Data processing: measuring – calibrating – or testing – Measurement system in a specific environment – Biological or biochemical
Reexamination Certificate
2000-06-27
2002-07-16
Marschel, Ardin H. (Department: 1631)
Data processing: measuring, calibrating, or testing
Measurement system in a specific environment
Biological or biochemical
C435S006120, C702S019000, C703S006000, C703S011000, C707S793000
Reexamination Certificate
active
06421613
ABSTRACT:
TECHNICAL FIELD
The present invention relates generally to plant molecular biology. More specifically, it relates to nucleic acids and methods for modulating their expression in plants.
BACKGROUND OF THE INVENTION
Genes encoding MCM proteins were originally reported from budding and fission yeast. Eukaryotic MCM sequences reported so far are homologous to any of the six MCM genes identified in yeast. Recently MCM proteins were reported in Archaea genomes as well, pointing to their ancient existence and critical role in several organisms.
MCM2-3-5 proteins are nuclear-localized in yeast and are required for DNA replication from specific replication origins. The MCM mutants exhibit defects in cell cycle progression in yeast. In Drosophila, mutations in MCM2 or MCM4 (dpa) inhibit cell proliferation and are lethal, with mutants showing prolonged S phase. In Arabidopsis following prolifera (PRL) gene disruption by transposon mutagenesis, semi-sterile plants with reduced megagametophyte viability were observed. Thus a clear role for the prolifera gene was established in plants in megagametophyte and embryo development.
At least in yeast, Xenopus, and mammals, there is a considerable amount of literature available about the MCM proteins and some models about their specific role at the molecular level in DNA replication. DNA replication is a very complex and tightly regulated process in eukaryotes and the machinery has been documented to be broadly conserved across eukaryotes. In eukaryotes, DNA replication actually is initiated from several replication origins and by some controlling mechanisms, that apparently involve other proteins, the replication process is restricted to a single round per each cell cycle. This ensures that cells do not end up with multiple sets of genomes resulting in polyploidy. Initiation of DNA replication follows the formation of a complex of proteins called Origin Recognition Complex (ORC) which may probably permit loading of other required proteins for the process such as cdc6 in yeast (cdc18 in
S. pombe
). Overexpression of cdc18 in
S. pombe
induces multiple rounds of S phase in the absence of mitosis and thus cdc6/cdc18 together ensure one round of DNA replication per cycle. Mutants of MCM proteins suggest that these proteins bind to chromatin in G1 phase and disassociate in S phase through the end of mitosis. This phase-specific association ensures that the replication origins remain competent only at the end of G1 phase and fire only once during S phase leading to one cycle of replication. In general, binding of MCM proteins to chromatin also require other proteins such as cdc6/cdc18 in conjunction possibly with some protein kinases cdk2/cdc2 and cdc7-dbf4 regulate firing of origins of replication.
Cell division plays a crucial role during all phases of plant development. The continuation of organogenesis and growth responses to a changing environment requires precise spatial, temporal and developmental regulation of cell division activity in meristems (and in cells with the capability to form new meristems, such as in lateral root formation). Such control of cell division is also important in organs themselves (i.e. separate from meristems per se), for example, in leaf expansion, secondary growth, and endoreduplication. It has been observed that endoreduplication occurs in about 20% of leaf epidermal cells in maize and in trichomes. In endosperm, endoreduplication has been well documented in maize. An important feature of cell differentiation in maize endosperm is nuclear enlargement through chromosome endoreduplication. This process may be related to many differentiation events, such as protein and starch synthesis and storage, accumulation of nucleotides, enzyme activation, and hormone synthesis, and consists of subsequent cycles of DNA replication without entering mitosis until high ploidy levels are attained. Variations in chromosome endoreduplication frequency in endosperm parenchyma have been described among maize populations.
What is needed in the art are methods and compositions to facilitate multiple rounds of DNA replication without nuclear division leading to polyploidy. Increasing the frequency of endoreduplication in the endospenn increases the size of endosperm cells and thus the endosperm size. Additionally, initiation of endoreduplication in leaf and stem cells, particularly in maize, leads to increased biomass and vegetative growth which has advantages in increasing plant yield. The present invention provides this and other advantages.
SUMMARY OF THE INVENTION
Generally, it is the object of the present invention to provide nucleic acids and proteins relating to the maize prolifera gene. It is an object of the present invention to provide: 1) antigenic fragments of the proteins of the present invention; 2) transgenic plants comprising the nucleic acids of the present invention; 3) methods for modulating, in a transgenic plant, the expression of the nucleic acids of the present invention.
Therefore, in one aspect, the present invention relates to an isolated nucleic acid comprising a member selected from the group consisting of (a) a polynucleotide having a specified sequence identity to a polynucleotide encoding a polypeptide of the present invention; (b) a polynucleotide which is complementary to the polynucleotide of (a); and, (c) a polynucleotide comprising a specified number of contiguous nucleotides from a polynucleotide of (a) or (b). The isolated nucleic acid can be DNA.
In another aspect, the present invention relates to recombinant expression cassettes, comprising a nucleic acid of the present invention operably linked to a promoter.
In another aspect, the present invention is directed to a host cell into which has been introduced the recombinant expression cassette.
In a further aspect, the present invention relates to an isolated protein comprising a polypeptide having a specified number of contiguous amino acids encoded by an isolated nucleic acid of the present invention.
In a further aspect, the present invention relates to a polynucleotide amplified from a
Zea mays
nucleic acid library using primers which selectively hybridize, under stringent hybridization conditions, to loci within polynucleotides of the present invention.
In another aspect, the present invention relates to an isolated nucleic acid comprising a polynucleotide of specified length which selectively hybridizes under stringent conditions to a polynucleotide of the present invention, or a complement thereof In some embodiments, the isolated nucleic acid is operably linked to a promoter.
In another aspect, the present invention relates to a recombinant expression cassette comprising a nucleic acid amplified from a library as referred to supra, wherein the nucleic acid is operably linked to a promoter. In some embodiments, the present invention relates to a host cell transfected with this recombinant expression cassette. In some embodiments, the present invention relates to a protein of the present invention that is produced from this host cell.
In yet another aspect, the present invention relates to a transgenic plant comprising a recombinant expression cassette comprising a plant promoter operably linked to any of the isolated nucleic acids of the present invention. The present invention also provides transgenic seed from the transgenic plant.
Definitions
Units, prefixes, and symbols may be denoted in their SI accepted form. Unless otherwise indicated, nucleic acids are written left to right in 5′ to 3′ orientation; amino acid sequences are written left to right in amino to carboxy orientation, respectively. Numeric ranges recited throughout the specification are inclusive of the numbers defining the range and include each integer within the defined range. Amino acids may be referred to herein by either their commonly known three letter symbols or by the one-letter symbols recommended by the IUPAC-IUB Biochemical Nomenclature Commission. Nucleotides, likewise, may be referred to by their commonly accepted single-letter codes. Unless otherw
Nadimpalli Ramgopal
Simmons Carl R.
Foley & Lardner
Hartter Amy
Jervis Herbert H.
Marschel Ardin H.
Pioneer Hi-Bred International , Inc.
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