KR20010110451A - Alteration of flowering time in plants - Google Patents

Abstract

The present invention provides genetic identification and uses information about the gene family of the FLOWERING LOCUS C (FLC) gene, which is the part that regulates the start of flowering in a plant. This information makes it possible to create transgenic plants that can selectively change the flowering time. Because these genes act to delay the flowering time in plants, an increase in FLC protein activity will delay the onset of flowering, while inhibition of FLC activity will hasten the onset of flowering. Representative sample numbers of gene families were described. Members of the gene family are also described as acting on other plant species.

Description

[0001] ALTERATION OF FLOWERING TIME IN PLANTS [0002]

An important developmental change in the life cycle of a plant is the change of growing plantlets from vegetative growth to flowering. The onset of flowering is crucial to the successful propagation of wild plants, and most plant species have an evolved system to precisely control flowering time. These systems monitor both environmental cues and plant development stages to regulate flowering.

Two environmental stimuli that are typically monitored are photoperiod and temperature. In the photoperiod-response plant experiment, daylength is detected in the leaves and flowering signals are thought to be transferred from the leaves to the fissured tissue (Zeevaar, Light and the Flowering Process , Process, eds., D Vince-Prue, B. Thomas and KE Cockshull, 137-142, Academic Press, Orlando, 1984.). By a process known as vernalization, exposure to low temperatures promotes flowering. The vernalization process has an effect by directing it to the splitting tissue, perhaps by making it suitable for receiving the flowering signal. (Lang, Encyclopedia of Plant Physiology , ed., W. Ruhland, 15 (Part 1), 1371-1536, Springer-Verlag , Berlin, 1965). Other environmental stimuli that may affect flowering include light intensity and nutritional status.

Also, the developmental state of the plant may affect the flowering time. Most species pass through the juvenile phase where flowering is inhibited and eventually reach the adult phase, which is suitable for flowering (Poeting, Science , 250, 923-930, 1990). This "phase change" leads to the appropriate size of plant for productive flowering.

In the literature on flowering, flowering flowering is often referred to as autonomous, not including consideration of environmental variables. However, it seems that autonomous and environmental paths are not completely separate. For example, a day-neutral species in a tobacco field blooms after forming a certain number of nodes. Thus, it can be classified as being entirely flown through autonomous pathways. However, a grafting study has shown that one-day neutral species and photoperiod-responsive tobacco species respond to similarly variable flowering signals (Lang et al., Proc. Natl Acad Sci USA , 74, 2412-2416, 1997; McDaniel et al., Plant J., 9, 55-61, 1996). The potential biochemical aspects of these pathways are therefore presumed to be conserved.

Genetic analysis in many species has revealed genes that influence timing of flowering. The most extensive genetic analysis of flowering time genes has been done in Arabidopsis thaliana . In Arabidopsis, the flowering time gene was revealed by two approaches. One approach was to induce early-flowering varieties of mutations affecting flowering time. Such mutations can lead to late-flowering or earlier-flowering. The maturation-flowering mutation reveals genes whose wild-type role of the gene promotes flowering, and early-flowering mutations reveal flowering inhibitory genes. Studies in Arabidopsis have revealed several different loci affecting the flowering time, as well as other aspects of their mutation, particularly loci and development, which affect the flowering time (eg det2, cop1, ga1 and phyB) (Koornneef et al, Ann Rev. Plant Physiol, Plant Mol, Biol, 49, 345-370, 1998;...... Weigel, Ann Rev. Genetics, 29, 19-39, 1995)

Another approach to identifying genes that affect flowering time is to measure the genetic basis that causes a natural variation in flowering time. The most commonly used varieties of Arabidopsis in the laboratory are early-flowering (breed), but most varieties are expiration-flowering. The late-flowering varieties differ from the early-flowering varieties in that the late-flowering varieties contain dominant alleles at two loci, FRIGIDA (FRI) and FLOWERING LOCUS C (FLC) , which inhibit flowering (Sanda et al., Plant Physiol, 111, 641-645, 1996; . Lee et al, Plant Journal, 6, 903-909, 1994;.... Clarke et al, Mol Gen. Genet, 242, 81-89, 1994; Koornneef et al , Plant Journal , 6, 911-919, 1994).

Physiological analysis of naturally occurring mutations in flowering time mutants and flowering time shows that flowering in Arabidopsis is regulated by various pathways (Koornneef et al., Ann. Rev. Plant Physiol., Plant Mol. Biol. , 49, 345-370, 1998). Plants containing a single group of mutants ( fca, fpa, fve, fy, ld ) and maturation -flowering FLC and FRI alleles that cause maturation -flowering have been shown to induce flowering in the induced condition Delayed, and delayed even more when the day is short. The vernalization treatment of these late-flowering lines can suppress the late-flowering phenotype. The other groups of cohort -flowering mutants ( co, fd, fe, fha, ft, fwa, gi ) show little or no difference in flowering time when grown in a short day compared to a long day. Moreover, this group showed little response to the sexualization process. Dual mutants containing mutations in each group bloom later than parents of single-mutants, whereas double mutants in one group do not flow much later than parents of each single-mutant (Koornneef et al Genetics , 148, 885-92, 1998). Thus, it is believed that there is an equal flowering pathway mediating the flowering reaction in response to environmental and developmental signals. The photoperiod pathway promotes flowering in a long day. Pathways referred to in the literature as autonomous (because the photoperiodic response is not affected by mutations in this pathway) is the age, or more specifically, the stage in which the plant regulates developmental stages suitable for blooming ≪ / RTI > A recent study of the developmental role of this pathway has shown that mutations in the autonomic pathway may alter the trichome pattern, which indicates that the adult transiton is delayed at the immature stage of these mutant plants (Telfer et al., Development, 124, 645-654, 1997).

Blocking of autonomous pathways by mutant alleles fca, fpa, fve, fy, and ld or by the presence of expiratory flowering FLC and FRI dominant alleles may be bypassed by the vernalization process (Koornneef et al. Ann Rev. Plant Physiol. Plant Mol. Biol. , 49, 345-370, 1998). Thus, FLC and FRI can be considered as genes that produce what is needed for the process of sexualization. Other species, especially Brassicas, are considered to have a "circuity" such as Arabidipsis. This similarity has been thoroughly analyzed in Brassicas for its relationship with dominant flowering inhibitors and vernalization treatments. The main difference between oilseed, Brassica napus and B. rapa, primary and biennial cultivars, is given by genes regulating flowering time in response to vernalization (Osborn et al., Genetics Society of America , 146, 1123-1129,1997). A comparison of the quantitative trait loci (QTLs) in the traits of the B. napus and B.rapa 1-year-old X 2-year-old cultivars shows that maturation in B. napus and B.rapa, - Osborn et al., Genetics Society of America, 146, 1123-1129, 1997). QTLs that determine two flowering times in B.rapa were isolated from the recombinant inbred group, and the QTLs that were most effective at flowering time were vernalization-responsive flowering time in rapa2 (VFR2). Furthermore, VFR2 is considered to correspond to the FLC of Arabidopsis: using the hybridizatin probe, which allows comparison of Arabidopsis and Brassicas after gene transfer into the early-flowering variety of the first year, The locus of VFR2 was determined, and it was found that only the probe corresponding to FLC had no recombination event with VER2 (<0.44 cm) indicating that VER2 is a FLC homolog.

Flowering time is very important for agriculture and horticulture. In horticultural crops, products are often flowers. For other types of food crops, forage crops, or fiber crops such as rice, wheat, corn, barley and oats, dicots such as beans, canola and cotton, sunflowers, tomatoes, broccoli and other legumes, Or fruits, seeds or pods resulting from flowering. Understanding the basis of molecular timing of flowering time regulation will lead to strategies to optimize flower, fruit and seed production by genetic manipulation to alter flowering time. For example, promoting the onset of flowering in certain crops will allow the crop to mature even in the shortest growing season, or will enable the cultivation of a variety of crops where only one crop is currently available .

In addition, there are crops that are useful for parts that are not flowering in plants. In these crops, inhibiting or substantially delaying flowering will increase the yield of these useful fractions. Examples of preferred plants where the flowering is delayed or suppressed include feed crops such as alfalfa and clover, cabbage and related vegetables such as Brassicas, spinach, and lettuce. Delaying or inhibiting flowering in crops where underground parts such as sugar beets or potatoes are used will increase production. In addition, inhibition of flowering in sugar beets will allow more energy to concentrate on sugar production. Similarly, the production of wood and biomass crops will be increased by delaying flowering. Therefore, there is a need and an important advantage to understand the basis of the molecular stage of flowering control.

SUMMARY OF THE INVENTION

The present invention includes a gene family of the FLOWERING LOCUS C (FLC) gene, which is one of the important regulators of flowering inhibition. The present invention includes DNA sequences for these genes, as well as polypeptides and proteins expressed from these genes.

The present invention also relates to transgenic plants whose flowering characteristics have been altered from non-transgenic plants of the same species by the presence of transgenes that influence the degree or timing of FLC protein activity in transgenic plants.

The object of the present invention is to provide a tool to those who wish to make new plant varieties that can change the flowering time in plant species. By altering the level of the FLC gene in a plant, the flowering time can be adjusted early or late as desired by the plant grower.

This makes it possible to transform plants in a very useful way. Because flowering is an important physiological step in flowering plants, the ability to manipulate flowering time in plant species makes it possible to increase plant growth or floral production in plants, depending on which more demanding.

Other objects and advantages of the present invention will become apparent from the following specification and attached drawings.

The present invention relates to the regulation of flowering time of plants by genetic engineering. In particular, the present invention relates to the regulation of flowering time by manipulating the activity of a family of FLOWERING LOCUS C (FLC) genes.

Figure 1 is a phylogenetic diagram for relative relevance among the members of the MADS box class of plant genes.

This specification is directly related to the nucleic acid and protein sequences for the genes of the Planting Locus C (FLC) family of plant genes. As described below, plants are generally found to have more than one FLC gene in their genome. However, these FLC genes are similar and homologous. It turns out here that these genes are used to make transgenic plants with altered flowering characteristics. With knowledge of the FLC gene described here, it is possible to slow the flowering time of plants by promoting flowering time of transgenic plants or by increasing FLC activity by inhibiting the activity of FLC in plants. Thus, this provides the plant grower with a special tool for manipulating the flowering time characteristics of the grain to better match the grower's wishes.

Described below are nucleic acid and amino acid sequences for several FLC genes. The work that produced this record began with the sequencing and sequencing of the gene from Arabidopsis thaliana , named FLC1. Several of these have discovered several other FLC genes using this information and other genetic information discussed below. Because it has one of the smallest genomes of all plants, the most extensively studied Arabidosis in plant genetics laboratories has been found to have at least three FLC genes, designated here as FLC1, FLC2, and FLC3. Using information from three FLC genes found in Arabidosis, two FLC genes from Brassica have been identified so far. These genes have several characteristics shared by all other plant FLC genes.

FLC genes are part of the gene category referred to as MADS box genes. The MADS box is a very well-preserved motif shared by a group of evolutionarily related transcription factors. The MADS name is an acronym for the original gene that was first found to share a common MADS box domain. There are a large number of MADS box genes that have been discovered so far and work is underway to organize these genes into subgroups. MADS box genes in plants are known to affect many aspects of plant development including plant structural development. The two FLC genes identified here (FLC2 and FLC3 of Arabidopsis ) have been previously sequenced as part of the Arabidopsis genome sequencing effort, although their action has not been previously known.

Figure 1, using data first accumulated by Martin Yanofsky and Elena Alvarez-Buylla, UC San Diego, is a phylogenetic tree for the relative relevance of the MADS-box proteins so far discovered from Arabidopsis and maize. It should be noted that the three FLC genes revealed are more closely related than those associated with other MADS box genes.

Described in the following sequence listing are the cDNA sequences of FLC1, FLC2, and FLC3 from Arabidopsis thaliana as well as BrFLClA and BrFLC1B from Brassica rapa and the amino acid sequences derived therefrom. What is also shown below is some sequence comparison information. These data show that the FLC1 and FLC2 genes from Arabidosis are 60% identical over their entire length, and are more than 50% identical over the entire region outside the MADS box region. The latter comparison is more important because all of the MADS box genes have relatively high conservation within the MADS box region itself. For purposes of this analysis, the MADS box region is considered to be the first 60 amino acids at the amino terminus of the protein sequence. The degree of sequence similarity is believed to be maintained across species. Brassica genes BrFLC1A and BrFLC1B are actually more similar to FLC1 than FLC1 is similar to FLC2. The identity of FLC2 to the Brassica gene is slightly less than 50% outside the MADS box, and among mutants of the FLC gene family, identity at the amino acid level is generally believed to be greater than 40%. Thus, an amino acid identity of greater than 40% outside the MADS box region is a marker indicating that it is a member of the FLC gene family.

Again, pay attention to the phylogenetic chart of FIG. It is important to note that all three consensus FLC genes from Arabidopsis are more closely related to each other than to the other Arabidopsis MADS box genes. Because a full spectrum of MADS box genes used by plants is known (at least in Arabidopsis from a nearly complete genome sequencing effort), the gene (s) as shown in Fig. 1, using commonly available sequencing and matching software, The members of the MADS box family can plan for the relevance of any newly sequenced genes. Members of the FLC gene family are genes located closer to FLC1, FLC2 or FLC3 of Arabidopsis than other MADS box genes from Arabidopsis by phylogenetic analysis.

One way to confirm that a gene is a member of the FLC gene family is to test the effect of the gene on the plant flowering time. Described in the examples below are actually tests that identify FLC genes that act to delay flowering in transgenic plants. Using Arabidopsis as a model, the activity of the putative FLC gene from other plant species can be identified by testing possible FLC genes that exhibit similar effects in transgenic plants.

It should be emphasized that this tool makes it possible to change the flowering time of a plant. The normal function of FLC is to delay or inhibit flowering. However, the availability of the FLC gene sequence makes it possible to change the flowering time of a plant in either direction. Several techniques for suppressing-modulating or promoting-the expression of endogenous plant genes are now known. In order to suppress-regulate gene expression, an antisense version of the coding sequence of the gene may be inserted into the plant, or the degree of gene expression may be lowered by co-suppression And incompletely understood phenomena by the insertion of artificial gene constructs into plants often suppress the expression of both the inserted gene and its other gene homologs. An extra copy of the gene may be inserted into the plant to facilitate the expression of the plant gene, preferably by transformation of the plant genome by a germ line, By appropriately selecting the length and characteristics, the activity of the protein produced by a gene inherent in plant cells and tissues can be reduced.

It is important to understand that plant genetic engineering techniques have evolved to the point where any genetic construct can be practically introduced into virtually any useful plant species. However, these processes still involve some random process, notably the processes in which the insertion of exogenous DNA into the genome of a plant occurs at random places in the plant genome. As a result, in a group of plants resulting from plant transformation processes, the results achieved in the case of different gene insertion processes will vary, and in some cases will change dramatically. For example, in the case of simple gene insertion of other replicons of endogenous plant genes, many of the produced plants will have slightly higher resistance protein activity, others will have no measurable change or rather measurable activity . On the other hand, some will have a substantial increase in activity. However, for each specific gene insertion, the results tend to be constant across generations, so this does not mean that a stable outcome can not be obtained. Thus, plants with virtually high activity have alleles with high activity that can be inherited by their offspring by normal Mendelian heredity.

As is known in the art, in order to produce a transgenic plant, it is necessary to construct a gene construct capable of expressing an exogenous or endogenous inserted protein coding sequence in the plant. In addition, a method of inserting a gene construct into a plant is required.

Tools and techniques for making gene constructs that express proteins in plants are now well known. A gene construct intended to induce the synthesis of a polypeptide or protein in a plant cell should contain a sequence of DNA known as a protein coding sequence, which specifically enumerates the sequence of the polypeptide or protein introduced into the resulting plant . In order for a protein coding sequence expressed in a plant to produce a polypeptide or protein, It should be placed under the control of the promoter expressed in the plant and also by the plant transcription termination sequence known as the polyadenylation sequence. Plant expression promoters are promoters derived from plant pathogens, such as plant-derived or virus (e.g., Cauliflower mosaic virus) or bacteria (such as Agrobacterium promoters such as the nopaline synthase promoter) that work in plants. Plant promoters from pathogens tend to be constitutive promoters, meaning that they always express protein coat sequences in all of the plant tissues. Other plant promoters are known to have tissue specificity (e.g., fruit or flower) or developmental specificity (e.g., plant growth stage such as neonatal specificity or senescence specificity), while others are intended to be attractable , Heat shock, or promoted metal ions). Some of these forms of the promoter may be used in the practice of the invention depending on the intended effect on the transgenic plant being produced.

Not all genetic constructs are intended to be produced as polypeptides or proteins within the cells of a transgenic plant. If the purpose of the manipulation is to lower the activity of the target protein in the plant, the genetic construct may be intended to lower the endogenous level of protein activity without production of the protein in the plant. One known method to accomplish this is to use antisense technology in which a mRNA strand that is complementary to a portion of the mRNA produced during the expression of the target gene is allowed to synthesize within the plant cell Genetic constructs are created. Antisense RNA interferes with the translation of target mRNA, and a small amount of protein is produced in affected plant cells.

Some methods for introducing genes into plants to make transgenic plants are exemplified. The most widely used methods are Agrobacterium-mediated transformation or accelerated particle-mediated transformation. A variety of Agrobacterium-mediated transformation techniques utilize the plant pathogens of the genus Agrobacterium to insert DNA from the plasmids in bacteria into the genome of plant cells. Particle-mediated plant transformation techniques use DNA coated with small carrier particles that are often accelerated into plant cells from a device called a gene gun. Complete methods of these approaches require techniques that can extract fully mature, morphologically normal plants from transformed cells. Thus, these techniques often include a selection or screening protocol to identify which plant cells have been transformed and a regenerative procollol to elicit whole plants from single modified plant cells. As mentioned above, these techniques have been used for many plant species and many, perhaps all economically important plant species. Other technologies such as electroporation have also been used to make transgenic plants. However, in the case of the invention fundamentally described herein, special techniques of plant transformation are not important. Once a plant is genetically engineered and a transgenic plant is created, the first plant transformation method is not a problem. The transgene inserted into the genome of a plant can be completely inherited by the offspring of a genetically engineered plant by the general rules of traditional plant cultivation. The term transgene is used herein to apply to an inserted genetic construct that has been transferred into the cells of the target plant. Thus, as used herein, the term transgenic plant refers to a plant carrying the same as the transgene.

Described here is information about plant gene sets, FLC genes. Although these genes have existed in their plants in their native or modified forms from now on, the explanation here is believed to be the first separate form of these genes. By separate forms, these genes have been isolated from their host plants. Currently, information from these genes can be used to engineer genetic and their components in vitro to create genetic constructs for use in some applications. Other intended uses relate to the diagnosis and analysis of plants that have not been transduced or introduced to analyze and form patterns of FLC gene activity to help grow or make plants with the characteristics of the desired flowering time.

As explained in more detail below, for example, plants lacking FLC activity as a result of FLC mutations bloom much earlier than plants with active FLC alleles. Plants with wild type FLC activity had about six times more leaves than plants with reduced FLC activity. Moreover, plants genetically engineered to express FLC at levels above normal show substantially delayed flowering. Because of the simplicity of genetic manipulation of plants, the following examples will be applied in Arabidopsis, and the same techniques will be applied to other plant species as well. In fact, the high sequence identity between FLC genes suggests that the FLC gene family portion from one plant species will be applied in accordance with general rules in other species of plants.

Flowering time control FLC gene is an inhibitor of flowering initiation in plants. Since other genes that regulate flowering work by regulating the activity of this gene, the start of flowering FLC polynucleotide fragments play a central role in regulating flowering delay. However, this gene is not the only factor that determines the flowering time in plants. For example, a locus named FRIGIDA (FRI) is a co-suppressor of flowering that cooperates with FLC (Lee et al., Plant Journal, 6, 903-909, 1994). Conversely, the LUMINIDEPENDENS gene, which promotes flowering, promotes flowering by reducing FLC levels. In mutants lacking LUMINIDEPENDENS activity that delayed flowering, the FLC level was substantially increased. Thus, manipulating flowering regulators is a reliable method of regulating flowering time.

It has also been found that FRI acts to increase FLC levels. In fact, both FLC and FRI genes are two dominant alleles that interact to slow down flowering. Using conventional nomenclature, lower case letters (eg, fri, flc) refer to recessive or inactive alleles, and capital letters (eg, FRI, FLC) refer to active dominant alleles. Both dominant alleles are normally required to slow down the flowering. Thus, early-flowering plants include gene combinations: fri / fri and flc1 / flc1; fri / fri and FLC1 / flc1; fri / fir and FLC1 / FLC1; FRI / fri and flc1 / flc1; And FRI / FRI and flc1 / flc1. The gene combinations that slow the flowering time are FRI / fri and FLC1 / flc1; FRI / FRI and FLC1 / flc1; FRI / fri and FLC1 / FLC1; And FIR / FRI and FLC1 / FLC1. Therefore, in testing of non-transgenic lines for FLC1-type mutants, plants that contain an unknown FLC1 allele should be screened for FRI alleles, preferably, It would be convenient to cross the tester strain containing the homozygous FRI allele.

The life span of a plant can be divided into at least two stages, a vegetative growth stage and a reproductive stage. For most commercially important crops, the growth continues to increase the plant size and number of leaves in the plant growth stage. Flowering begins at the fertilization stage. At this point, most plant growth refers to the growth (or development) of flowers, fruits and seeds. Commercially important crops have been grown to have the desired characteristics, including uniformity at the time of harvest. This resulted in high uniformity in the number of leaves present in each plant in the plant group grown in the same plant. Changes in the onset of flowering, due to the uniformity of the current number of leaves, can often be measured by the number of leaves attached to the plant. For example, if the initiation of flowering in a plant is activated, the plant will have fewer leaves than the same plant grown under the same conditions that were not activated to begin early flowering. Moreover, plants that are activated to start early bloom can be said to have a shorter plant maturation stage than the same plants grown under the same conditions that were not activated to start early blooming. Likewise, if the initiation of flowering is inhibited so that the plant begins flowering later, the plant will have many leaves as compared to the same plant grown under the same conditions, where flowering is not inhibited to start late. Moreover, it can be said that the plants with inhibited flowering start will have prolonged vegetative growth steps compared to the same plants grown under the same conditions without inhibiting flowering start. Also, the change in the flowering start time can be measured as the action time.

Plants into which the replication of the FLC gene has been introduced may also contain a flowering-time regulating coating region of a wild-type (i.e. endogenous) functioning to inhibit the onset of flowering. Once introduced into the plant genome, the FLC gene can act to augment the activity of the endogenous flowering-time regulatory coding region in order to delay the onset of flowering. For example, a second clone of the flowering-time regulatory coding region may be introduced into a plant to increase the amount of flowering-time-regulated FLC protein in the plant. Expression of the FLC protein portion encoded by the flowering-time regulatory coding region portion can also induce the activation of flowering in the plant. The portion of the polypeptide that induces flowering in plants to activate flowering may be referred to as a dominant negative mutant and is described in detail herein.

The invention also provides a genetically modified plant characterized by having a phenotype for the modified time of flowering start. Thus, the modified plants of the present invention, which have been modified by inserting FLC genes expressing new or additional FLC proteins in plants or by inhibiting the activity of endogenous FLC genes in plants, are compared with the same plant in which the transgene is not inserted And has another flowering start time. The onset of flowering (average) in transgenic plants begins preferably at least about 3 days, more preferably at least about 7 days, and most preferably at least about 12 days, after initiating flowering in the same plant without the transgene . Alternatively, the onset (average) of flowering in the transgenic plants may be at least about 3 days, more preferably at least about 7 days, most preferably at least about 12 days prior to commencement of flowering in the same plant without the transgene do. Preferably, the same genetically modified plant and the same plant without the transgene are grown under the same conditions. Actual data from Arabidopsis plants, as described below, demonstrate that the flowering time in plants is possible for more dramatic changes from three weeks to three months or vice versa. The difference in the onset of flowering in plants compared to the same plant without the introduced gene is due to the number of leaves in the genetically modified plant and the number of leaves in the same plant without the transgene By measuring the difference between the two. Transgenic plants have a higher number of leaves, preferably at least about 10%, more preferably at least about 50%, and most preferably at least about 80%, compared to the same plant without the transgene. Alternatively, the transgenic plants have at least about 10%, more preferably at least about 50%, and most preferably at least about 80% fewer leaves, compared to the same plant without the transgene. Preferably, the same genetically modified plant and the same plant without the transgene are grown under the same conditions.

In one embodiment of the invention, a FLC gene homologue from an allele and other species in the FLC gene sequence as well as a nucleotide sequence encoding a FLOWERING LOCUS C (FLC) gene coated region from Arabidopsis thaliana or a portion thereof There is provided a nucleic acid molecule comprising a polynucleotide having a nucleotide sequence. Homology is not limited to the following methods, but nucleic acids may be used in nucleic acid hybridization techniques, computer searches of databases, computer or manual comparisons of amino acid and nucleotide sequences, and proteins using antibodies specific for FLC Means a cognate relationship that can be determined by searching. The two nucleotide sequences are " similar " if the proportions of corresponding residues can be aligned to match. The two nucleotide sequences preferably have an identity of greater than about 31%, more preferably at least about 50%, even more preferably at least about 70%, and most preferably at least about 80%.

Thus, the present invention includes genes and proteins that are members of the FLC family of plant genes and shares an important primary structure with the FLC1 sequence provided below. The two amino acid sequences (i. E., The amino acid sequence of the homologue and the sequence of FLC1) are designed such that the residues constituting the MADS domain, i.e. 1-60 amino acids, are aligned in that region and the total lengths of the two amino acid sequences are common The number of amino acids is maximized. Although the amino acids in each sequence should maintain their proper order, the spacing in either or both sequences should be such that the residues of the MADS domain are located in a register and aligned to maximize the number of shared amino acids Allowed. Amino acid identity percentages are higher than two numbers: (a) the number of amino acids that two sequences have in common within the sequence, divided by the number of amino acids in FLC1 and multiplied by 100; Or (b) the number of amino acids that two sequences have in common within the sequence, divided by the number of amino acids in the candidate polypeptide and multiplied by 100. The flowering-time-regulating polypeptide has an identity of preferably at least 40%, more preferably at least about 50%, and most preferably at least about 70%, over the entire length of the FLC1 protein.

Sequence homology or identity is less important at the nucleotide level. As is well understood in this field, degeneracy of the genetic code allows many DNA protein coding sequences to coat the same amino acid sequence. For the convenience of a variety of cloning or for reasons of codon usage or preference in target plants, the coating sequence of the original protein coding sequence may be used as part of a plant expressing cassettes without altering the amino acid sequence of the resulting protein It is possible to deform it and is also general.

The isolated nucleic acid molecule of the present invention can be obtained by several methods. For example, they can be separated through well-known processes in the art. These include, but are not limited to: 1) hybridization of a genomic or cDNA library (libarary) with a detectably labeled probe that expresses all or a portion of any of the FLC genes to find similar nucleic acid sequences hybridization); 2) antibodies that screen expression libraries to look for similar structural features; 3) synthesis by gene amplification technology (PCR); And 4) chemical synthesis of nucleic acid molecules. Sequences for specific coding regions of the gene can also be found in the computer database of GenBank, National Institutes of Health. As described below, the coding region can be ligated to a vector after being separated.

Standard stringency hybridization conditions for identifying isolated nucleic acid molecules using a detectable probe, or for identifying polynucleotide fragments whose complement is hybridized with FLC1, are described in Chruch et al. Proc. Natl. Acad. Sci. USA, 81, 1991 1984, in a solution containing 0.5 M phosphate buffer, pH 7.2, 7% sodium dodecyl sulfate (SDS), 10 mM EDTA, Time culturing, and washing in a solution containing 2 x SSC (1X SSC: 150 mM NaCl / 15 mM sodium citrate, pH 7.0) and 0.1% SDS at 45 캜 for 3 hours and 20 minutes. Preferably, a polynucleotide (e.g., a probe) will hybridize with the nucleotide sequence set forth in SEQ ID NO: 1 under standard strict hybridization conditions. In general, polynucleotides (e.g., probes) do not necessarily have to be complementary to all nucleotides of the polynucleotide fragment, as long as the hybridization occurs under the conditions mentioned above. Optionally, high stringency conditions can be used when the temperature of the hybridization and washing steps is increased to 65, 60, 55 or 50 &lt; 0 &gt; C. Moreover, the time required for hybridization can be varied from the 12 hours described above. Typically, low stringency conditions allow the identification of FLC genes in other species by permitting the crossing of related but not identical FLC genes. Hybridization and washing temperatures were carried out in the preferred order, preferably at a hybridization temperature of 68 占 폚, a washing temperature of 65 占 폚; Hybridization and Washing Temperature 60 ° C; Hybridization and Washing Temperature 55 ° C; The hybridization and washing temperature is 50 [deg.] C, most preferably the hybridization and washing temperature is 45 [deg.] C.

The plants included in the present invention are flowering plants, including both monocotyledonous plants and dicotyledonous plants, capable of accommodating variant techniques. Examples of monocots include, but are not limited to, vegetables such as asparagus, onion and garlic; Cereals such as corn, barley, wheat, rice, sorghum, pearl millet, rye and oats; And grasses such as feed plants and grasses. Examples of dicotyledons include, but are not limited to, tomatoes, beans, soybeans, peppers, lettuce, peas, alfalfa, clover, pearl (eg, cabbage, broccoli, cauliflower, ), Carrots, beets, branches, spinach, cucumbers, pumpkins, melons, cantaloupes, sunflowers; feed and feed crops; Fiber crops such as cotton; And various decorative plants such as flowers and shrubs.

The examples described below demonstrate that increased expression of FLC delays or suppresses flowering and loss of FLC action causes accelerated flowering while there are other various methods that can alter flowering time by altering FLC expression . For example, transgenes encoding the dominant-negative form of the flowering-time-regulating polypeptide could be introduced into the plant genome. The dominant-negative mutant is a protein that actively suppresses the action of normal endogenous proteins. Thus, the function of the gene can be blocked without inactivating the structural gene itself or its RNA. This method was successful in the case of transcription factors in the MADS domain family such as FLC (Gauthier-Rouviere et al., Exp Cell Res, 209, 208-215, 1993; Mizukami et al., Plant Cell, 845, 1996). In these embodiments, which made dominant-negative using MADS box genes, the most effective constituents were those lacking the C-terminal domain of the polypeptide. Similar to the case of most MADS box genes, FLC shares the same underlying structure, so similar C-terminal truncations may be effective in FLC. This cleavage contains 1-150 amino acids of the entire FLC1 protein, which corresponds to 1-450 nucleotides of the FLC1 gene.

The above techniques generally describe the present invention. The present invention will now be described more fully hereinafter with reference to the accompanying drawings, in which exemplary embodiments of the invention are shown.

Example 1

Loss of FLC action causes early flowering.

The initiation of maturation in Arabidopsis was initiated principally by two dominant genes FLC1 (Lee, et al., Plant Journal 6, 903-909 1994; Koornneef, et al. Plant Journal 6, 911-919 1994) and FRIGIDA (FRI) , Mol . Gen Genet 237, 171-176 1993; Clarke and Dean. Mol Gen Genet 242, 81-89 1994). We show that the late-flowering phenotype of FRI is suppressed in the Landsberg erecta (Ler) strain of Arabidopsis by the recessive allele of FLC1 (Lee et al., Plant Pathology 6, 903-909 1994; Koornneef, et al. Plant Journal 6, 911-919 1994). Similarly, maturation induced by mutation in the LUMINIDEPENDENS (LD) gene is also inhibited by the Ler allele of FLC1 in the background of Ler (Lee, et al. Plant Journal 6, 903-909 1994; Koornneer, et al Plant Journal 6, 911-919 1994). To determine whether the ability of the Ler alleles of FLCl to inhibit the maturation -flowering phenotype of the FRI and ld mutations is accounted for by the loss-of-function mutation of FLC1, mutated populations of Arabidopsis were made to screen for plants containing the flc1 mutation . In one mutation, 5,000 late-flowering ld mutant plants (ld-3) were mutated to ethylmethanesulfonate. One early-flowering plant isolated from the complementation test from the 50,000 M2 plants selected from this cluster proved to be an allele of the recessive allele of FLC1 found in the Ler background. The second screen was performed using the maturation-flowering line of the homozygous homologue of the maturation-flowering allele FRI in the Columbia (Col) background. 90,000 seeds were treated with 5-6 krad of high-speed neutron radiation. 300,000 M2 plants were screened for early flowering and four new alleles flc1 were isolated.

Damage was found in four of the introduced allele flc1 . The data for the early-flowering phenotype of all three fast-neutron mutated alleles flc1 are shown in Tables 1 and 2. All four mutant allele genes, flc1, were predicted to cause total loss of function. In summary, loss of FLC1 gene function results in total inhibition of the effect of expiration-flowering of FRI in both long-term and single-effect. In early-flowering lines that do not contain FRI in the background, the loss of FLC1 gene action affects the flowering time under only a single condition. Thus, wild-type or active FLC1 clearly serves as an inhibitor of flowering.

Effect of loss-of-function flc1 mutations in early-flowering lines, including FRI in the background Number of rosette leaves in flowering system Long - including FRI 74 (12) ^* long - flc1 -2 11.8 (.75) long - flc1 -3 12.0 (0.6) long - flc1 -4 11.8 (0.4) single - wild type with FRI > 100 single - flc1 -3.4 (2.7)

* The numbers in parentheses are the standard deviation. The number of leaves formed before flowering is the standard of flowering time. Thus, when containing FRI in the background, the flc1 mutant plants will bloom consistently at 1/6 of the flowering time of the wild-type plants, including FLC1 .

Effects of functional-loss flc1 mutations in the early-flowering system without FRI Number of rosette leaves in flowering system -Coll Wild Type 13.7 (0.8) ^* long- flc1 -3 13.2 (0.7) Single -Col Wild Type 55.4 (4.7) Single- flc1 -3 42.4 (3.5)

* The numbers in parentheses are the standard deviation. The number of leaves formed before flowering is the standard of flowering time.

The lines described here were obtained from the Arabidopsis Biological Resource Center, Columbus, Ohio. Unless otherwise noted, growth conditions are commonly known and used conditions for those skilled in the art. Because of the effect of the plowing treatment on the onset of flowering, growth conditions did not include prolonged exposure of plants to low temperatures (e.g., 0 ° C to 8 ° C). The techniques used to genetically analyze Arabidopsis thaliana are known to those of skill in the art (see, for example, Koornneef et al., Genetic Analysis, In: " Arabidopsis Protocols ", Methods in Molecular Biology Series, vol. , Martinez-Zapater et al., (Eds.), Human Press, Totowa, New Jersey, pp. 105-227,

Example 2

Separation of FLC by position cloning

F1 generated from the crossing of Ler (fri / fri: flc1 / flc1) and Col ( fri / fri: FLC1 / FLC1 ) to create a segregation cluster useful for high resolution mapping and FLC1 positional cloning Plants were crossed with test lines containing FRI in Ler (FRI / FRI; flc1 / flc1) . This test line contained the late-flowering allele of the FRI , but also included the flc1-Ler allele, leading to premature flowering. By the mating of F1 and FRI in the Ler early flowering and late flowering 1 hour by the descendants, and FRI interaction FLC1 separated by the presence of one FLC1 -Col; late (i. E., FRI / fri FLC1 / flc1) But flowered late in the presence of flc1-Ler (i.e., FRI / fri; flc1 / flc1 ). Test-crossing offspring (4500 plants) were screened with simple repetitive microsatellite markers nga158 and nga151 previously shown on flank FLC1 (Lee et al., 1994b) (Bell and Ecker, 1994 ). Plants containing recombination between nga158 and nga151 were then tested with a third simple repeat nucleotide sequence marker nga249, which revealed that FLC1 was present in the interval between nga249 and nga151.

The region between nga249 and nga151 was contained within four yeast artificial chromosome (YAC) clones. To produce additional markers, we measured the DNA sequence of the end clone of YAC and designed a primer to amplify the corresponding sequence from Ler and Col. These DNAs from Ler and Col were sequenced to confirm single-nucleotide changes and then used to generate cleaved amplified polymorphic sequence (dCAPS) markers (Michaels and Amasino, 1998; Neff et al., 1998). The marker derived from the right and left ends of the YAC CIC1B8 they were found in any of the recombination on one side of FLC1, which will be described that is present in the 620kb interval FLC1 is extended by a CIC1B8.

A group of 13 BAC, TAC and P1 clones has been identified by the Kazusa Arabidopsis genome project extending to CIC1B8. These clones have an insertion length of about 70-100 kb and were used as probes for DNA blotting with EcoRV-cut DNA from plants containing the fast-neutron induced flc1 mutation. Two overlapping clones, named K6M1 and NYB9, found several deleted bands in flc1-2 . Any 10-20 kb fragments of K6M1 and NYB9 resulting from partial cleavage with Sau3A1 were used to create a library in a binary vector (Hajdukiewicz et al., 1994), and each clone from this library It was used to modify the FRI in the Ler system. This library consists of DNA from the Col background, which contains the late-flowering allele of FLC1 . Thus, FRI in a Ler plant modified with a construct containing the Col allele of FLC1 will induce maturation . One of the clones from this library, 211-31, produced very expired-flowering T1 plants. More than a third of these plants did not bloom after 8 months of growth and progressed to old age.

Sequencing revealed three putative genes at 211-31. To identify which gene represents FLC1 , the three candidate genes are the two additional fast-neutron mutated flc1 alleles, namely, flc1-3 and flc1-4 , and one EMS-producing allele, flc1- 1 &lt; / RTI &gt; All of the alleles mutated by the fast-neutron showed polymorphism on bands due to the MADS box transcription factor. The determination of the DAN sequences of flc1-1, flc1-3 and flc1-4 revealed that they all contained damage to the first axon of the MADS box transcription factor. flc1-3 contains a 104 bp deletion portion that removes the start codon and flc1-4 contains a 7 bp deletion portion that causes frame shift after the first 20 amino acids. flc1-1 contains a single-base transition at the first exon / intron junction that changes the conserved GT donor site to AT and disrupts splicing. FLC1 cDNA was isolated by RT-PCR from the Col background, and the sequence is shown in the sequence listing.

Example 3

Generation of ARABIDOPSIS transgenic for late flowering

To determine the usefulness of FLC1 in altering flowering time, transgenic Arabidopsis was constructed containing two different FLC1 constructs. The first construct, 211-31, was made of genomic DNA containing FLC1 and a unique promoter. The second construct, pSM7, contained the genomic coding region of FLC1 under the control of the structural 35S promoter from cauliflower mosaic virus (Odell, et al., Nature 313, 810-2 1985).

211-31 was transformed into the FRI of Ler by Agrobacterium-mediated transformation (Bechtold, et al., CR Acad Sci. Paris, 316: 1194, 1993). The unmodified FRI of Ler blooms after the formation of about 14 initial rosette leaves (Lee et al., Plant Journal 6, 903-909 1994). The FRI strain to 211-31 in the Ler plants by mutual synergistic effect of FRI and FLC1 which to delay the flowering and showed it significant delay in flowering (Table 3). More than 90% of the transformants formed more than 50 leaves before flowering and 38% were not flowered at all even when grown under conditions rich in red light that strongly promoted flowering in Arabidopsis . Thus, overexpression of FLC1 can inhibit flowering throughout the life of the plant. In plants modified by 211-31, biological resources were increased ten-fold due to the continuation of the biological growth phase. This explains that over-expression of the FLC gene can turn early-flowering plants into late-flowering plants.

pSM7 has been transformed into a wild-type Ler by Agrobacterium mediated transformation to measure the effect of structural expression of FLC1 in the normal early-flowering line (Bechtold, et al., CR Acad Sci. Paris, 316: 1194 , 1993). The results are summarized in Table 5. 30% of the transgenic plants obtained did not bloom significantly later than Ler parents, 35% bloomed moderately late, and 35% bloomed very late. Changes in the flowering time of transgenic plants may be due to differences in the degree of expression by insertion of other sites in the genome. This indicates that FLCl expression is sufficient to substantially delay flowering, even in the absence of FRI activity. Moreover, delays in flowering time due to structural FLC1 expression are not sensitive to vernalization treatments (vernalization treatment is effective in promoting flowering in naturally late-flowering lines including FRI and FLC1 ). Thus, by replacing the original FLC promoter with a structural promoter, it became possible to create expired-flowering plants with a late-flowering characteristic that is not sensitive to environmental signals. In addition, delay in flowering time is related to substantial increase in living resources.

Flowering time of plants transformed with FLC1 ^* containing FRI Number of rosette leaves at flowering Rate of transformed plants 12-20 leaves 8% 20-40 leaves 0% 40-80 leaves 54%> 80 leaves 38%

Plants containing FRI , which lack FLC1 function and are not modified in Ler , bloom after forming 12-14 rosette leaves. On the other hand, more than one FLC1 has been introduced and 92% of the transformed plants have formed at least three times more leaves before flowering. The number of leaves formed before flowering is the standard of flowering time. Thus, the introduction of the FLC1 gene into this line requires that modified plants consistently require at least 6 times longer time for flowering. In some strains, no flowering occurred.

Increase in biomass of plant modified with FLC1 ^* containing FRI System weight The modified with FRI 1.5g (.14) 211-31 in Ler FRI 15.5g in Ler (2.1)

* Fresh weight of plants was increased 10-fold when transformed into FLC1 . The numbers in parentheses are the standard deviation.

Flowering time of Ler plants modified by structurally expressed FLC1 ^* Number of rosette leaves at flowering Rate of transformed plants 7-10 leaves 30% 10-20 leaves 35%> 20 leaves 35%

* Wild type Ler blooms after forming 7-8 leaves. 70% of the original variants exhibited delayed flowering, and 35% formed more than twice the leaf before flowering. The number of leaves formed before flowering is the standard of flowering time. Therefore, the flowering time in the system which expresses FLC1 structurally is delayed three times or more.

Example 4

Examination of FLC1 homologues from BRASSICA

FLC activity regulates flowering time in Arabidopsis species. To investigate this possibility, FLC1 homologues were isolated from Brassica rapa mRNA using RT-PCR using primers designed according to the Arabidopsis FLC1 sequence. The nucleotide sequences of B. rapa homologues ( BrFLClA and BrFLCIB ) are shown in the sequence listing below. Amino acid identity between FLC1 and BrFLC1A and BrFLC1B is shown in Table 6. Over-expression constructs were constructed by placing BrFLClA and BrFLCIB under the control of the structural 35S promoter. These constructs were introduced into the early-flowering line of Arabidopsis, and the flowering time of T1 plants was specified. Many Arabidopsis plants, including the Brassica overexpressing construct, were delayed in flowering and formed more than three times as much leaves as before flowering than were regulated not to be introduced. Thus, FLC1 analogs isolated from B. rapa, like overexpression of FLC1 from Arabidopsis, can delay flowering in Arabidopsis when expressed structurally.

Amino acid identity between FLC1 and Brassica rapa FLC1 homologues Identity within the MADS domain ^* Identity outside the MADS domain Full identity BrFLC1A 88% 81% 83% BrFLC1B 93% 81% 85%

* For this comparison, amino acids 1-60 are considered to be the MADA box domain. Because of the high conservation within the MADA box domain between family members, there are the same inner and outer sides of the MADA box domain.

Example 5

Identification of genes such as FLC1 in ARABIDOPSIS

Database studies using the FLC1 sequence identified two other MADA box domain genes, FLC2 and FLC3 , which are important homologs in FLC1 . These genes have not been identified yet, but have already been identified previously by efforts to sequence the Arabidopsis genome, named FLC2 and FLC3 . CDNA was prepared for FLC2 and FLC3 using RT-PCR. The nucleotide sequences of FLC2 and FLC3 are shown in the following sequence listing. The identity of the amino acids between FLC1 and FLC2 and FLC3 is shown in Table 7.

The identity of amino acids between FLC1 and FLC2 and FLC3 Identity within the MADS domain ^* Identity outside the MADS domain Full identity FLC2 83% 50% 60% FLC3 83% 50% 60%

* For this comparison, amino acids 1-60 are considered to be the MADA box domain. Because of the high conservation within the MADA box domain between family members, there are the same inner and outer sides of the MADA box domain.

Example 6

In ARABIDOPSIS, FLC2 acts to inhibit flowering.

In a population of mutants mutated by T-DNA insertion using established techniques (Krysan, Young, Sussman, T-DNA as an insertion mutagen in Arabidopsis . Plant Cell , v.11, p.2283-2290, 1999) Functional deletion alleles were identified in FLC2 by PCR-based screening. Two alleles, flc2-1 and flc2-2, were found. In the allele flc2-1 , T-DNA was inserted at position 4138 of the genome sequence, and in the case of allele flc2-2 , T-DNA was inserted at position 442. The effects of loss of function mutation on flowering time were investigated under long day and single conditions. The results are shown in Table 8. Like FLC1 , FLC2 serves to delay flowering in Arabidopsis. Under long-day and single conditions, flc2 mutants flow faster than the corresponding wild-type; Thus, the wild type role of FLC2 is to delay flowering.

Effects of flc2 functional loss mutation on flowering time Range of number of rosette leaves at flowering Long-day-long-day wild-type 8-9 - flc 2-1 6-7 long-day -flc 2-2 6-7 Single-wild type 25-30 days -flc 2-1 7-9 single--flc2-2 9-11

Example 7

Overexpression of FLC2 delays flowering in ARABIDOPSIS.

To determine the effect of overexpression of FLC2 on flowering time, the coding region of the genome of FLC2 was placed under the control of the structural 35S promoter and inserted into Arabidopsis thaliana using common techniques (Bechtold et al., CR Acad. Sci Paris , 316, 1194, 1993). These results are shown in Table 9. Like FLC1 , overexpression of FLC2 is sufficient to delay flowering. Many modified plants formed 2-3 times more leaves before flowering.

Effect of overexpression of FLC2 on flowering time ^* Number of rosette leaves at flowering Rate of transformed plants 12-16 Leaves 55% 17-33 Leaves 45%

* Wild type that is not deformed blooms with 7-8 leaves.

Taken as a whole, this experimental data has established that the FLC family of genes acts to delay flowering time in plants. This data demonstrates that there are one or more FLC genes in many plants, even if not most plants. FLC genes can also be used alone to alter flowering time in plants, while FLC genes interact with other genes that regulate other flowering time, such as FRI. FLC genes encode proteins with high degree of sequence identity in the same plant species as well as in other plant species. Thus, useful tools can be provided that can manipulate and help regulate plant flowering over a wide range of plant species.

Detailed descriptions of all patents, patent applications, disclosures, and nucleic acid and protein data, including GenBank accession numbers and EMBL accession numbers, cited herein, are incorporated herein by reference. It is also understood that restrictions on the state of the art, accidental sequence errors or deletions can occur without affecting the usefulness of the data presented. Various modifications and alterations of this invention will be apparent to those skilled in the art without departing from the scope and spirit of this invention. It should also be understood that the present invention should not be unduly limited to the exemplary embodiments herein.

Claims (20)

A transgenic plant comprising a transgene encoding a member of the FLC gene family in the genome, wherein the transgenic plant has a modified flowering time when compared to non-transgenic plants of the same species. The transgenic plant according to claim 1, characterized in that the transgenic plant is flowering earlier than the non-transgenic plants of the same species. The transgenic plant according to claim 1, characterized in that the transgenic plant blooms later than the non-transgenic plants of the same species. 2. The transgenic plant according to claim 1, wherein the member of the FLC gene family is selected from the group consisting of FLC1, FLC2, and FLC3 from Arabidopsis thaliana , and BrFLClA and BrFLCIB from Brassica rapa . The seed of the transgenic plant of claim 1. In seeds for transgenic plants, Wherein said seed comprises a promoter capable of expressing in plants in a genome and an introduced gene comprising a protein coding region for a plant FLC protein, Wherein the plant FLC protein has (i) a MADS box domain, (ii) the FLC1 or FLC2 protein from Arabidopsis, SEQ ID NO: 2 or SEQ ID NO: 4 is at least 40% identical outside the MADS box domain domain , Iii) a seed characterized by being effective in delaying the onset of flowering in a transgenic plant when expressed in a transgenic plant, as compared to a non-transgenic plant with the same genetic background. Plant grown from the seed of claim 6. The method according to claim 6, Wherein the FLC protein is at least 50% identical to the amino acid sequence of the FLC1 gene outside the MADS box domain. Comprises a transgene comprising a protein coding region for a member of the expressible promoter and FLC protein family in the plant in the genome, and a member of the FLC protein family is FLC1 from Arabidopsis thaliana than any other MADS box domain protein from Arabidopsis thaliana Or seeds for transgenic plants that are more phylogenetically related to the FLC2 protein. A transgenic plant grown from the seed of claim 9. In seeds for transgenic plants, A promoter expressed in a plant that is operably linked to a sequence encoding the seed to be complementary to a sufficient portion of the protein coding region for the plant FLC protein in the genome to lower the extent of endogenous FLC protein activity in the plant grown from said seed, Comprising an introduced gene, Wherein the plant FLC protein has (i) a MADS box domain, (ii) the FLC1 or FLC2 protein from Arabidopsis, SEQ ID NO: 2 or SEQ ID NO: 4 is at least 40% identical outside the MADS box domain domain , Iii) Seeds for transgenic plants, characterized in that they are effective in delaying the onset of flowering in transgenic plants when expressed in transgenic plants, as compared to non-transgenic plants with the same genetic background. Arabidopsis, an isolated nucleotide sequence comprising the coding sequence for the FLC1 gene from SEQ ID NO: 1. Arabidopsis, a separate DNA sequence comprising a DNA sequence encoding the FLC1 protein from SEQ ID NO: 2. A promoter expressed in a plant operably linked to a protein coding sequence for the protein of the FLC gene family, Wherein the plant FLC protein has at least 40% identity with the FLC1 (SEQ ID NO: 2) or FLC2 (SEQ ID NO: 4) protein from Arabidopsis, i) the MADS box domain, ii) A genetic construct that, when expressed in an introduced plant, is effective in delaying the onset of flowering in a transgenic plant as compared to a non-transgenic plant with the same genetic background. 14. A plant comprising the genetic construct of claim 14 in the genome. 15. The method of claim 14, Wherein said FLC protein is selected from the group consisting of FLC1, FLC2, and FLC3 from Arabidopsis thaliana , BrFLClA and BrFLCIB from Brassica rapa . 15. The method of claim 14, Wherein the FLC gene is at least 50% identical in amino acid sequence to the FLC1 protein from Arabidopsis, SEQ ID NO: 1. A promoter expressed in a plant that is operably linked to a sequence sufficiently complementary to a protein coding sequence for the FLC gene family protein to lower the FLC protein activity in the transgenic plant, Wherein the plant FLC protein has at least 40% identity with the amino acid sequence of i) the MADS box domain, ii) the Arabidopsis, the FLC1 protein from SEQ ID NO: 1, and iii) A genetic construct that is effective at delaying the onset of flowering in transgenic plants compared to non-transgenic plants with background. In a transgenic plant containing a transgene for a member of the FLC gene family, the start of flowering of the genetically modified plant may be initiated at the beginning of flowering in a non-transgenic plant of the same genetic background, Transgenic plants that occur at least about 7 days before or after. A method of producing a transgenic plant having altered flowering characteristics, the method comprising: contacting a plant cell with a transgene comprising a plant expressing promoter and a plant FLC gene; Identifying a plant cell harboring the inserted transgene; Regenerating the transgenic plant from the plant cell, Wherein the transgenic plant has at least about 10% fewer or more leaves than the non-transgenic plant of the same genetic background without the transgene, the number of leaves is such that the transgenic plant and the non- &Lt; / RTI &gt; is measured under growth conditions.