Multicellular living organisms and unmodified parts thereof and – Method of introducing a polynucleotide molecule into or...
Reexamination Certificate
2000-03-29
2004-11-09
Nelson, Amy J. (Department: 1638)
Multicellular living organisms and unmodified parts thereof and
Method of introducing a polynucleotide molecule into or...
C800S298000, C800S295000, C800S320000, C800S317000, C435S468000, C435S320100, C536S023600
Reexamination Certificate
active
06815578
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
The RAD50 gene of
Saccharomyces cerevisiae
plays a crucial role in meiotic recombination as well as DNA repair during vegetative growth (Kupiec, M. and Simchen, G.,
Mol. Gen. Genet.
193: 525-531, 1984). The yeast RAD50 gene encodes a 153 kDa protein (Rad50) that contains an ATP-binding site (Walker-B box or P-loop) in the N-terminal region and exhibits ATP-dependent DNA binding in vitro (Raymond, W. E. and Kleckner, N.,
Nucleic Acid Res.
16: 3851-3856, 1993). The Rad50 protein also exhibits two, 250 amino acid segments of heptad-repeat sequence, which form alpha helical coiled coil structures (Alani et al.,
Genetics
122: 47-57, 1989). In yeast, RAD50 deletion mutants show a mitotic hyper-recombinational phenotype. The same mutant exhibits reduced meiotic double strand break formation and recombination (reviewed in Malkova et al.,
Genetics
143: 741-754, 1996 and Jeggo, P.,
Radiation Res.
150: S80-S91, 1998).
Interestingly, similar phenotypes were observed in the deletion mutants for two other yeast genes MRE11 and XRS2, suggesting an involvement of these genes in double-strand break repair and homologous recombination (Malkova et al.,
Genetics
143: 741-754, 1996; Jeggo, P.,
Radiation Res.
150: S80-S91, 1998). Subsequently, Jozhuka and Ogawa demonstrated the interaction of yeast Rad50 and Mre11 proteins (Johzuka, K. and Ogawa, H.,
Genetics
139: 1521-1532, 1995). Tsukamoto et al., showed the involvement of yeast RAD50, MER11 and XRS2 as well as HDF1 (yeast homologue of Ku70) in illegitimate or non-homologous end-joining (Tsukamoto, Y. et al.,
Mol. Gen. Genet.
255: 543-547, 1997).
Recently, mammalian homologues of yeast RAD50 have been cloned and characterized extensively (Kim, K, et al.,
J. Biol. Chem.,
271: 29255-29264, 1996; Dolganov et al.,
Mol. Cell. Biol.
16: 4832-4841, 1996; Carney J. P. et al.,
Cell
93: 477-486, 1998; Trujillo, K. M. et al.,
J. Biol. Chem.
273: 21447-21450, 1998). Similarly, the
Arabidopsis thaliana
chromosome II BACF22D22 region (Accession No. AC006223) has been found to contain an open reading frame which encodes a protein with homology to yeast RAD50 (GI:4263721).
Control of homologous recombination or non-homologous end joining by modulating Rad50 provides the means to modulate the efficiency with which heterologous nucleic acids are incorporated into the genomes of a target plant cell. Control of these processes has important implications in the creation of novel recombinantly engineered crops such as maize. The present invention provides this and other advantages.
SUMMARY OF THE INVENTION
The present invention describes the maize Rad50 protein, which clearly possesses features characteristic of other Rad50 proteins, and has a calculated molecular weight of ~152.5 kDa. The maize Rad50 protein is characterized by the presence of an ATP binding site in the N-terminal region, a second nucleotide binding site in the C-terminal region, putative nuclear localization signals, and heptad-repeats. The presence of extensive leucine zipper structures appears to be another striking feature of the Rad50 proteins. These are also found in the maize Rad50protein and are indicated in bold in Example 4. The present invention also describes a maize Rad50 polynucleotide sequence. The maize Rad50 orthologue of the present invention was used as a probe to map the maize RAD50 gene(s) to the short arm of chromosome 4.
Generally, it is the object of the present invention to provide nucleic acids and proteins relating to maize Rad50. 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 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 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 otherwise provided for, software, electrical, and electronics terms as used herein are as defined in The New IEEE Standard Dictionary of Electrical and Electronics Terms (5
th
edition, 1993). The terms defined below are more fully defined by reference to the specification as a whole.
By “amplified” is meant the construction of multiple copies of a nucleic acid sequence or multiple copies complementary to the nucleic acid sequence using at least one of the nucleic acid sequences as a template. Amplification systems include the polymerase chain reaction (PCR) system, ligase chain reaction (LCR) system, nucleic acid sequence based amplification (NASBA, Cangene, Mississauga, Ontario), Q-Beta Replicase systems, transcription-based amplification system (TAS), and strand displacement amplification (SDA). See, e.g.,
Diagnostic Molecular Microbiology: Principles and Applications
, D. H. Persing et al., Ed., American Society for Microbiology, Washington, D.C. (1993). The product of amplification is termed an amplicon.
The term “antibody” includes reference to antigen binding forms of antibodies (e.g., Fab, F(ab)
2
). The term “antibody” frequently refers to a polypeptide substantially encoded by an immunoglobulin gene or immunoglobulin genes, or fragments thereof which
Mahajan Pramod B.
Shi Jinrui
Ibrahim Medina A.
Nelson Amy J.
Pioneer Hi-Bred International , Inc.
Pioneer Hi-Bred International Inc.
LandOfFree
Polynucleotide encoding MRE11 binding polypeptide and uses... does not yet have a rating. At this time, there are no reviews or comments for this patent.
If you have personal experience with Polynucleotide encoding MRE11 binding polypeptide and uses..., we encourage you to share that experience with our LandOfFree.com community. Your opinion is very important and Polynucleotide encoding MRE11 binding polypeptide and uses... will most certainly appreciate the feedback.
Profile ID: LFUS-PAI-O-3356918