  • the TATABOX4 motif can be the motif at nucleotides 1235 to 1241 of SEQ ID NO:9 or a TATABOX4 motif heterologous to that in SEQ ID NO:9.
  • the ATHB6COREAT motif can be the motif at nucleotides 607 to 615 of SEQ ID NO:9 or an ATHB6COREAT motif heterologous to that in SEQ ID NO:9.
  • such a regulatory region can also include a 5 ' UTR.
  • the 5 ' UTR can be the 5 ' UTR at nucleotides 1268 to 1400 of SEQ ID NO:9 or can be a heterologous UTR.
  • a regulatory region has a nucleotide sequence with 90% or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO: 10 or a fragment thereof, wherein the nucleic acid contains a TCAl Motif, TATAbox motif, and ATCT motif.
  • the TCAlMotif can be the motif at nucleotides 589 to 598 of SEQ ID NO: 10 or a TCAlMotif heterologous to that in SEQ ID NO: 10.
  • the TATAbox motif can be the motif at nucleotides 628 to 634 of SEQ ID NO: 10 or a TATAbox motif heterologous to that in SEQ ID NO: 10.
  • the ATCT motif can be the motif at nucleotides 585 to 594 of SEQ ID NO: 10 or an ATCT motif heterologous to that in SEQ ID NO: 10.
  • such a regulatory region can also include a 5 ' UTR.
  • the 5 ' UTR can be the 5' UTR at nucleotides 662 to 689 of SEQ ID NO: 10 or can be a heterologous UTR.
  • a regulatory region has a nucleotide sequence with 90% or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO: 11 or a fragment thereof, wherein the nucleic acid contains a GATABOX and CARGCW8GAT motif.
  • the GATABOX motif can be the motif at nucleotides 1113 to 1116 of SEQ ID NO: 11 or a GATABOX motif heterologous to that in SEQ ID NO: 11.
  • the CARGCW8GAT motif can be the motif at nucleotides 596 to 605 of SEQ ID NO: l 1 or a CARGCW8GAT motif heterologous to that in SEQ ID NO: 11.
  • such a regulatory region can also include a 5 ' UTR.
  • the 5 ' UTR can be the 5 ' UTR at nucleotides 1119 to 1200 of SEQ ID NO: 11 or can be a heterologous UTR.
  • a regulatory region has a nucleotide sequence with 90%) or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO: 12 or a fragment thereof, wherein the nucleic acid contains a TATABOX4,
  • the TATABOX4 motif can be the motif at nucleotides 668 to 674 of SEQ ID NO: 12 or a TATABOX4 motif heterologous to that in SEQ ID NO: 12.
  • the UP2ATMSD motif can be the motif at nucleotides 220 to 227 or 765 ti 773 of SEQ ID NO: 12 or an UP2ATMSD motif heterologous to those in SEQ ID NO: 12.
  • the ELI-box3 motif can be the motif at nucleotides 10 to 18 or 156 to 164 of SEQ ID NO: 12 or an ELI-box3 motif heterologous to those in SEQ ID NO: 12.
  • such regulatory regions can also include a 5' UTR.
  • the 5' UTR can be the 5' UTR at nucleotides 816 to 855 of SEQ ID NO: 12 or can be a heterologous UTR.
  • a regulatory region has a nucleotide sequence with 90% or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO: 13 or a fragment thereof, wherein the nucleic acid contains a TATA BOX,
  • the TATA BOX motif can be the motif at nucleotides 280 to 298 of SEQ ID NO: 13 or a TATA BOX motif heterologous to that in SEQ ID NO: 13.
  • the CTRMC AM V35 S motif can be the motif at nucleotides 351 to 359 of SEQ ID NO: 13 or a CTRMC AM V35 S motif heterologous to that in SEQ ID NO: 13.
  • the ACE motif can be the motif at nucleotides 100 to 106 of SEQ ID NO: 13 or an ACE motif heterologous to that in SEQ ID NO: 13.
  • CACGTGMOTIF motif can be the motif at nucleotides 102 to 107 of SEQ ID NO: 13 or a CACGTGMOTIF motif heterologous to that in SEQ ID NO: 13.
  • a regulatory region can also include a 5 ' UTR.
  • the 5 ' UTR can be the 5 ' UTR at nucleotides 325 to 429 of SEQ ID NO: 13 or can be a heterologous UTR.
  • a regulatory region has a nucleotide sequence with 90% or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO: 14 or a fragment thereof (e.g., the nucleotide sequence set forth in SEQ ID NO:28, which is nucleotides 193 to 700 of SEQ ID NO: 14), wherein the nucleic acid contains a ACGTABREMOTIFAOSOSEM, ABREL ATERD 1 , and ABRE motif.
  • ACGTABREMOTIFAOSOSEM motif can be the motif at nucleotides 389 to 396 of SEQ ID NO: 14 or an ACGTABREMOTIFAOSOSEM motif heterologous to that in SEQ ID NO: 14.
  • the ABREL ATERD 1 motif can be the motif at nucleotides 305 to 309 of SEQ ID NO: 14 or an ABREL ATERD 1 motif heterologous to that in SEQ ID NO: 14.
  • the ABRE motif can be the motif at nucleotides 269 to 275 of SEQ ID NO: 14 or an ABRE motif heterologous to that in SEQ ID NO: 14.
  • such regulatory regions can also include a 5' UTR.
  • the 5' UTR can be the 5' UTR at nucleotides 560 to 700 of SEQ ID NO: 14 or can be a heterologous UTR.
  • a regulatory region has a nucleotide sequence with 90%) or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO: 15 or a fragment thereof, wherein the nucleic acid contains a TATABOX4,
  • the TATABOX4 motif can be the motif at nucleotides 796 to 802 of SEQ ID NO: 15 or a TATABOX4 motif heterologous to that in SEQ ID NO: 15.
  • the UP2ATMSD motif can be the motif at nucleotides 6 to 13 of SEQ ID NO: 15 or an UP2ATMSD motif heterologous to that in SEQ ID NO: 15.
  • the TC richVepeats motif can be the motif at nucleotides 179 to 188 of SEQ ID NO: 15 or a TC richVepeats motif heterologous to that in SEQ ID NO: 15.
  • the GAGAGMGSA1 motif can be the motif at nucleotides 725 to 742 of SEQ ID NO: 15 or a GAGAGMGSA1 motif heterologous to those in SEQ ID NO: 15.
  • regulatory regions can also include a 5 ' UTR.
  • the 5 ' UTR can be the 5 ' UTR at nucleotides 829 to 1000 of SEQ ID NO: 15 or can be a heterologous UTR.
  • a regulatory region has a nucleotide sequence with 90% or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO: 16 or a fragment thereof, wherein the nucleic acid contains a PRECONSCRHSP70A, TATAbox, ABRE, and HSE motif.
  • the PRECONSCRHSP70A motif can be the motif at nucleotides 735 to 758 of SEQ ID NO: 16 or a PRECONSCRHSP70A motif heterologous to that in SEQ ID NO: 16.
  • the TATAbox motif can be the motif at nucleotides 723 to 727 of SEQ ID NO: 16 or a TATAbox motif heterologous to that in SEQ ID NO: 16.
  • the ABRE motif can be the motif at nucleotides 656 to 661 or 689 to 694 of SEQ ID NO: 16 or an ABRE motif heterologous to those in SEQ ID NO: 16.
  • the HSE motif can be the motif at nucleotides 380 to 389 of SEQ ID NO : 16 or a HSE motif heterologous to that in SEQ ID NO: 16.
  • such a regulatory region can also include a 5' UTR.
  • the 5' UTR can be the 5' UTR at nucleotides 754 to 840 of SEQ ID NO: 16 or can be a heterologous UTR.
  • a regulatory region has a nucleotide sequence with 90% or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO: 17 or a fragment thereof, wherein the nucleic acid contains a GT1MOTIFPSRBCS, TATAbox, and RYREPEATVFLEB4 motif.
  • the GT1MOTIFPSRBCS motif can be the motif at nucleotides 66 to 77 of SEQ ID NO: 17 or a GT1MOTIFPSRBCS motif heterologous to that in SEQ ID NO: 17.
  • the TATAbox motif can be the motif at nucleotides 682 to 688 of SEQ ID NO: 17 or a TATAbox motif heterologous to that in SEQ ID NO: 17.
  • the RYREPEATVFLEB4 motif can be the motif at nucleotides 344 to 351 of SEQ ID NO: 17 or a RYREPEATVFLEB4 motif heterologous to that in SEQ ID NO: 17.
  • a regulatory region can also include a 5' UTR.
  • the 5' UTR can be the 5 ' UTR at nucleotides 697 to 773 of SEQ ID NO : 17 or can be a heterologous UTR.
  • a regulatory region has a nucleotide sequence with 90% or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO: 18 or a fragment thereof, wherein the nucleic acid contains an UPRMOTIFIIAT, PRECONSCRHSP70A or GCC ⁇ Box motif.
  • the UPRMOTIFIIAT motif can be the motif at nucleotides 220 to 238 of SEQ ID NO: 18 or an UPRMOTIFIIAT motif heterologous to that in SEQ ID NO: 18.
  • the PRECONSCRHSP70A motif can be the motif at nucleotides 213 to 236 of SEQ ID NO: 18 or a PRECONSCRHSP70A motif heterologous to that in SEQ ID NO : 18.
  • the GCC ⁇ Box motif can be the motif at nucleotides 184 to 190 of SEQ ID NO: 18 or a GCC ⁇ Box motif heterologous to that in SEQ ID NO: 18.
  • such a regulatory region can contain an intron (e.g., at nucleotides 473 to 1680 of SEQ ID NO: 18).
  • such a regulatory region can also include a 5' UTR.
  • the 5' UTR can be the 5' UTR at nucleotides 329 to 472 or 1681 to 1700 of SEQ ID NO: 18 or can be a heterologous UTR.
  • a regulatory region has a nucleotide sequence with 90% or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO: 19 or a fragment thereof, wherein the nucleic acid contains a TATABOX4,
  • the TATABOX4 motif can be the motif at nucleotides 407 to 413 of SEQ ID NO: 19 or a TATABOX4 motif heterologous to that in SEQ ID NO: 19.
  • the ATHB2ATCONSENSUS motif can be the motif at nucleotides 258 to 266 of SEQ ID NO: 19 or a ATHB2ATCONSENSUS motif heterologous to that in SEQ ID NO: 19.
  • the SORLREP2AT motif can be the motif at nucleotides 304 to 312 of SEQ ID NO: 19 or a SORLREP2AT motif heterologous to that in SEQ ID NO: 19. In some cases, such a regulatory region can also include a 5' UTR.
  • a regulatory region has a nucleotide sequence with 90%) or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO:20 or a fragment thereof, wherein the nucleic acid contains a PRECONSCRHSP70A, UPRMOTIFIIAT, ABRE, and MBS motif.
  • the PRECONSCRHSP70A motif can be the motif at nucleotides 576 to 599 of SEQ ID NO:20 or a PRECONSCRHSP70A motif heterologous to that in SEQ ID NO:20.
  • the UPRMOTIFIIAT motif can be the motif at nucleotides 242 to 260, 384 to 402, 460 to 478, or 544 to 562 of SEQ ID NO:20 or an UPRMOTIFIIAT motif heterologous to those in SEQ ID NO:20.
  • the ABRE motif can be the motif at nucleotides 257 to 262, 399 to 404, or 475 to 480 of SEQ ID NO:20 or an ABRE motif heterologous to that in SEQ ID NO:20.
  • the MBS motif can be the motif at nucleotides 280 to 285 of SEQ ID NO:20 or a MBS motif heterologous to that in SEQ ID NO:20.
  • such a regulatory region can also include a 5' UTR.
  • the 5' UTR can be the 5' UTR at nucleotides 637 to 814 of SEQ ID NO:20 or can be a heterologous UTR.
  • a regulatory region has a nucleotide sequence with 90% or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO:21 or a fragment thereof, wherein the nucleic acid contains a MYBPLANT motif.
  • the MYBPLANT motif can be the motif at nucleotides 454 to 461, 509 to 516, 858 to 868 of SEQ ID NO:20 or a MYBPLANT motif heterologous to those in SEQ ID NO:21.
  • such a regulatory region can also include a 5' UTR.
  • a regulatory region has a nucleotide sequence with 90% or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO:22 or a fragment thereof, wherein the nucleic acid contains a PRECONSCRHSP70A, ACIIPVPAL2, and CAAT-box motif.
  • the PRECONSCRHSP70A motif can be the motif at nucleotides 36 to 59 of SEQ ID NO:22 or a PRECONSCRHSP70A motif heterologous to that in SEQ ID NO:22.
  • the ACIIPVPAL2 motif can be the motif at nucleotides 266 to 277 of SEQ ID NO:22 or an ACIIPVPAL2 motif heterologous to that in SEQ ID NO:22.
  • the CAAT-box motif can be the motif at nucleotides 487 to 491 of SEQ ID NO:22 or a CAAT-box motif heterologous to that in SEQ ID NO:22.
  • such a regulatory region can contain an intron (e.g., at nucleotides 413 to 1500 of SEQ ID NO: 22).
  • such a regulatory region can also include a 5' UTR.
  • the 5' UTR can be the 5' UTR at nucleotides 353 to 412 of SEQ ID NO:22 or can be a heterologous UTR.
  • a regulatory region has a nucleotide sequence with 90%) or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO:23 or a fragment thereof, wherein the nucleic acid contains a TATABOX4 and ABREATCONSENSUS motif.
  • the TATABOX4 motif can be the motif at nucleotides 153 to 159 of SEQ ID NO:23 or a TATABOX4 motif heterologous to that in SEQ ID NO:23.
  • the ABREATCONSENSUS motif can be the motif at nucleotides 85 to 92 of SEQ ID NO:23 or an ABREATCONSENSUS motif heterologous to that in SEQ ID NO:23.
  • such a regulatory region can contain an intron (e.g., at nucleotides 250 to 835 of SEQ ID NO: 23).
  • such a regulatory region can also include a 5' UTR.
  • the 5' UTR can be the 5' UTR at nucleotides 184 to 249 or 836 to 880 of SEQ ID NO:23 or can be a heterologous UTR.
  • a regulatory region has a nucleotide sequence with 90% or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO:24 or a fragment thereof, wherein the nucleic acid contains a TATABOX4, TATC- box, P1BS, and OSE2ROOTNODULE motif.
  • the TATABOX4 motif can be the motif at nucleotides 939 to 945 of SEQ ID NO:24 or a TATABOX4 motif heterologous to that in SEQ ID NO:24.
  • the TATC-box motif can be the motif at nucleotides 941 to 947 of SEQ ID NO:24 or a TATC-box motif heterologous to that in SEQ ID NO:24.
  • the P1BS motif can be the motif at nucleotides 493 to 500 of SEQ ID NO:24 or a P1BS motif heterologous to that in SEQ ID NO:24.
  • the OSE2ROOTNODULE motif can be the motif at nucleotides 619 to 624 of SEQ ID NO:24 or an OSE2ROOTNODULE motif heterologous to that in SEQ ID NO:24.
  • such a regulatory region can also include a 5' UTR.
  • the 5' UTR can be the 5' UTR at nucleotides 968 to 1000 of SEQ ID NO:24 or can be a heterologous UTR.
  • a regulatory region has a nucleotide sequence with 90% or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO:25 or a fragment thereof, wherein the nucleic acid contains a TATABOX2,
  • the TATABOX2 motif can be the motif at nucleotides 1617 to 1623 of SEQ ID NO:25 or a TATABOX2 motif heterologous to that in SEQ ID NO:25.
  • the ABREATCONSENSUS motif can be the motif at nucleotides 710 to 717 of SEQ ID NO:25 or an ABREATCONSENSUS motif heterologous to that in SEQ ID NO:25.
  • the IBOX motif can be the motif at nucleotides 1523 to 1528 of SEQ ID NO:25 or an IBOX motif heterologous to that in SEQ ID NO:25.
  • the ABRELATERD 1 motif can be the motif at nucleotides 1474 to 1478, 1046 to 1050, or 1096 to 1100 of SEQ ID NO:25 or an ABRELATERD 1 motif heterologous to those in SEQ ID NO:25.
  • the MBS motif can be the motif at nucleotides 1379 to 1375 of SEQ ID NO:25 or a MBS motif heterologous to that in SEQ ID NO:25.
  • such a regulatory region can also include a 5 ' UTR.
  • the 5 ' UTR can be the 5' UTR at nucleotides 1665 to 1763 of SEQ ID NO:25 or can be a heterologous UTR.
  • a regulatory region has a nucleotide sequence with 90% or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO:26 or a fragment thereof, wherein the nucleic acid contains a MYBIAT, TATABOX, and MBS motif.
  • the MYBIAT motif can be the motif at nucleotides 193 to 208, 340 to 345, 405 to 410, or 527 to 532 of SEQ ID NO:26 or a MYBIAT motif heterologous to those in SEQ ID NO:26.
  • the TATABOX motif can be the motif at nucleotides 754 to 759 of SEQ ID NO:26 or a TATABOX motif heterologous to that in SEQ ID NO:26.
  • the MBS motif can be the motif at nucleotides 672 to 677 of SEQ ID NO:26 or a MBS motif heterologous to that in SEQ ID NO:26. In some cases, such a regulatory region can also include a 5 ' UTR.
  • a regulatory region has a nucleotide sequence with 90% or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO:27 or a fragment thereof, wherein the nucleic acid contains a TATABOX5,
  • the TATABOX5 motif can be the motif at nucleotides 185 to 190 of SEQ ID NO:27 or a TATABOX5 motif heterologous to that in SEQ ID NO:27.
  • the CARGCW8GAT motif can be the motif at nucleotides 120 to 129 of SEQ ID NO:27 or a CARGCW8GAT motif heterologous to that in SEQ ID NO:27.
  • the E2FCONSENSUS motif can be the motif at nucleotides 54 to 61 of SEQ ID NO:27 or an E2FCONSENSUS motif heterologous to that in SEQ ID NO:27.
  • the LEAFYATAG motif can be the motif at nucleotides 57 to 63 of SEQ ID NO:27 or a LEAFYATAG motif heterologous to that in SEQ ID NO:27.
  • the ROOTMOTIFTAPOXl motif can be the motif at nucleotides 45 to 49 of SEQ ID NO:27 or a ROOTMOTIFTAPOXl motif heterologous to that in SEQ ID NO:27.
  • such a regulatory region can also include a 5 ' UTR.
  • the 5 ' UTR can be the 5 ' UTR at nucleotides 213 to 400 of SEQ ID NO:27 or can be a heterologous UTR.
  • a regulatory region has a nucleotide sequence with 90%) or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO:28 or a fragment thereof, wherein the nucleic acid contains an ABRELATERD1 motif
  • the ABRELATERD 1 motif can be the motif at nucleotides 79 to 83, 113 to 117, 198 to 202, or 271 to 275 of SEQ ID NO:28 or an ABRELATERD 1 motif heterologous to those in SEQ ID NO:28.
  • such a regulatory region can also include a 5' UTR.
  • the 5' UTR can be the 5' UTR at nucleotides 368 to506 of SEQ ID NO:28 or can be a heterologous UTR.
  • a regulatory region has a nucleotide sequence with 90% or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO:29 or a fragment thereof, wherein the nucleic acid contains an ABRELATERD 1 motif.
  • the ABRELATERD 1 motif can be the motif at nucleotides 125 to 129 or 542 to 546 of SEQ ID NO:29 or an ABRELATERD 1 motif heterologous to those in SEQ ID NO:29.
  • such a regulatory region can also include a 5' UTR.
  • the 5' UTR can be the 5 ' UTR at nucleotides 1111 to 1200 of SEQ ID NO:29 or can be a
  • a regulatory region has a nucleotide sequence with 90% or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO:30 or a fragment thereof (e.g., nucleotides 269 to 763 or 419 to 763 of SEQ ID NO: 30) wherein the nucleic acid contains a TATABOX motif.
  • the TATABOX motif can be the motif at nucleotides 664 to 669 of SEQ ID NO:30 or a TATABOX motif heterologous to that in SEQ ID NO:30.
  • such a regulatory region can also include a 5' UTR.
  • the 5' UTR can be the 5' UTR at nucleotides 698 to736 of SEQ ID NO:30 or can be a heterologous UTR.
  • a regulatory region has a nucleotide sequence with 90%) or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO:31 or a fragment thereof (e.g., the nucleotide sequence set forth in SEQ ID NO:32, which is nucleotides 1205 to 3004 of SEQ ID NO:31) that contains an UP2ATMSD and TATABOX motif.
  • the UP2ATMSD motif can be the motif at nucleotides 407 to 414 of SEQ ID NO:32 or an UP2ATMSD motif heterologous to that in SEQ ID NO:32.
  • the TATABOX motif can be the motif at nucleotides 381 to 388 of SEQ ID NO:32 or a TATABOX motif heterologous to that in SEQ ID NO:32.
  • a regulatory region can contain an intron (e.g., at nucleotides 526 to 1699 of SEQ ID NO: 32).
  • such a regulatory region can also include a 5' UTR.
  • the 5' UTR can be the 5' UTR at nucleotides 408 to 525 of SEQ ID NO:32 or can be a heterologous UTR.
  • a regulatory region has a nucleotide sequence with 90% or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO:33 or a fragment thereof, wherein the nucleic acid contains a TATABOX4,
  • RGATAOS R-GATA (GATA motif binding factor) binding site
  • 3-AFl ⁇ binding ⁇ site light responsive element or P-box gibberellin-responsive element motif.
  • TATABOX4 motif can be the motif at nucleotides 852 to 858 of SEQ ID NO:33 or a TATABOX4 motif heterologous to that in SEQ ID NO:33
  • the 3-AFl ⁇ binding ⁇ site light responsive element motif can be the motif at nucleotides 100 to 1 lOof SEQ ID NO:33 or a 3-AFl ⁇ binding ⁇ site light responsive element motif heterologous to that in SEQ ID NO:33.
  • the P-box gibberellin-responsive element motif can be the motif at nucleotides 385 to 395 of SEQ ID NO:33 or a P-box gibberellin-responsive element motif heterologous to that in SEQ ID NO:33.
  • the RGATAOS motif can be the motif at nucleotides 822 to 830 of SEQ ID NO:33 or a RGATAOS motif heterologous to that in SEQ ID NO:33.
  • a regulatory region can also include a 5' UTR.
  • the 5' UTR can be the 5' UTR at nucleotides 881 to 946 of SEQ ID NO:33 or can be a heterologous UTR.
  • a regulatory region has a nucleotide sequence with 90% or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO:34 or a fragment thereof. In some cases, such a regulatory region can also include a 5' UTR.
  • a regulatory region has a nucleotide sequence with 90%) or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO:35 or a fragment thereof. In some cases, such a regulatory region can also include a 5' UTR.
  • a regulatory region has a nucleotide sequence with 90%o or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO:36 or a fragment thereof. In some cases, such a regulatory region can also include a 5' UTR. [00117] In some embodiments, a regulatory region has a nucleotide sequence with 90% or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO:37 or a fragment thereof, wherein the nucleic acid contains a TATAbox motif.
  • the TATAbox motif can be the motif at nucleotides 670 to678 of SEQ ID NO:37 or a TATAbox motif heterologous to that in SEQ ID NO:37.
  • such a regulatory region can contain an intron (e.g., at nucleotides 741 to 823 of SEQ ID NO: 37).
  • such a regulatory region can also include a 5' UTR.
  • the 5' UTR can be the 5' UTR at nucleotides 713 to 740 or 824 to 846 of SEQ ID NO:37 or can be a heterologous UTR.
  • a regulatory region has a nucleotide sequence with 90% or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO:38 or a fragment thereof, wherein the nucleic acid contains a TATAbox or CAAT- box motif.
  • the TATAbox motif can be the motif at nucleotides 785 to793 of SEQ ID NO :38 or a TATAbox motif heterologous to that in SEQ ID NO:38
  • the CAAT-box motif can be the motif at nucleotides 722 to729 of SEQ ID NO:38 or a CAAT-box motif heterologous to that in SEQ ID NO:38.
  • such a regulatory region can also include a 5' UTR.
  • the 5' UTR can be the 5' UTR at nucleotides 817 to 925 of SEQ ID NO:38 or can be a heterologous UTR.
  • a regulatory region has a nucleotide sequence with 90%) or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO:39 or a fragment thereof. In some cases, such a regulatory region can also include a 5' UTR.
  • a regulatory region has a nucleotide sequence with 90%o or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO:40 or a fragment thereof, wherein the nucleic acid contains a TATAbox and enhancer motif.
  • the TATAbox motif can be the motif at nucleotides 773 to 738 of SEQ ID NO:40 or a TATAbox motif heterologous to that in SEQ ID NO:40.
  • the enhancer motif can be the motif at nucleotides 303 to 308 or 602 to 607 of SEQ ID NO:40 or an enhancer motif heterologous to those in SEQ ID NO:40.
  • such a regulatory region can also include a 5' UTR.
  • the 5' UTR can be the 5' UTR at nucleotides 806 to 835 of SEQ ID NO:40 or can be a heterologous UTR.
  • a regulatory region has a nucleotide sequence with 90% or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO:41 or a fragment thereof. In some cases, such a regulatory region can also include a 5' UTR.
  • a regulatory region has a nucleotide sequence with 90% or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO:42 or a fragment thereof. In some cases, such a regulatory region can also include a 5' UTR.
  • a regulatory region has a nucleotide sequence with 90%) or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO:43 or a fragment thereof. In some cases, such a regulatory region can also include a 5' UTR.
  • a regulatory region has a nucleotide sequence with 90%o or greater sequence identity to the polynucleotide sequence set forth in SEQ ID NO:44 or a fragment thereof, wherein the nucleic acid contains a SP8BFIBSP8AIB, CACGCAATGMGH3, and SP8BFIBSP8AIB motif.
  • the SP8BFIBSP8AIB motif can be the motif at nucleotides 1122 to 1140 of SEQ ID NO:44 or a SP8BFIBSP8AIB motif heterologous to that in SEQ ID NO:44.
  • the CACGCAATGMGH3 motif can be the motif at nucleotides 1068 to 1075 of SEQ ID NO:44 or a CACGCAATGMGH3 motif heterologous to that in SEQ ID NO:44.
  • the SP8BFIBSP8AIB motif can be the motif at nucleotides 841 to 848 of SEQ ID NO:44 or a SP8BFIBSP8AIB motif heterologous to that in SEQ ID NO:44.
  • such a regulatory region can also include a 5' UTR.
  • Promoters of the document were tested for activity by cloning the sequence into an appropriate vector, transforming plants with the construct and assaying for marker gene expression.
  • Recombinant DNA constructs were prepared which comprise the promoter sequences of the document inserted into a vector suitable for transformation of plant cells.
  • the construct can be made using standard recombinant DNA techniques (Sambrook et al. 1989) and can be introduced to the species of interest by Agrobacterium-rnQdiatcd transformation or by other means of transformation as referenced below.
  • the vector backbone can be any of those typical in the art such as plasmids, viruses, artificial chromosomes, BACs, YACs and PACs and vectors of the sort described by
  • the construct comprises a vector containing a promoter sequence of the present document operationally linked to any marker gene.
  • the promoter was identified as a promoter by the expression of the marker gene.
  • GFP Green Fluorescent Protein
  • the vector may also comprise a marker gene that confers a selectable phenotype on plant cells.
  • the marker may encode biocide resistance, particularly antibiotic resistance, such as resistance to kanamycin, G418, bleomycin, hygromycin, or herbicide resistance, such as resistance to chlorosulfuron or phosphinotricin.
  • Vectors can also include origins of replication, scaffold attachment regions (SARs), markers, homologous sequences, introns, etc.
  • the promoter and promoter control elements of the present document can be used alone or combined with each other to produce the desired preferential transcription.
  • the promoters of the document can be combined with other known sequences to obtain other useful promoters to modulate, for example, tissue transcription specific or transcription specific to certain conditions.
  • Such preferential transcription can be determined using the techniques or assays described above.
  • Promoters can contain any number of control elements.
  • a promoter can contain multiple transcription binding sites or other control elements.
  • One element may confer tissue or organ specificity; another element may limit transcription to specific time periods, etc.
  • promoters will contain at least a basal or core promoter as described above. Any additional element can be included as desired.
  • a fragment comprising a basal or "core" promoter can be fused with another fragment with any number of additional control elements.
  • promoters that are induced under stress conditions and can be combined with those of the present document: ldhl (oxygen stress; tomato; see Germain and Ricard (1997) Plant Mol Biol 35:949-54), GPx and CAT (oxygen stress; mouse; see Franco et al. (1999) Free Radic Biol Med 27: 1122-32), ci7 (cold stress;
  • promoters are induced by the presence or absence of light can be used in combination with those of the present document: Topoisomerase II (pea; see Reddy et al. (1999) Plant Mol Biol 41 : 125-37), chalcone synthase (soybean; see Wingender et al. (1989) Mol Gen Genet 218:315-22) mdm2 gene (human tumor; see Saucedo et al. (1998) Cell Growth Differ 9: 119-30), Clock and BMAL1 (rat; see Namihira et al. (1999) Neurosci Lett 271 : 1-4, PHYA (Arabidopsis; see Canton and Quail (1999) Plant Physiol 121 : 1207-16), PRB-lb
  • the promoters and control elements of the following genes can be used in combination with the present document to confer tissue specificity: MipB (iceplant; Yamada et al. (1995) Plant Cell 7:1129-42) and SUCS (root nodules; broadbean; Kuster et al. (1993) Mol Plant Microbe Interact 6:507-14) for roots, OsSUTl (rice ; Hirose et al. (1997) Plant Cell Physiol 38: 1389-96) for leaves, Msg (soybean; Stomvik et al. (1999) Plant Mol Biol 41 :217-31) for siliques, cell (Arabidopsis; Shani et al. (1997) Plant Mol Biol 34(6):837 -42) and ACT11 (Arabidopsis; Huang et al. (1997) Plant Mol Biol 33: 125-39) for inflorescence.
  • MipB iceplant; Yamada et al. (1995) Plant Cell 7:1129
  • Still other promoters are affected by hormones or participate in specific physiological processes, which can be used in combination with those of present document.
  • Some examples are the ACC synthase gene that is induced differently by ethylene and brassinosteroids (mung bean; Yi et al. (1999) Plant Mol Biol 41 :443-54), the TAPG1 gene that is active during abscission (tomato; Kalaitzis et al. (1995) Plant Mol Biol 28:647-56), and the 1-aminocyclopropane-l-carboxylate synthase gene
  • control elements or the configuration or control elements can be determined or optimized to permit the desired protein-polynucleotide or polynucleotide interactions to occur.
  • the binding sites are spaced to allow each factor to bind without steric hindrance.
  • the spacing between two such hybridizing control elements can be as small as a profile of a protein bound to a control element.
  • two protein binding sites can be adjacent to each other when the proteins bind at different times during the transcription process.
  • control elements when two control elements hybridize the spacing between such elements will be sufficient to allow the promoter polynucleotide to hairpin or loop to permit the two elements to bind.
  • the spacing between two such hybridizing control elements can be as small as a t-RNA loop, to as large as 10 kb.
  • the spacing is no smaller than 5 bases; more typically, no smaller than 8; more typically, no smaller than 15 bases; more typically, no smaller than 20 bases; more typically, no smaller than 25 bases; even more typically, no smaller than 30, 35, 40 or 50 bases.
  • Such spacing between promoter control elements can be determined using the techniques and assays described above.
  • a plant transformation construct containing a promoter of the present document may be introduced into plants by any plant transformation method.
  • Methods and materials for transforming plants by introducing a plant expression construct into a plant genome in the practice of this document can include any of the well-known and demonstrated methods including electroporation (U.S. Pat. No. 5,384,253);
  • microprojectile bombardment U.S. Pat. No. 5,015,580; U.S. Pat. No. 5,550,318; U.S. Pat. No. 5,538,880; U.S. Pat. No. 6,160,208; U.S. Pat. No. 6,399,861; and U.S. Pat. No. 6,403,865; Agrobacterium-mediatGd transformation (U.S. Pat. No. 5,824,877; U.S. Pat. No. 5,591,616; U.S. Pat. No. 5,981,840; and U.S. Pat. No. 6,384,301); and protoplast transformation (U.S. Pat. No. 5,508,184).
  • the present promoters and/or promoter control elements may be delivered to a system such as a cell by way of a vector.
  • a vector may range from simply introducing the promoter or promoter control element by itself randomly into a cell to integration of a cloning vector containing the present promoter or promoter control element.
  • a vector need not be limited to a DNA molecule such as a plasmid, cosmid or bacterial phage that has the capability of replicating autonomously in a host cell. All other manner of delivery of the promoters and promoter control elements of the document are envisioned.
  • the various T-DNA vector types are a preferred vector for use with the present document. Many useful vectors are commercially available.
  • Marker sequences typically include genes that provide antibiotic resistance, such as tetracycline resistance, hygromycin resistance or ampicillin resistance, or provide herbicide resistance.
  • Specific selectable marker genes may be used to confer resistance to herbicides such as glyphosate, glufosinate or bromoxynil (Comai et al. (1985) Nature 317: 741-744; Gordon-Kamm et al. (1990) Plant Cell 2: 603-618; and Stalker et al. (1988) Science 242: 419-423).
  • Other marker genes exist which provide hormone responsiveness.
  • the promoter or promoter control element of the present document may be operably linked to a polynucleotide to be transcribed. In this manner, the promoter or promoter control element may modify transcription by modulating transcript levels of that polynucleotide when inserted into a genome.
  • the promoter or promoter control element need not be linked, operably or otherwise, to a polynucleotide to be transcribed.
  • the promoter or promoter control element may be inserted alone into the genome in front of a polynucleotide already present in the genome. In this manner, the promoter or promoter control element may modulate the transcription of a polynucleotide that was already present in the genome.
  • This polynucleotide may be native to the genome or inserted at an earlier time.
  • the promoter or promoter control element may be inserted into a genome alone to modulate transcription. See, for example, Vaucheret, H et al. (1998) Plant J 16: 651-659. Rather, the promoter or promoter control element may be simply inserted into a genome or maintained extrachromosomally as a way to divert transcription resources of the system to itself. This approach may be used to
  • polynucleotide to be transcribed is not limited.
  • the polynucleotide may include sequences that will have activity as RNA as well as sequences that result in a polypeptide product. These sequences may include, but are not limited to antisense sequences, RNAi sequences, ribozyme sequences, spliceosomes, amino acid coding sequences, and fragments thereof. Specific coding sequences may include, but are not limited to endogenous proteins or fragments thereof, or heterologous proteins including marker genes or fragments thereof.
  • constructs of the present document would typically contain a promoter operably linked to a transcribable nucleic acid molecule operably linked to a 3' transcription termination nucleic acid molecule.
  • constructs may include but are not limited to additional regulatory nucleic acid molecules from the 3 '-untranslated region (3' UTR) of plant genes (e.g., a 3' UTR to increase mRNA stability of the mRNA, such as the PI-II termination region of potato or the octopine or nopaline synthase 3' termination regions).
  • Constructs may include but are not limited to the 5' untranslated regions (5' UTR) of an mRNA nucleic acid molecule which can play an important role in translation initiation and can also be a genetic component in a plant expression construct.
  • 5' UTR 5' untranslated regions
  • non-translated 5' leader nucleic acid molecules derived from heat shock protein genes have been demonstrated to enhance gene expression in plants (see for example, U.S. Pat. No. 5,659,122 and U.S. Pat. No. 5,362,865, all of which are hereby incorporated by reference).
  • These additional upstream and downstream regulatory nucleic acid molecules may be derived from a source that is native or heterologous with respect to the other elements present on the promoter construct.
  • one embodiment of the document is a promoter such as provided in SEQ ID NOs: 1 - 44 or a fragment thereof, operably linked to a transcribable nucleic acid molecule so as to direct transcription of said transcribable nucleic acid molecule at a desired level or in a desired tissue or developmental pattern upon introduction of said construct into a plant cell.
  • the transcribable nucleic acid molecule comprises a protein-coding region of a gene
  • the promoter provides for transcription of a functional mRNA molecule that is translated and expressed as a protein product.
  • Constructs may also be constructed for transcription of antisense RNA molecules or other similar inhibitory RNA in order to inhibit expression of a specific RNA molecule of interest in a target host cell.
  • Exemplary transcribable nucleic acid molecules for incorporation into constructs of the present document include, for example, nucleic acid molecules or genes from a species other than the target gene species, or even genes that originate with or are present in the same species, but are incorporated into recipient cells by genetic engineering methods rather than classical reproduction or breeding techniques.
  • Exogenous gene or genetic element is intended to refer to any gene or nucleic acid molecule that is introduced into a recipient cell.
  • the type of nucleic acid molecule included in the exogenous nucleic acid molecule can include a nucleic acid molecule that is already present in the plant cell, a nucleic acid molecule from another plant, a nucleic acid molecule from a different organism, or a nucleic acid molecule generated externally, such as a nucleic acid molecule containing an antisense message of a gene, or a nucleic acid molecule encoding an artificial or modified version of a gene.
  • the promoters of the present document can be incorporated into a construct using marker genes as described, and tested in transient analyses that provide an indication of gene expression in stable plant systems.
  • the term "marker gene” refers to any transcribable nucleic acid molecule whose expression can be screened for or scored in some way. Methods of testing for marker gene expression in transient assays are known to those of skill in the art. Transient expression of marker genes has been reported using a variety of plants, tissues, plant cell(s), and DNA delivery systems.
  • types of transient analyses can include but are not limited to direct gene delivery via electroporation or particle bombardment of tissues in any transient plant assay using any plant species of interest.
  • transient systems would include, but are not limited to, electroporation of protoplasts from a variety of tissue sources or particle bombardment of specific tissues of interest.
  • the present document encompasses the use of any transient expression system to evaluate promoters or promoter fragments operably linked to any transcribable nucleic acid molecules, including but not limited to selected reporter genes, marker genes, or genes of agronomic interest.
  • plant tissues envisioned to test in transients via an appropriate delivery system would include, but are not limited to, leaf base tissues, callus, cotyledons, roots, endosperm, embryos, floral tissue, pollen, and epidermal tissue.
  • Promoters and control elements of the present document are useful for modulating metabolic or catabolic processes. Such processes include, but are not limited to, secondary product metabolism, amino acid synthesis, seed protein storage, increased biomass, oil development, pest defense and nitrogen usage.
  • expression constructs can be
  • each of these regulatory elements may be combined with the present vector if desired.
  • Translation of eukaryotic mRNA is often initiated at the codon that encodes the first methionine.
  • the vector of the present document may contain additional components.
  • an origin of replication allows for replication of the vector in a host cell.
  • homologous sequences flanking a specific sequence allow for specific recombination of the specific sequence at a desired location in the target genome.
  • T- DNA sequences also allow for insertion of a specific sequence randomly into a target genome.
  • the vector may also be provided with a plurality of restriction sites for insertion of a polynucleotide to be transcribed as well as the promoter and/or promoter control elements of the present document.
  • the vector may additionally contain selectable marker genes.
  • the vector may also contain a transcriptional and translational initiation region, and a transcriptional and translational termination region functional in the host cell.
  • the termination region may be native with the transcriptional initiation region, may be native with the polynucleotide to be transcribed, or may be derived from another source. Convenient termination regions are available from the Ti-plasmid of A.
  • tumefaciens such as the octopine synthase and nopaline synthase termination regions. See also, Guerineau et al. (1991) Mol. Gen. Genet. 262: 141-144; Proudfoot (1991) Cell 64:671-674; Sanfacon et al. (1991) Genes Dev. 5: 141-149; Mogen et al. (1990) Plant Cell 2: 1261-1272; Munroe et al. (1990) Gene 91 : 151-158; Ballas et al. (1989) Nucleic Acids Res. 17:7891-7903; Joshi et al. (1987) Nucleic Acid Res. 15:9627-9639.
  • the polynucleotide to be transcribed may be optimized for increased expression in a certain host cell.
  • the polynucleotide can be synthesized using preferred codons for improved transcription and translation. See U.S. Patent Nos. 5,380,831, 5,436,391; see also and Murray et al. (1989) Nucleic Acids Res. 17:477-498.
  • Additional sequence modifications include elimination of sequences encoding spurious polyadenylation signals, exon intron splice site signals, transposon- like repeats, and other such sequences well characterized as deleterious to expression.
  • the G-C content of the polynucleotide may be adjusted to levels average for a given cellular host, as calculated by reference to known genes expressed in the host cell.
  • the polynucleotide sequence may be modified to avoid hairpin secondary mR A structures.
  • luciferase expression vectors and luciferase gene cassettes are available from Promega Corp. (Madison, Wisconsin).
  • GFP vectors are available from Aurora Biosciences.
  • the promoters according to the present document can be inserted into a host cell.
  • a host cell includes but is not limited to a plant, mammalian, insect, yeast, and prokaryotic cell, preferably a plant cell.
  • the method of insertion into the host cell genome is chosen based on convenience.
  • the insertion into the host cell genome may either be accomplished by vectors that integrate into the host cell genome or by vectors which exist independent of the host cell genome.
  • the promoters of the present document can exist autonomously or independent of the host cell genome.
  • Vectors of these types are known in the art and include, for example, certain type of non-integrating viral vectors, autonomously replicating plasmids, artificial chromosomes, and the like.
  • transient expression of a promoter may be desired.
  • the promoter sequences, promoter control elements or vectors of the present document may be transformed into host cells. These transformations may be into protoplasts or intact tissues or isolated cells. Preferably expression vectors are introduced into intact tissue.
  • General methods of culturing plant tissues are provided for example by Maki et al. (1993) "Procedures for Introducing Foreign DNA into Plants” In Methods in Plant Molecular Biology & Biotechnology, Glich et al. Eds. pp. 67-88 CRC Press; and by Phillips et al. (1988) "Cell-Tissue Culture and In-Vitro Manipulation” In Corn & Corn Improvement, 3rd Edition Sprague et al. eds., pp. 345-387, American Society of
  • Methods of introducing polynucleotides into plant tissue include the direct infection or co-cultivation of plant cell with Agrobacterium tumefaciens, Horsch et al. (1985) Science, 227: 1229. Descriptions of Agrobacterium vector systems and methods for Agrobacterium-mediatGd gene transfer provided by Gruber et al. supra.
  • polynucleotides are introduced into plant cells or other plant tissues using a direct gene transfer method such as microprojectile-mediated delivery, DNA injection, electroporation and the like. More preferably polynucleotides are introduced into plant tissues using the microprojectile media delivery with the biolistic device. See, for example, Tomes et al., "Direct DNA transfer into intact plant cells via microprojectile bombardment” In: Gamborg and Phillips (Eds.) Plant Cell, Tissue and Organ Culture: Fundamental Methods, Springer Verlag, Berlin (1995).
  • Agrostis stolonifera Poa pratensis, Stenotaphrum secundatum
  • wheat Triticum aestivum
  • switchgrass Panicum vigatum
  • alfalfa Medicago sativa
  • polynucleotides and vectors described herein can be used to transform a number of monocotyledonous and dicotyledonous plants and plant cell systems, including species from one of the following families: Acanthaceae, Alliaceae, Alstroemeriaceae, Amaryllidaceae, Apocynaceae, Arecaceae, Asteraceae,
  • Berberidaceae Bixaceae, Brassicaceae, Bromeliaceae, Cannabaceae, Caryophyllaceae, Cephalotaxaceae, Chenopodiaceae, Colchicaceae, Cucurbitaceae, Dioscoreaceae, Ephedraceae, Erythroxylaceae, Euphorbiaceae, Fabaceae, Lamiaceae, Linaceae, Lycopodiaceae, Malvaceae, Melanthiaceae, Musaceae, Myrtaceae, Nyssaceae,
  • Suitable species may include members of the genus Abelmoschus, Abies,
  • Acer Agrostis, Allium, Alstroemeria, Ananas, Andrographis, Andropogon, Artemisia, Arundo, Atropa, Berberis, Beta, Bixa, Brassica, Calendula, Camellia, Camptotheca, Cannabis, Capsicum, Carthamus, Catharanthus, Cephalotaxus, Chrysanthemum, Cinchona, CitruUus, Coffea, Colchicum, Coleus, Cucumis, Cucurbita, Cynodon, Datura, Dianthus, Digitalis, Dioscorea, Elaeis, Ephedra, Erianthus, Erythroxylum, Eucalyptus, Festuca, Fragaria, Galanthus, Glycine, Gossypium, Helianthus, Hevea, Hordeum, Hyoscyamus, Jatropha, Lactuca, Linum, Lolium, Lupinus, Lycopersicon, Lycopodium, Manihot
  • Suitable species include Panicum spp. or hybrids thereof, Sorghum spp. or hybrids thereof, sudangrass, Miscanthus spp. or hybrids thereof, Saccharum spp. or hybrids thereof, Erianthus spp., Populus spp., Andropogon gerardii (big bluestem), Pennisetum purpureum (elephant grass) or hybrids thereof (e.g., Pennisetum purpureum x Pennisetum typhoidum), Phalaris arundinacea (reed canarygrass), Cynodon dactylon (bermudagrass), Festuca arundinacea (tall fescue), Spartina pectinata (prairie cord- grass), Medicago sativa (alfalfa), Arundo donax (giant reed) or hybrids thereof, Secale cereale (rye), Salix spp.
  • a suitable species can be a wild, weedy, or cultivated sorghum species such as, but not limited to, Sorghum almum, Sorghum amplum, Sorghum angustum, Sorghum arundinaceum, Sorghum bicolor (such as bicolor, guinea, caudatum, kafir, and durra), Sorghum brachypodum, Sorghum bulbosum, Sorghum burmahicum, Sorghum controversum, Sorghum drummondii, Sorghum ecarinatum, Sorghum exstans, Sorghum grande, Sorghum halepense, Sorghum interjectum, Sorghum intrans, Sorghum laxiflorum, Sorghum leiocladum, Sorghum macrospermum, Sorghum matarankense, Sorghum miliaceum, Sorghum nigrum, Sorghum nitidum, Sorghum plumosum, Sorghum prop
  • purpureosericeum Sorghum stipoideum, Sorghum sudanensese, Sorghum timorense, Sorghum trichocladum, Sorghum versicolor, Sorghum virgatum, Sorghum vulgare, or hybrids such as Sorghum x almum, Sorghum x sudangrass or Sorghum x drummondii.
  • Suitable species also include Helianthus annuus (sunflower), Carthamus tinctorius (safflower), Jatropha curcas (jatropha), Ricinus communis (castor), Elaeis guineensis (palm), Linum usitatissimum (flax), and Brassica juncea.
  • Suitable species also include Beta vulgaris (sugarbeet), and Manihot esculenta (cassava).
  • Suitable species also include Lycopersicon esculentum (tomato),
  • Lactuca sativa (lettuce), Musa paradisiaca (banana), Solanum tuberosum (potato), Brassica oleracea (broccoli, cauliflower, brusselsprouts), Camellia sinensis (tea), Fragaria ananassa (strawberry), Theobroma cacao (cocoa), Coffea arabica (coffee), Vitis vinifera (grape), Ananas comosus (pineapple), Capsicum annum (hot & sweet pepper), Allium cepa (onion), Cucumis melo (melon), Cucumis sativus (cucumber), Cucurbita maxima (squash), Cucurbita moschata (squash), Spinacea oleracea (spinach), Citrullus lanatus (watermelon), Abelmoschus esculentus (okra), and Solanum melongena (eggplant).
  • Suitable species also include Papaver somniferum (opium poppy),
  • Papaver orientale, Taxus baccata, Taxus brevifolia, Artemisia annua, Cannabis sativa, Camptotheca acuminate, Catharanthus roseus, Vinca rosea, Cinchona officinalis, Colchicum autumnale, Veratrum californica, Digitalis lanata, Digitalis purpurea, Dioscorea spp., Andrographis paniculata, Atropa belladonna, Datura stomonium, Berberis spp., Cephalotaxus spp., Ephedra sinica, Ephedra spp., Erythroxylum coca, Galanthus wornorii, Scopolia spp., Lycopodium serratum ( Huperzia errata), Lycopodium spp., Rauwolfia serpentina, Rauwolfia spp., Sanguinaria canadensis, Hyoscyamus spp., Calendula officinalis, Chr
  • Suitable species also include Parthenium argentatum (guayule), Hevea spp. (rubber), Mentha spicata (mint), Mentha piperita (mint), Bixa orellana, and Alstroemeria spp.
  • Suitable species also include Rosa spp. (rose), Dianthus caryophyllus
  • Petunia spp. (carnation), Petunia spp. (petunia) and Poinsettia pulcherrima (poinsettia).
  • Suitable species also include Nicotiana tabacum (tobacco), Lupinus albus (lupin), Uniola paniculata (oats), bentgrass (Agrostis spp.), Populus tremuloides (aspen), Pinus spp. (pine), Abies spp. (fir), Acer spp. (maple, Hordeum vulgare (barley), Poa pratensis (bluegrass), Lolium spp. (ryegrass) and Phleum pratense (timothy).
  • the methods and compositions can be used over a broad range of plant species, including species from the dicot genera Brassica, Carthamus, Glycine, Gossypium, Helianthus, Jatropha, Parthenium, Populus, and Ricinus; and the monocot genera Elaeis, Festuca, Hordeum, Lolium, Oryza, Panicum, Pennisetum, Phleum, Poa, Saccharum, Secale, Sorghum, Triticosecale, Triticum, and Zea.
  • a plant is a member of the species Panicum virgatum (switchgrass), Sorghum bicolor (sorghum, sudangrass), Miscanthus giganteus (miscanthus), Saccharum sp. (energycane), Populus balsamifera (poplar), Zea mays (corn), Glycine max (soybean), Brassica napus (canola), Triticum aestivum (wheat), Gossypium hirsutum (cotton), Oryza sativa (rice), Helianthus annuus (sunflower), Medicago sativa (alfalfa), Beta vulgaris (sugarbeet), or Pennisetum glaucum (pearl millet).
  • the polynucleotides and vectors described herein can be used to transform a number of monocotyledonous and dicotyledonous plants and plant cell systems, wherein such plants are hybrids of different species or varieties of a specific species (e.g., Saccharum sp. x Miscanthus sp., Panicum virgatum x Panicum amarum, Panicum virgatum x Panicum amarulum, and Pennisetum purpureum x Pennisetum typhoidum).
  • Saccharum sp. x Miscanthus sp. Panicum virgatum x Panicum amarum
  • Panicum virgatum x Panicum amarulum Panicum virgatum x Panicum amarulum
  • Pennisetum purpureum x Pennisetum typhoidum Pennisetum purpureum x Pennisetum typhoidum
  • expression constructs can be used for gene expression in callus culture for the purpose of expressing marker genes encoding peptides or polypeptides that allow identification of transformed plants.
  • a promoter that is operatively linked to a polynucleotide to be transcribed is transformed into plant cells and the transformed tissue is then placed on callus-inducing media. If the transformation is conducted with leaf discs, for example, callus will initiate along the cut edges. Once callus growth has initiated, callus cells can be transferred to callus shoot-inducing or callus root-inducing media.
  • callus root-inducing promoters will be activated on callus root-inducing media, etc.
  • Examples of such peptides or polypeptides useful as transformation markers include, but are not limited to barstar, glyphosate, chloramphenicol acetyltransferase (CAT), kanamycin, spectinomycin, streptomycin or other antibiotic resistance enzymes, green fluorescent protein (GFP), and ⁇ -glucuronidase (GUS), etc.
  • Some of the promoters provided in SEQ ID NOs: 1 - 18 will also be capable of sustaining expression in some tissues or organs after the initiation or completion of regeneration. Examples of these tissues or organs are somatic embryos, cotyledon, hypocotyl, epicotyl, leaf, stems, roots, flowers and seed.
  • Integration into the host cell genome also can be accomplished by methods known in the art, for example, by the homologous sequences or T-DNA discussed above or using the Cre-lox system (A.C. Vergunst et al. (1998) Plant Mol. Biol. 38:393).
  • the promoters of the present application can be used to further understand developmental mechanisms. For example, promoters that are specifically induced during callus formation, somatic embryo formation, shoot formation or root formation can be used to explore the effects of overexpression, repression or ectopic expression of target genes, or for isolation of trans-acting factors.
  • the vectors of the present application can be used not only for expression of coding regions but may also be used in exon-trap cloning, or promoter trap procedures to detect differential gene expression in various tissues (see Lindsey et al. (1993) Transgenic Research 2:3347. Auch and Reth (1990) Nucleic Acids Research 18: 6743).
  • Entrapment vectors first described for use in bacteria (Casadaban and
  • Entrapment vectors can be introduced into pluripotent ES cells in culture and then passed into the germline via chimeras (Gossler et al. 1989) Science 244: 463; Skarnes (1990) Biotechnology 8: 827).
  • Promoter or gene trap vectors often contain a reporter gene, e.g., lacZ, lacking its own promoter and/or splice acceptor sequence upstream. That is, promoter gene traps contain a reporter gene with a splice site but no promoter. If the vector lands in a gene and is spliced into the gene product, then the reporter gene is expressed.
  • IVET innate promoter traps
  • various bacterial genome fragments are placed in front of a necessary metabolic gene coupled to a reporter gene.
  • the DNA constructs are inserted into a bacterial strain otherwise lacking the metabolic gene, and the resulting bacteria are used to infect the host organism. Only bacteria expressing the metabolic gene survive in the host organism; consequently, inactive constructs can be eliminated by harvesting only bacteria that survive for some minimum period in the host.
  • broadly active constructs can be eliminated by screening only bacteria that do not express the reporter gene under laboratory conditions.
  • the bacteria selected by such a method contain constructs that are selectively induced only during infection of the host.
  • the IVET approach can be modified for use in plants to identify genes induced in either the bacteria or the plant cells upon pathogen infection or root colonization.
  • For information on IVET see the articles by Mahan et al. (1993) Science 259:686-688, Mahan et al. (1995) Proc. Natl. Acad. Sci. USA 92:669-673, Heithoff et al. (1997) Proc. Natl. Acad. Sci USA 94:934-939, and Wang et al. (1996) Proc. Natl. Acad. Sci USA 93: 10434.
  • a nucleic acid molecule as shown in SEQ ID NOs: 1 - 44 is incorporated into a construct such that a promoter of the present document is operably linked to a transcribable nucleic acid molecule that is a gene of agronomic interest.
  • the term "gene of agronomic interest” refers to a transcribable nucleic acid molecule that includes but is not limited to a gene that provides a desirable characteristic associated with plant morphology, physiology, growth and development, yield, nutritional enhancement, disease or pest resistance, or environmental or chemical tolerance. The expression of a gene of agronomic interest is desirable in order to confer an agronomically important trait.
  • a gene of agronomic interest that provides a beneficial agronomic trait to crop plants may be, for example, including, but not limited to genetic elements comprising herbicide resistance, increased yield, increased biomass, insect control, fungal disease resistance, virus resistance, nematode resistance, bacterial disease resistance, starch production, modified oils production, high oil production, modified fatty acid content, high protein production, fruit ripening, enhanced animal and human nutrition, biopolymers, environmental stress resistance, pharmaceutical peptides, improved processing traits, improved digestibility, industrial enzyme production, improved flavor, nitrogen fixation, hybrid seed production, and biofuel production.
  • the genetic elements, methods, and transgenes described in the patents listed above are hereby incorporated by reference.
  • a transcribable nucleic acid molecule can effect the above mentioned phenotypes by encoding a RNA molecule that causes the targeted inhibition of expression of an endogenous gene, for example via antisense, inhibitory RNA (RNAi), or cosuppression-mediated mechanisms.
  • the RNA could also be a catalytic RNA molecule (i.e., a ribozyme) engineered to cleave a desired endogenous mRNA product.
  • Promoters and control elements providing modulation of transcription under oxidative, drought, oxygen, wound, and methyl jasmonate stress are particularly useful for producing host cells or organisms that are more resistant to biotic and abiotic stresses.
  • modulation of genes, transcripts, and/or polypeptides in response to oxidative stress can protect cells against damage caused by oxidative agents, such as hydrogen peroxide and other free radicals.
  • Drought induction of genes, transcripts, and/or polypeptides are useful to increase the viability of a plant, for example, when water is a limiting factor.
  • genes, transcripts, and/or polypeptides induced during oxygen stress can help the flood tolerance of a plant.
  • the promoters and control elements of the present document can modulate stresses similar to those described in, for example, stress conditions are VuPLDl (drought stress; Cowpea; see Pham-Thi et al. (1999) Plant Mol Biol 39: 1257- 65), pyruvate decarboxylase (oxygen stress; rice; see Rivosal et al. (1997) Plant Physiol 114(3): 1021-29), chromoplast specific carotenoid gene (oxidative stress; Capsicum; see Bouvier et al. (1998) J Biol Chem 273: 30651-59).
  • Promoters and control elements providing preferential transcription during wounding or induced by methyl jasmonate can produce a defense response in host cells or organisms.
  • preferential modulation of genes, transcripts, and/or polypeptides under such conditions is useful to induce a defense response to mechanical wounding, pest or pathogen attack or treatment with certain chemicals.
  • Promoters and control elements of the present document also can trigger a response similar to those described for cf9 (viral pathogen; tomato; see O'Donnell et al. (1998) Plant J 14(1): 137-42), hepatocyte growth factor activator inhibitor type 1 (HAI- 1), which enhances tissue regeneration (tissue injury; human; Koono et al. (1999) J Histochem Cytochem 47: 673-82), copper amine oxidase (CuAO), induced during ontogenesis and wound healing (wounding; chick-pea; Rea et al. (1998) FEBS Lett 437: 177-82), proteinase inhibitor II (wounding; potato; see Pena-Cortes et al.
  • HAI-1 hepatocyte growth factor activator inhibitor type 1
  • CuAO copper amine oxidase
  • genes, transcripts, and/or polypeptides that increase oxidative, flood, or drought tolerance may require up-regulation of transcription.
  • promoter or control elements which provide preferential transcription in wounding or under methyl jasmonate induction, produce transcript levels that are statistically significant as compared to cell types, organs or tissues under other conditions.
  • promoter and control elements produce transcript levels that are above background of the assay.
  • Promoters and control elements providing preferential transcription when induced by light exposure can be utilized to modulate growth, metabolism, and development; to increase drought tolerance; and decrease damage from light stress for host cells or organisms.
  • modulation of genes, transcripts, and/or polypeptides in response to light is useful
  • the promoters and control elements of the present document also can trigger responses similar to those described in: abscisic acid insensitive3 (ABI3) (dark- grown Arabidopsis seedlings, see Rohde et al. (2000) Plant Cell 12: 35-52), asparagine synthetase (pea root nodules, see Tsai and Coruzzi (1990) EMBO J9: 323-32), mdm2 gene (human tumor, see Saucedo et al. (1998) Cell Growth Differ 9: 119-30).
  • ABS3 abscisic acid insensitive3
  • genes, transcripts, and/or polypeptides that increase drought or light tolerance may require up-regulation of transcription.
  • promoter or control elements which provide preferential transcription in cells, tissues or organs exposed to light, produce transcript levels that are statistically significant as compared to cells, tissues, or organs under decreased light exposure (intensity or length of time).
  • promoter and control elements For preferential up-regulation of transcription, promoter and control elements produce transcript levels that are above background of the assay.
  • Promoters and control elements providing preferential transcription when induced by dark or decreased light intensity or decreased light exposure time can be utilized to time growth, metabolism, and development, to modulate photosynthesis capabilities for host cells or organisms.
  • modulation of genes, transcripts, and/or polypeptides in response to dark is useful, for example, (1) to induce growth or development, such as fruit development and maturity, despite lack of light;
  • present promoters and control elements can also trigger response similar to those described in the section above.
  • Up-regulation and down-regulation of transcription are useful for these applications.
  • genes, transcripts, and/or polypeptides that increase or decrease growth and development may require up-regulation of transcription.
  • promoter or control elements which provide preferential transcription under exposure to dark or decrease light intensity or decrease exposure time, produce transcript levels that are statistically significant.
  • promoter and control elements For preferential up-regulation of transcription, promoter and control elements produce transcript levels that are above background of the assay.
  • Promoters and control elements providing preferential transcription in a leaf can modulate growth, metabolism, and development or modulate energy and nutrient utilization in host cells or organisms.
  • preferential modulation of genes, transcripts, and/or polypeptide in a leaf is useful, for example,
  • Up-regulation and down-regulation of transcription are useful for these applications.
  • genes, transcripts, and/or polypeptides that increase growth, for example, may require up-regulation of transcription.
  • promoter or control elements which provide preferential transcription in the cells, tissues, or organs of a leaf, produce transcript levels that are statistically significant as compared to other cells, organs or tissues.
  • promoter and control elements For preferential up-regulation of transcription, produce transcript levels that are above background of the assay.
  • Promoters and control elements providing preferential transcription in a root can modulate growth, metabolism, development, nutrient uptake, nitrogen fixation, or modulate energy and nutrient utilization in host cells or organisms.
  • preferential modulation of genes, transcripts, and/or polypeptide in a root is useful,
  • Up-regulation and down-regulation of transcription are useful for these applications.
  • genes, transcripts, and/or polypeptides that increase or decrease growth, for example, may require up-regulation of transcription.
  • promoter or control elements which provide preferential transcription in cells, tissues, or organs of a root, produce transcript levels that are statistically significant as compared to other cells, organs or tissues.
  • promoter and control elements For preferential up-regulation of transcription, promoter and control elements produce transcript levels that are above background of the assay.
  • Promoters and control elements providing preferential transcription in a stem or shoot can modulate growth, metabolism, and development or modulate energy and nutrient utilization in host cells or organisms.
  • preferential modulation of genes, transcripts, and/or polypeptide in a stem or shoot is useful, for example,
  • Up-regulation and down-regulation of transcription are useful for these applications.
  • genes, transcripts, and/or polypeptides that increase growth, for example, may require up-regulation of transcription.
  • promoter or control elements which provide preferential transcription in the cells, tissues, or organs of a stem or shoot, produce transcript levels that are statistically significant as compared to other cells, organs or tissues.
  • promoter and control elements For preferential up-regulation of transcription, promoter and control elements produce transcript levels that are above background of the assay.
  • Promoters and control elements providing preferential transcription in a silique or fruit can time growth, development, or maturity; or modulate fertility; or modulate energy and nutrient utilization in host cells or organisms.
  • preferential modulation of genes, transcripts, and/or polypeptides in a fruit is useful
  • seeds such as, storage molecules, starch, protein, oil, vitamins, anti-nutritional components, such as phytic acid;
  • compositions such as lysine rich proteins
  • Up-regulation and down-regulation of transcription are useful for these applications. For instance, genes, transcripts, and/or polypeptides that increase or decrease growth, for example, may require up-regulation of transcription.
  • promoter or control elements which provide preferential transcription in the cells, tissues, or organs of siliques or fruits, produce transcript levels that are statistically significant as compared to other cells, organs or tissues.
  • promoter and control elements For preferential up-regulation of transcription, promoter and control elements produce transcript levels that are above background of the assay.
  • Promoters and control elements providing preferential transcription in a callus can be useful to modulating transcription in dedifferentiated host cells.
  • preferential modulation of genes, transcripts, in callus is useful to modulate transcription of a marker gene, which can facilitate selection of cells that are transformed with exogenous polynucleotides.
  • genes, transcripts, and/or polypeptides that increase marker gene detectability may require up-regulation of transcription.
  • promoter and control elements produce transcript levels that are above background of the assay.
  • Promoters and control elements providing preferential transcription in flowers can modulate pigmentation; or modulate fertility in host cells or organisms.
  • preferential modulation of genes, transcripts, and/or polypeptides in a flower is useful,
  • genes, transcripts, and/or polypeptides that increase or decrease pigmentation may require up-regulation of transcription
  • promoter or control elements which provide preferential transcription in flowers, produce transcript levels that are statistically significant as compared to other cells, organs or tissues.
  • promoter and control elements For preferential up-regulation of transcription, promoter and control elements produce transcript levels that are above background of the assay.
  • Promoters and control elements providing preferential transcription in an immature bud/floret or inflorescence can time growth, development, or maturity; or modulate fertility or viability in host cells or organisms.
  • preferential modulation of genes, transcripts, and/or polypeptide in an immature bud and/or inflorescence is useful,
  • Up-regulation and down-regulation of transcription are useful for these applications.
  • genes, transcripts, and/or polypeptides that increase or decrease growth, for example, may require up-regulation of transcription.
  • promoter or control elements which provide preferential transcription in immature buds/florets and inflorescences, produce transcript levels that are statistically significant as compared to other cell types, organs or tissues.
  • promoter and control elements For preferential up-regulation of transcription, promoter and control elements produce transcript levels that are above background of the assay.
  • Promoters and control elements providing preferential transcription during senescence can be used to modulate cell degeneration, nutrient mobilization, and scavenging of free radicals in host cells or organisms.
  • Other types of responses that can be modulated include, for example, senescence associated genes (SAG) that encode enzymes thought to be involved in cell degeneration and nutrient mobilization (Arabidopsis; see Hensel et al. (1993) Plant Cell 5: 553-64), and the CP-2/cathepsin L gene (rat; Kim and Wright (1997) Biol Reprod 57: 1467-77), both induced during senescence.
  • SAG senescence associated genes
  • CP-2/cathepsin L gene rat; Kim and Wright (1997) Biol Reprod 57: 1467-77
  • Up-regulation and down-regulation of transcription are useful for these applications.
  • genes, transcripts, and/or polypeptides that increase or decrease scavenging of free radicals, for example, may require up-regulation of transcription.
  • promoter or control elements which provide preferential transcription in cells, tissues, or organs during senescence, produce transcript levels that are statistically significant as compared to other conditions.
  • promoter and control elements For preferential up-regulation of transcription, promoter and control elements produce transcript levels that are above background of the assay.
  • Promoters and control elements providing preferential transcription in a germinating seed can time growth, development, or maturity; or modulate viability in host cells or organisms.
  • preferential modulation of genes, transcripts, and/or polypeptide in a germinating seed is useful,
  • Up-regulation and down-regulation of transcription are useful for these applications.
  • genes, transcripts, and/or polypeptides that increase or decrease growth, for example, may require up-regulation of transcription.
  • promoter or control elements which provide preferential transcription in a germinating seed, produce transcript levels that are statistically significant as compared to other cell types, organs or tissues.
  • promoter and control elements produce transcript levels that are above background of the assay.
  • Wassilewskija (WS) plants are transformed with Ti plasmids containing nucleic acid sequences to be expressed, as noted in the respective examples, in the sense orientation relative to the 35S promoter in a Ti plasmid.
  • a Ti plasmid vector useful for these constructs, CRS 338 contains the Ceres-constructed, plant selectable marker gene phosphinothricin acetyltransferase (PAT), which confers herbicide resistance to transformed plants.
  • PAT phosphinothricin acetyltransferase
  • Ten independently transformed events are typically selected and evaluated for their qualitative phenotype in the Ti generation.
  • Horticulture, Ltd., Bellevue, WA is mixed with 16L Therm-O-Rock vermiculite (Therm- O-Rock West, Inc., Chandler, AZ) in a cement mixer to make a 60:40 soil mixture.
  • To the soil mixture is added 2 Tbsp Marathon 1% granules (Hummert, Earth City, MO), 3 Tbsp OSMOCOTE® 14-14-14 (Hummert, Earth City, MO) and 1 Tbsp Peters fertilizer 20-20-20 (J.R. Peters, Inc., Allentown, PA), which are first added to 3 gallons of water and then added to the soil and mixed thoroughly.
  • 4-inch diameter pots are filled with soil mixture. Pots are then covered with 8-inch squares of nylon netting.
  • Plant Maintenance 3 to 4 days after planting, lids and shade cloth are removed. Plants are watered as needed. After 7-10 days, pots are thinned to 20 plants per pot using forceps. After 2 weeks, all plants are subirrigated with Peters fertilizer at a rate of 1 Tsp per gallon of water. When bolts are about 5-10 cm long, they are clipped between the first node and the base of stem to induce secondary bolts. Dipping
  • infiltration is performed 6 to 7 days after clipping.
  • Agrobacterium starter blocks are obtained (96-well block with
  • Agrobacterium cultures grown to an OD 60 o of approximately 1.0) and inoculated one culture vessel per construct by transferring 1 mL from appropriate well in the starter block. Cultures are then incubated with shaking at 27°C. Cultures are spun down after attaining an OD 6 oo of approximately 1.0 (about 24 hours). 200 mL infiltration media is added to resuspend Agrobacterium pellets. Infiltration media is prepared by adding 2.2 g MS salts, 50 g sucrose, and 5 2 mg/ml benzylaminopurine to 900 ml water.
  • Tissues are dissected by eye or under magnification using INOX 5 grade forceps and placed on a slide with water and coversliped. An attempt is made to record images of observed expression patterns at earliest and latest stages of development of tissues listed below. Specific tissues will be preceded with High (H), Medium (M), Low (L) designations. Flower Pedicel, receptacle, nectary, sepal, petal, filament, anther, pollen, carpel, style, papillae, vascular, epidermis, stomata, trichome
  • Ovule Pre- fertilization inner integument, outer integument, embryo sac,
  • Post- fertilization zygote, inner integument, outer integument, seed coat, primordia, chalaza, micropyle, early endosperm, mature endosperm, embryo
  • Embryo Suspensor preglobular, globular, heart, torpedo, late mature, pro vascular, hypophysis, radicle, cotyledons, hypocotyl
  • Leaf Petiole mesophyll, vascular, epidermis, trichome, primordia, stomata, stipule, margin
  • Tl Mature These are the Tl plants resulting from independent transformation events. These are screened between stage 6.50-6.90 (i.e. the plant is flowering and 50-90% of the flowers that the plant will make have developed), which is 4-6 weeks of age. At this stage the mature plant possesses flowers, siliques at all stages of development, and fully expanded leaves. The plants are initially imaged under UV with a Leica Confocal microscope to allow examination of the plants on a global level. If expression is present, they are re -imaged using scanning laser confocal microscopy.
  • T2 Seedling Progeny are collected from the Tl plants giving the same expression pattern and the progeny (T2) are sterilized and plated on agar-solidified medium containing M&S salts. In the event that there is no expression in the Tl plants, T2 seeds are planted from all lines. The seedlings are grown in Percival incubators under continuous light at 22°C for 10-12 days. Cotyledons, roots, hypocotyls, petioles, leaves, and the shoot meristem region of individual seedlings were screened until two seedlings were observed to have the same pattern. In general, the same expression pattern was found in the first two seedlings. However, up to 6 seedlings were screened before "no expression pattern" was recorded. All constructs are screened as T2 seedlings even if they did not have an expression pattern in the Tl generation.
  • T2 Mature The T2 mature plants were screened in a similar manner to the Tl plants. The T2 seeds were planted in the greenhouse, exposed to selection and at least one plant screened to confirm the Tl expression pattern. In instances where there were any subtle changes in expression, multiple plants were examined and the changes noted in the tables.
  • T3 Seedling This was done similar to the T2 seedlings except that only the plants for which we are trying to confirm the pattern are planted.
  • Nicotinic acid 2.8 g/L Proline, 300 mg/L Casamino acid, 30 g/L Sucrose, 2 mg/L 2, 4- Dichloro-Phenoxyacetic Acid, 4g/L Gel rite, pH 5.6); 10 seeds each Petri dish. Each Petri dish contains 30 ml N6-P medium. Dishes are sealed with antifungal tape to allow air exchange. Place the plates at 28°C under cold fluorescent light. Many granular calli should be formed within 4 weeks. Calli of good quality consist of small and spherical cells with dense cytoplasm, which are competent for transformation. The calli can be used directly for Agrobacterium infection, or subculture them for later use. Infection and co-cultivation of calli with Agrobacterium cells
  • YEB liquid medium by growing it overnight in a shaker. Appropriate antibiotics are included at 50 mg/L or higher. Put on 28°C shaker overnight. The next day, reinoculate 25 xL of overnight culture into 5 mL of liquid YEB with selection and grow overnight at 28°C. The next day, use this culture for transformation. Transfer the liquid culture to 1.5 mL microtube and centrifuge it at 10,000 RPM for 2 minutes.
  • N6-AS liquid medium (3.98g/L N6 basal salt mixture, 0.8 mg/L KI, 0.025 mg/L CoCl 2 .6H 2 0, 0.025 mg/L CuSO 4 .5H 2 0, 0.25 mg/L NaMoO 4 .2H 2 0, 2 mg/L Glycine, lOOmg/L Myo-inositol, 5 mg/L Thiamine.HCl, 1 mg/L Pyridoxine.HCl, 1 mg/L Nicotinic acid, 1 g/L Casamino acid, 30 g/L Sucrose, 10 g/L Glucose, 2 mg/L 2, 4- Dichloro-Phenoxyacetic Acid, pH 5.6).
  • the optimal OD 6 oo reading varies significantly depending on Agrobacterium strains and sometimes vectors. The optimal reading means that there is no over-growth of
  • the optimum OD 6 oo for rice is 0.2.
  • Each 100 mm x 20 mm dish contains 10-20 mL N6-AS media. Seal the dish with antifungal tape. Cover plates with aluminum foil because acetosyringone present in the media is light sensitive. Co-culture calli and Agrobacterium cells at 22°C in the dark until
  • Agrobacterium mass can be seen by the naked eye.
  • KimwipesTM paper in a large spherical Petri plate to blot Culture calli on dishes containing semisolid N6-P medium with 250mg/L carbenicillin at 28°C under cold fluorescent light for 6 days. Each 100mm x 20mm dish contains 30 mL N6-P medium, use approximately 4 dishes of calli for each construct. Dishes are sealed with antifungal tape. Transfer calli from the resting media on to the selection media containing 250mg/L carbenicillin and 5 mg/L purified BialaphosTM (for selection of BAR gene) or 100 mg/L Paromomycin sulfate (for seclection of NPTII gene) for 14 days. For second round of selection, subculture calli for another 14 days. Transformed calli can typically be seen clearly at the end of this selection. Third selection is done if the second selection does not produce enough resistant calli.
  • N6-R plant regeneration medium (3.98 g/L N6 basal salt mixture, 0.8 mg/L KI, 0.025 mg/L CoCL 2 .6H 2 0, 0.025 mg/L CuSO 4 .5H 2 0, 0.25 mg/L NaMoO 4 .2H 2 0, 2 mg/L Glycine, 100 mg/L Myo-inositol, 5 mg/L Thiamine.HCl, 1 mg/L Pyridoxine.HCl, 1 mg/L Nicotinic acid, 1 g/L Casamino acid, 25 g/L Sucrose, 25 g/L Sorbitol, 2 mg/L 6-Benzylaminopurine, 0.05 mg/L 1- Naphthaleneacetic acid, 7 g/L Agarose (Omnipur), pH 5.6).
  • Each 100mm x 20mnm dish contains 30 mL N6-R medium, 4 or 6 callus lines on each dish. Dishes are sealed with antifungal tape. Culture calli at 28°C under cold fluorescent light until shoots and roots are formed. Typically, shoots should be seen within 3 weeks. Transfer plantlets to Magenta boxes containing 30 mL 1 ⁇ 2 MSI A (2.165 g/L MS salts, lml/L of lOOOx B5 vitamins stock, 15 g/L Sucrose, 5 g/L Agar, pH 5.7). Grow plantlets at 28°C under old fluorescent light for 10-14 days.
  • Soil, Chino, CA is mixed with 4L Turface in a cement mixer to make a 60:40 soil mixture.
  • 1 tsp Marathon 1% granules Hummert, Earth City, MO
  • 2 Tbsp OSMOCOTE® 14-14-14 Hummert, Earth City, MO.
  • Tbsp Peters fertilizer 20-20-20 J.R. Peters, Inc., Allentown, PA
  • 6-inch diameter Azalea pots are used for transplanting with 1-2 plants per pot.
  • Plant Maintenance Plants are well watered throughout the duration of the lifecycle. The bottom of the flat is cleaned and new water added twice a week.
  • GFP Green Fluorescent Protein
  • GFP expression in rice callus can be observed as early as 4-7 days after co-cultivation.
  • the rice callus used for co-cultivation is observed under Zeiss Stemi SVII APO dissecting microscope for GFP expression.
  • GFP 500 filter for viewing GFP expression we use GFP 500 filter in the microscope.
  • the images observed under the microscope can be transferred, captured and stored to a computer using the Axiocam (Zeiss) camera and Axiovision software.
  • Each independently transformed event is divided into single tillers which then undergo Typhoon scanner laser imaging.
  • One positively GFP-expressing tiller per event is selected for subsequent GFP analysis by Confocal microscopy and ultimately for transplantation for further mature tissue analysis.
  • Typhoon Scan Plants are initially scanned with a Typhoon Scanner to examine the GFP expression of the plants on a global level. If expression is present, images are collected by Typhoon scanning laser imaging and scanning laser confocal microscopy. Scanned images from the Typhoon scanner are taken as 2-D images of the entire plant and can be opened using the program ImageQuant.
  • Ziess UV stereoscope Reproductive tissues that are too large to use with the confocal microscope are prepared using a dissection microscope under high magnification using INOX 5 grade forceps and placed on a slide. An attempt is made to record images of observed expression patterns in mature rice reproductive tissues.
  • TO plants resulting from a single tiller from each independent transformation event having predetermined positive GFP expression are screened between stage 3-5 (i.e. between late vegetative to panicle initiation and floral maturation), which is 6-8 weeks of age. At this stage the mature plant possesses young panicle inflorescence to adolescent flowers, fully expanded leaves, multiple nodes and mature stem and root tissue.
  • stage 3-5 i.e. between late vegetative to panicle initiation and floral maturation
  • the plants are initially imaged using the Typhoon scanner and then imaged in detail using the Leica Confocal microscope and Ziess UV Stereoscope to allow examination of the mature plants on a global level.
  • Seed is collected from the TO plants and stored for further use in induction experiments.
  • Tl Mature expression Expression is observed specifically from the ovule.
  • GenBank At5g24910 similar to Cytochrome P450 72A1 (SP:Q05047) [Catharanthus roseus]; similar to fatty acid omega-hydroxylase cytochrome P450 4A11 - Homo sapiens, PIRT53015; supported by cDNA: gi_16604323_gb_AY058060.1_; go function: cytochrome P450 activity
  • this promoter sequence could be useful to engineer enhancements to seed, including, but not limited to, alterations in germination timing, abiotic and biotic tolerances, and seed size, and to engineer seed sterility. Analysis of Promoter PD3536 activity
  • GenBank contains SRF-type transcription factor (DNA-binding and dimerisation domain); go function: DNA binding [goid 0003677]; go function: transcription factor activity [goid
  • this promoter sequence could be useful to engineer modifications to early embryo and seedling development. Analysis of Promoter PD3558 activity
  • Promoter candidate LP 71811437
  • Tl Mature expression Expression observed in green tissues.
  • GenBank Photosystem I reaction center subunit V, chloroplast precursor (PSI-G) (Photosystem
  • go_process photosynthesis [goid 0015979]
  • Source Promoter Organism Populus balsamifera subsp. trichocarpa
  • this promoter sequence could be useful to engineer increased biomass , among other things, increasing photosynthetic efficiency.
  • Promoter candidate LP 75181013
  • Tl Mature expression Expression was detected in a stem-specific manner, in all tissues, in the mature plant.
  • Selection Criteria Predicted to be stem-specific.
  • GenBank Pv clone 1806607 cellulose synthase-like family C; pfam: Glycosyl transferase family 2
  • Sub-trait Area Nitrogen use efficiency, phosphate use efficiency, Increased sugar accessibility,
  • this promoter sequence could be useful to engineer pathways that enhance the distribution of nutrients and/or water from the roots to the green tissues of the plant. It could also be used to specifically target the modulation of cell-wall genes to, among other things, modulate components involved in regulating energy production from the plant, such as, for example, enhancing accessible sugars. Analysis of Promoter PD3421 activity
  • Promoter candidate LP 58417441
  • Events 1-8 were screened for above ground tissues only, and the low-level of stem expression was not observed by the whole-plant scanner. Events 9-16 were screened for above and below ground expression, and the strength of the root-expression lead to confocal screening of the stem.
  • Tl Mature expression Expression was detected in the vasculature of the root and stem, but not in the leaves.
  • GenBank locus tag At3g 16920
  • n 16 Events
  • n 14 (1-2,4-6,8-16) FLOWER
  • Tl Mature expression Expression was detected in both above ground and below ground vegetative tissues, in both the epidermal, cortical, and vasculature tissues. No expression detected in pollen or seed.
  • Gen Bank At4g30650 similar to SP
  • go_process response to cold [goid 0009409]; go_process:
  • this promoter sequence could be useful to engineer agronomic trait improvements in both above ground (plant architecture, biomass yield, photosynthetic efficiency, herbivory resistance and disease resistance, among others) and below ground tissues (root architecture, insect and disease resistance, and water and nutrient uptake from soils, among others) while minimizing expression in gametes and germplasm, thus minimizing the exposure of expressed trait protein to the environment
  • Promoter candidate LP 58417447
  • Events 1-8 were analyzed for above-ground expression only. Events 9-16 were analyzed for whole-plant expression.
  • Tl Mature expression Expression was detected within the vasculature of the root
  • Selection Criteria Nominated based upon microarray data. The cDNA downstream of this promoter element is predicted to be highly expressed in stems.
  • Sub-trait Area Nitrogen Uptake, Phosphorous Uptake, Water Uptake
  • this promoter sequence could be useful to engineer increased volume or efficiency of nutrient and/or water uptake and transport from soil and into the above ground regions of the plant. Analysis of Promoter PD3333 activity
  • Promoter candidate LP 57007779
  • Events 1-8 were screened for above ground expression only. Events 9-15 were scored for root expression.
  • Tl Mature expression Expression was detected exclusively in the root; specifically in the outer layers of root tissue, including the epidermis and exodermis.
  • microarray data suggest root-specific expression of the mRNA downstream of this promoter element.
  • GenBank AT3G16450 similar to SP
  • ⁇ Arabidopsis thaliana ⁇ contains Pfam profile: PF01419 jacalin-like lectin domain
  • this promoter sequence could be useful to engineer pathways to modulate root growth and the uptake of substances from the soil, including, but not limited to, water, nitrogen, phosphorous, and other minerals. It could also be used to engineer traits that minimize pathogen invasion, including microbial and nematode infections. Analysis of Promoter PD3407 activity
  • GFP expression was detected in the cortical fiber cells, the endodermis, and the vasculature of the root.
  • GenBank Ole e I family protein
  • Sub-trait Area Enhanced water uptake, Enhanced nitrogen uptake
  • this promoter sequence could be useful to improve: the growth and architecture of roots, the strength of the root system, and the transport of water and nutrients into the above ground portion of the plant for enhanced biomass. Analysis of Promoter PD3389 activity
  • Sub-trait Area Salt tolerance, Drought tolerance, Phosphate and Nitrate Use Efficiency, Phosphate and Nitrate Utilization
  • this promoter sequence could be useful to improve: the biomass of the plants under normal and stressful conditions through the overexpression of trait- specific transgenes.
  • Promoter candidate I.D 58350521
  • Promoter candidate LP 89744767
  • GenBank pfam: Ubiquitin; go function: protein degradation;
  • Source Promoter Organism Sorghum bicolor
  • Sub-trait Area Salt tolerance, Drought tolerance, Phosphate and Nitrate Use Efficiency, Phosphate and Nitrate Utilization, Plant Architecture, Enhanced Photosynthesis, Cell-wall composition and conversion, Male sterility, Female sterility, Total sterility
  • this promoter sequence could be useful to improve: the biomass of the plants under normal and stressful conditions through the overexpression of transgenes that improve water use, nutrient use, alter composition, alter plant architecture, disrupt reproductive biology, and improve the carbon-nitrogen balance.
  • GenBank PFAM AP2; Herpes_gp2;
  • Source Promoter Organism Sorghum bicolor
  • Sub-trait Area Total sterility
  • this promoter sequence could be useful to engineer sterility through the expression of genes that regulate floral development. Reduced fertility may also lead to increased biomass, increased sugar content and inhibit ergot infection in sorghum.
  • Promoter candidate LP 75273275
  • GenBank PFAM FLO LFY; LFY floral meristem identity control protein
  • Source Promoter Organism Sorghum bicolor
  • Sub-trait Area Male sterility, Female sterility, Total sterility
  • this promoter sequence could be useful to engineer sterility through the expression of genes that regulate floral development. Reduced fertility may also lead to increased biomass, increased sugar content and inhibit ergot infection in sorghum.
  • Promoter candidate LP 89744768
  • GenBank pfam: Ubiquitin; go function: protein degradation;
  • Source Promoter Organism Sorghum bicolor
  • Sub-trait Area Salt tolerance, Drought tolerance, Phosphate and Nitrate Use Efficiency, Phosphate and Nitrate Utilization, Plant Architecture, Enhanced Photosynthesis, Cell- wall composition and conversion, Male sterility, Female sterility, Total sterility
  • this promoter sequence could be useful to improve: the biomass of the plants under normal and stressful conditions through the overexpression of transgenes that improve water use, nutrient use, alter composition, alter plant architecture, disrupt reproductive biology, and improve the carbon-nitrogen balance.
  • promoter PD3423 (SEQ ID NO:33), expression in Arabidopsis was characterized as above ground, vegetative.
  • promoter PD3801 (SEQ ID NO:43),
  • PD3428 1 1 Arabidopsis EMBRYO,OVULE,MATURE ROOT, LEAF, STEM, Biomass, yield
  • PD3695 30 Oryza sativa SPIKELET,LEAF,MAIN CULM,ROOT Plant size, Low nitrogen tolerance
  • Nitrogen use efficiency Nitrogen utilization, Biomass, Disease resistance
  • PD2946 25 Arabidopsis SILIQUE,FLOWER COLD,HEAT, Drought, biomass, cold, heat, yield thaliana - DROUGHT
  • Nitrogen use efficiency yield Nitrogen utilization
  • Photosynthetic efficiency biomass C/N Partitioning
  • Cold growth Freezing tolerance
  • Chronic drought Growth rate
  • Plant architecture Early flowering
  • PD3548, PD3408, PD3492, PD3092, PD3428, PD3395, PD3583, PD3607, PD3716, PD3623, PD3684, PD3689, PD3407, PD3490, PD3701, PD3777, PD3343, PD3201, PD2946, PD2998, PD3389, PD3324, PD3304, PD3695, P02916, PD3505, PD3423, PD3558, PD3536, PD3485, PD3333, PD3422, PD3540, PD3421, PD3684, PD3492, PD3801, or PD3579 are generated by introducing mutations into the nucleotide sequence set forth in SEQ ID NO: l-SEQ ID NO:44 as disclosed in U.S.
  • Each of the plant transformation vectors are prepared essentially as described above, except that the full length promoter is replaced by a mutagenized derivative.
  • Plants e.g., rice, switchgrass, or Arabidopsis
  • Plants are transformed with each of the plant transformation vectors and analyzed for expression of the GFP marker to identify those mutagenized derivatives having promoter activity.
  • PD3548, PD3408, PD3492, PD3092, PD3428, PD3395, PD3583, PD3607, PD3716, PD3623, PD3684, PD3689, PD3407, PD3490, PD3701, PD3777, PD3343, PD3201, PD2946, PD2998, PD3389, PD3324, PD3304, PD3695, P02916, PD3505, PD3423, PD3558, PD3536, PD3485, PD3333, PD3422, PD3540, PD3421, PD3684, PD3492, PD3801, or PD3579 are isolated by designing primers to clone fragments of the promoters set forth in SEQ ID NO: l-44.
  • a plurality of cloned fragments of PD3796, PD3800, PD3579, PD2695, YP2959, PD3256, PD3548, PD3408, PD3492, PD3092, PD3428, PD3395, PD3583, PD3607, PD3716, PD3623, PD3684, PD3689, PD3407, PD3490, PD3701, PD3777, PD3343, PD3201, PD2946, PD2998, PD3389, PD3324, PD3304, PD3695, P02916, PD3505, PD3423, PD3558, PD3536, PD3485, PD3333, PD3422, PD3540, PD3421, PD3684, PD3492, PD3801, or PD3579 ranging in size from 50 nucleotide up to the full length sequence set forth in SEQ ID NO: 1-44 are obtained
  • a fragment of PD3796 of about 50, 100, 150, 200, 250, 300, 350, 400, 450, 500, 550, 600, 650, 700, 750, 800, 850, 900, 950, 975, 1000, 1050, 1100, 1150, 1200, 1250, 1300, 1350, 1400, 1450, 1475 or 1490 nucleotides in length from various parts of PD3796 (SEQ ID NO: l) are obtained and inserted into a plant transformation vector operably linked to a GFP marker gene.
  • Such fragments of PD3796 can include one or more of a TATABOX4, GAREAT, and CARGCW8GAT motif
  • PD3800 450, 500, 550, 600, 650, 700, 750, 800, 850, 900, 950, 975, 1000, 1050, 1100, 1150, 1200, 1250, 1300, 1350, 1400, 1450, 1475 or 1490 nucleotides in length from various parts of PD3800 (SEQ ID NO:2) are obtained and inserted into a plant transformation vector operably linked to a GFP marker gene.
  • Such fragments of PD3800 can include one or more of a CARGNCAT and one or more of a CARGCW8GAT motif.
  • 375, or 390 nucleotides in length from various parts of PD3560 are obtained and inserted into a plant transformation vector operably linked to a GFP marker gene.
  • Such fragments of PD3560 can include one or more of an AREl, SBOXATRBCS, TE2F2NTPCNA, GADOWNAT, ACGTABREMOTIFA20SEM, and TATABOX motif.
  • PD3561 nucleotide sequence derived from various parts of PD3561 (SEQ ID NO:4) are obtained and inserted into a plant transformation vector operably linked to a GFP marker gene.
  • Such fragments of PD3561 can include one or more of a
  • YP2959 SEQ ID NO:5
  • Such fragments of YP2959 can include one or more of an ATHB1ATCONSENSUS, GAREAT, P1BS, GARE20SREP 1 , ABRE, and TATC-box motif.

Landscapes

  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Genetics & Genomics (AREA)
  • Chemical & Material Sciences (AREA)
  • Organic Chemistry (AREA)
  • Engineering & Computer Science (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Biochemistry (AREA)
  • Biotechnology (AREA)
  • General Engineering & Computer Science (AREA)
  • Molecular Biology (AREA)
  • Biophysics (AREA)
  • Biomedical Technology (AREA)
  • General Health & Medical Sciences (AREA)
  • Wood Science & Technology (AREA)
  • Zoology (AREA)
  • Plant Pathology (AREA)
  • Physics & Mathematics (AREA)
  • Microbiology (AREA)
  • Cell Biology (AREA)
  • Botany (AREA)
  • Gastroenterology & Hepatology (AREA)
  • Medicinal Chemistry (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Breeding Of Plants And Reproduction By Means Of Culturing (AREA)
  • Micro-Organisms Or Cultivation Processes Thereof (AREA)

Abstract

La présente invention concerne des séquences de promoteur et des éléments de contrôle de promoteur, des constructions polynucléotidiques comprenant les promoteurs et les éléments de contrôle, et des procédés d'identification des promoteurs, des éléments de contrôle ou de leurs fragments. L'invention concerne en outre l'utilisation de ces promoteurs ou éléments de contrôle de promoteur pour moduler les niveaux de transcrits dans des plantes et des plantes contenant ces promoteurs ou éléments de contrôle de promoteur.
PCT/US2011/044033 2010-07-16 2011-07-14 Promoteur, éléments de contrôle de promoteur et leurs combinaisons et utilisations WO2012009551A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
US36490310P 2010-07-16 2010-07-16
US61/364,903 2010-07-16

Publications (1)

Publication Number Publication Date
WO2012009551A1 true WO2012009551A1 (fr) 2012-01-19

Family

ID=45469804

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/US2011/044033 WO2012009551A1 (fr) 2010-07-16 2011-07-14 Promoteur, éléments de contrôle de promoteur et leurs combinaisons et utilisations

Country Status (1)

Country Link
WO (1) WO2012009551A1 (fr)

Cited By (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US9828608B2 (en) 2010-10-27 2017-11-28 Ceres, Inc. Transgenic plants having altered biomass composition
EP3434771A1 (fr) * 2012-12-19 2019-01-30 Monsanto Technology LLC Éléments régulateurs pour végétaux et leurs utilisations
US10323256B2 (en) 2011-12-09 2019-06-18 Ceres, Inc. Transgenic plants having altered biomass composition
WO2020097092A1 (fr) * 2018-11-06 2020-05-14 Pioneer Hi-Bred International, Inc. Éléments de modulation d'expression et procédés d'utilisation

Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20090144848A1 (en) * 2000-05-08 2009-06-04 Kovalic David K Nucleic acid molecules and other molecules associated with plants and uses thereof for plant improvement
WO2009099899A2 (fr) * 2008-02-01 2009-08-13 Ceres, Inc. Promoteur, éléments de commande de promoteur et combinaisons, et utilisation de ceux-ci
WO2009149304A2 (fr) * 2008-06-04 2009-12-10 Edenspace Systems Corporation Éléments régulateurs de gène de plante

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20090144848A1 (en) * 2000-05-08 2009-06-04 Kovalic David K Nucleic acid molecules and other molecules associated with plants and uses thereof for plant improvement
WO2009099899A2 (fr) * 2008-02-01 2009-08-13 Ceres, Inc. Promoteur, éléments de commande de promoteur et combinaisons, et utilisation de ceux-ci
WO2009149304A2 (fr) * 2008-06-04 2009-12-10 Edenspace Systems Corporation Éléments régulateurs de gène de plante

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
DATABASE GENBANK 28 February 2006 (2006-02-28), Database accession no. AC169370 *

Cited By (10)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US9828608B2 (en) 2010-10-27 2017-11-28 Ceres, Inc. Transgenic plants having altered biomass composition
US11667925B2 (en) 2010-10-27 2023-06-06 Ceres, Inc. Transgenic plants having altered biomass composition
US10323256B2 (en) 2011-12-09 2019-06-18 Ceres, Inc. Transgenic plants having altered biomass composition
US10815496B2 (en) 2011-12-09 2020-10-27 Ceres, Inc. Transgenic plants having altered biomass composition
US10822616B2 (en) 2011-12-09 2020-11-03 Ceres, Inc. Transgenic plants having altered biomass composition
US11299747B2 (en) 2011-12-09 2022-04-12 Ceres, Inc. Transgenic plants having altered biomass composition
EP3434771A1 (fr) * 2012-12-19 2019-01-30 Monsanto Technology LLC Éléments régulateurs pour végétaux et leurs utilisations
AU2018203164B2 (en) * 2012-12-19 2020-03-12 Monsanto Technology Llc Plant regulatory elements and uses thereof
WO2020097092A1 (fr) * 2018-11-06 2020-05-14 Pioneer Hi-Bred International, Inc. Éléments de modulation d'expression et procédés d'utilisation
US20210388369A1 (en) * 2018-11-06 2021-12-16 Pioneer Hi-Bred International, Inc. Expression modulating elements and methods of use

Similar Documents

Publication Publication Date Title
US11560569B2 (en) Promoter, promoter control elements, and combinations, and uses thereof
US9896693B2 (en) Promoter, promoter control elements, and combinations, and uses thereof
US7378571B2 (en) Promoter, promoter control elements, and combinations, and uses thereof
US20050223422A1 (en) Promoter, promoter control elements, and combinations, and uses thereof
US20050246785A1 (en) Promoter, promoter control elements, and combinations, and uses thereof
US20060021083A1 (en) Promoter, promoter control elements, and combinations, and uses thereof
US20090181851A1 (en) Promoter, Promoter Control Elements, And Combinations, And Uses Thereof
US10851383B2 (en) Promoter, promoter control elements, and combinations, and uses thereof
US8278434B2 (en) Promoter, promoter control elements, and combinations, and uses thereof
WO2012009551A1 (fr) Promoteur, éléments de contrôle de promoteur et leurs combinaisons et utilisations
US20100299784A1 (en) Promoter, promoter control elements, and combinations, and uses thereof
US11634723B2 (en) Promoter, promoter control elements, and combinations, and uses thereof
US20230287442A1 (en) Promoter, promoter control elements, and combinations, and uses thereof
US7217864B2 (en) Shade responsive promoter, promoter control elements, and combinations, and uses thereof
US20080134357A1 (en) Promoter, promoter control elements, and combinations, and uses thereof
CA2402857A1 (fr) Promoteurs de gene atrsp

Legal Events

Date Code Title Description
121 Ep: the epo has been informed by wipo that ep was designated in this application

Ref document number: 11807522

Country of ref document: EP

Kind code of ref document: A1

NENP Non-entry into the national phase

Ref country code: DE

122 Ep: pct application non-entry in european phase

Ref document number: 11807522

Country of ref document: EP

Kind code of ref document: A1