Multicellular living organisms and unmodified parts thereof and – Method of introducing a polynucleotide molecule into or... – The polynucleotide alters plant part growth
Reexamination Certificate
1999-10-08
2002-04-23
Nelson, Amy J. (Department: 1638)
Multicellular living organisms and unmodified parts thereof and
Method of introducing a polynucleotide molecule into or...
The polynucleotide alters plant part growth
C435S320100, C435S468000, C536S023600, C800S260000, C800S286000, C800S298000
Reexamination Certificate
active
06376751
ABSTRACT:
FIELD OF THE INVENTION
The present invention is directed to plant genetic engineering. In particular, it relates to modulation of expression of genes controlling reproductive development in plants.
BACKGROUND OF THE INVENTION
Arabidopsis primary shoots undergo a series of developmental phase changes as they mature and age (Schultz, et al., Development 119:745-765 (1993)). Their development can be broadly categorized into three major phases based on node morphologies: first, the rosette or vegetative phase, with nodes closely compressed and bearing a petiolated leaf and an axillary bud; second, the early-inflorescence phase, with nodes separated by internode elongation and bearing a sessile leaf and a coflorescence; and third, the late-inflorescence phase, with nodes bearing solitary flowers. Thus, two major phase transitions are involved in Arabidopsis main shoot development: the transition from rosette to early inflorescence when the rosette begins to bolt and the transition from early to late inflorescence (or from inflorescence to flower) when the primary shoot switches from producing cauline leaves and coflorescences to flowers. Ultimately, the primary shoot meristem becomes senescent and ceases producing flowers from its flanks (Shannon, et al.,
Plant Cell
3:877-892 (1991)).
The transition from rosette to early inflorescence is considered to be the vegetative-to-reproductive transition. It is regulated by many flowering-time genes, that is, floral repression and floral promotion genes (or early- and late-flowering genes, respectively) (Koornneef et al.,
Mol. Gen. Genet
. 229:57-66 (1991); Zagofta, et al.,
Aust. J. Plant Physiol
. 19:411-418 (1992)). Loss-of-function mutations in floral repression genes, such as EARLY FLOWER 1 (ELF1), cause early flowering, whereas mutations in floral promotion genes, such as CONSTANS (CO), delay transition from the rosette-to-inflorescence stage. In addition, two EMBRYONIC FLOWER (EMF) genes, EMF1 and EMF2, are proposed to be involved in this process as floral repressors, suppressing the onset of reproductive development (Sung et al.,
Science
258:1645-1647 (1992); Martinez-Zapater et al. In
Arabidopsis
, E. M. Meyerowitz and C. R. Somerville, eds (Cold Spring Harbor, N.Y.: Cold Spring Harbor Laboratory Press), pp 403-433 (1994); Castle, et al.,
Flowering Newslet
. 19:12-19 (1995); Yang, et al.,
Dev. Biol
. 169:421-435 (1995)). Based on this floral repressor concept, vegetatively growing plants must decrease EMF1 and EMF2 activities to initiate reproductive growth. It has been proposed that the floral repression genes maintain, whereas floral promotion genes inhibit, EMF1 and EMF2 activities. A balance of these gene actions would cause a gradual decline in EMF activities and determine the time of vegetative-to-reproductive transition.
The transition from inflorescence to flower is regulated by flower meristem identity genes, such as LEAFY (LFY), APETALA1 (AP1), AP2, and CAULIFLOWER (CAL) (Irish, et al.,
Plant Cell
2:741-753 (1990); Mandel, et al.,
Nature
360:273-277 (1992); Bowman, et al.,
Development
119:721-743 (1993); Jofuku, et al.,
Plant Cell
6:1211-1225 (1994)). Mutants with defective LFY, AP1, AP2, or AP1 CAL genes are impaired in flower initiation; thus, inflorescence-like or flowerlike shoots, instead of flowers, initiate peripherally from the apical meristem during the late-inflorescence phase. In addition to these genes, the TERMINAL FLOWER1 (TFL1) gene is reported to negatively regulate meristem identity gene function in inflorescence development. Both the primary shoot and the lateral shoots in tfl1 mutants terminate in a flower, reflecting a precocious inflorescence-to-flower transition (Alvarez et al.,
Plant J
. 2:103-116 (1992)). Molecular data have shown that the LFY gene is ectopically expressed in the entire apical meristem of tfl1 primary and lateral shoots, which is consistent with the tfl1 phenotype (Bradley, et al.,
Science
275:80-83 (1997)). Thus, TFL1 functions to maintain inflorescence development. Mutants impaired in EMF1 or EMF2 produce a reduced inflorescence and a terminal flower, indicating a role for the EMF genes in delaying the inflorescence-to-flower transition.
The development of Arabidopsis floral organs also depends on normal EMF gene function. As in ap1 and ap2 mutants, weak emf mutants, such as emf1-1 and all of the emf2 mutants, lack petals (Yang, et al., Dev. Biol. 169:421-435 (1995)). The strong emf mutant, emf1-2, is impaired in the development of all floral organs: only carpelloid organs form. The effects of emf mutations on inflorescence and flower development suggest that EMF1 and EMF2 continue to function during reproductive development.
In light of the above, it is clear that EMF genes play an important role in reproductive development in plants. Control of the expression of the genes is therefore useful in controlling flowering and other functions in plants. These and other advantages are provided by the present application.
SUMMARY OF THE INVENTION
The present invention provides methods of modulating reproductive development (e.g., flowering and other traits) in plants. The methods involve providing a plant comprising a recombinant expression cassette containing an EMF1 nucleic acid linked to a plant promoter.
In some embodiments, expression of the EMF1 nucleic acids of the invention are used to enhance expression of an endogenous EMF1 gene or gene product activity. In these embodiments, the nucleic acids are used to inhibit or delay transition to a reproductive state and can be used to promote vegetative growth of the plant. Alternatively, transcription of the EMF1 nucleic acid inhibits expression of an endogenous EMF1 gene or the activity of the encoded protein. These embodiments are particularly useful in promoting the transition to a reproductive state and, for instance, promoting uniform flowering in plants.
In the expression cassettes, the plant promoter may be a constitutive promoter, for example, the CaMV 35S promoter. Alternatively, the promoter may be a tissue-specific or an inducible promoter. For instance, the promoter sequence from the EMF1 genes disclosed here can be used to direct expression in relevant plant tissues.
The invention also provides seed or fruit produced by the methods described above. The seed or fruit of the invention comprise a recombinant expression cassette containing an EMF1 nucleic acid.
Definitions
The phrase “nucleic acid sequence” refers to a single or double-stranded polymer of deoxyribonucleotide or ribonucleotide bases read from the 5′ to the 3′ end. It includes chromosomal DNA, self-replicating plasmids, infectious polymers of DNA or RNA and DNA or RNA that performs a primarily structural role.
A “promoter” is defined as an array of nucleic acid control sequences that direct transcription of an operably linked nucleic acid. As used herein, a “plant promoter” is a promoter that functions in plants. Promoters include necessary nucleic acid sequences near the start site of transcription, such as, in the case of a polymerase II type promoter, a TATA element. A promoter also optionally includes distal enhancer or repressor elements, which can be located as much as several thousand base pairs from the start site of transcription. A “constitutive” promoter is a promoter that is active under most environmental and developmental conditions. An “inducible” promoter is a promoter that is active under environmental or developmental regulation. The term “operably linked” refers to a functional linkage between a nucleic acid expression control sequence (such as a promoter, or array of transcription factor binding sites) and a second nucleic acid sequence, wherein the expression control sequence directs transcription of the nucleic acid corresponding to the second sequence.
The term “plant” includes whole plants, plant organs (e.g., leaves, stems, flowers, roots, etc.), seeds and plant cells and progeny of same. The class of plants which can be used in the method of the invention is generally as broad as the class of higher plant
Aubert Dominique
Chen Lingjing
Sung Z. Renee
Mehta Ashwin D.
Nelson Amy J.
The Regents of the University of California
Townsend and Townsend / and Crew LLP
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