Multicellular living organisms and unmodified parts thereof and – Method of introducing a polynucleotide molecule into or... – The polynucleotide confers pathogen or pest resistance
Reexamination Certificate
1998-01-09
2002-02-26
Bui, Phuong T. (Department: 1648)
Multicellular living organisms and unmodified parts thereof and
Method of introducing a polynucleotide molecule into or...
The polynucleotide confers pathogen or pest resistance
C800S278000, C435S006120, C435S069100, C435S069700, C435S410000, C435S418000, C435S419000, C435S320100, C530S370000, C536S023100, C536S023400, C536S023600, C536S024100, C536S024320, C536S024330
Reexamination Certificate
active
06350933
ABSTRACT:
FIELD OF THE INVENTION
The present invention relates generally to plant molecular biology. In particular, it relates to nucleic acids and methods for conferring pest resistance in plants, particularly lettuce.
BACKGROUND OF THE INVENTION
Recently, several resistance genes have been cloned by several groups from several plants. Many of these genes are sequence related. The derived amino acid sequences of the most common class, RPS2, RPM1 (bacterial resistances in
Arabidopsis
(Mindrinos et al.
Cell
78:1089-1099 (1994)); Bent et al.
Science
265:1856-1860 (1994); Grant et al.,
Science
269:843-846 (1995)), L6 (fungal resistance in flax; Lawrence, et al.,
The Plant Cell
7:1195-1206 (1995)), and N, (virus resistance in tobacco; Whitham, et al.,
Cell
78:1101-1115 (1994); and U.S. Pat. No. 5,571,706), all contain leucine-rich repeats (LRR) and nucleotide binding sites (NBS).
The NBS is a common motif in several mammalian gene families encoding signal transduction components (e.g., Ras) and is associated with ATP/GTP-binding sites.
The NBS is a common motif in several mammalian gene families encoding signal transduction components (e.g., Ras) and is associated with ATP/GTP-binding sites.
LRR domains can mediate protein-protein interactions and are found in a variety of proteins involved in signal transduction, cell adhesion and various other functions. LRRs are leucine rich regions often comprising 20-30 amino acid repeats where leucine and other aliphatic residues occur periodically. LRRs can function extracellularly or intracellularly.
Since the onset of civilization, plant diseases have had catastrophic effects on crops and the well-being of the human population. Plant diseases continue to effect enormous human and economic costs. An increasing human population and decreasing amounts of arable land make all approaches to preventing and treating plant pathogen destruction critical. The ability to control and enhance a plant's protective responses against pathogens would be of enormous benefit. Tissue-specific and temporal control of mechanisms responsible for plant cell death would also be of great practical and economic value. The present invention fulfills these and other needs.
What is needed in the art are plant disease resistance genes and means to create transgenic disease resistance plants, particularly in lettuce. Further, what is needed in the art is a means to DNA fingerprint cultivars and germplasm with respect to their disease resistance haplotypes for use in plant breeding programs. The present invention provides these and other advantages.
SUMMARY OF THE INVENTION
The present invention provides isolated nucleic acid constructs. These constructs comprise an RG (resistance gene) polynucleotide which encodes an RG polypeptide having at least 60% sequence identity to an RG polypeptide selected from the group consisting of: an RG1 polypeptide, an RG2 polypeptide, an RG3 polypeptide, and an RG4 polypeptide. RG1, RG2, RG3, RG4, and the like, represent individual “RG families.” Each “RG family,” as defined herein, is a group of polypeptide sequences that have at least 60% amino acid sequence identity. Individual members of an RG family, i.e., individual species of the genus, typically map to the same genomic locus. The invention provides for constructs comprising nucleotides encoding the RG families of the invention, which can include sequences encoding a leucine rich region (LRR), and/or a nucleotide binding site (NBS), or both.
The invention provides for an isolated nucleic acid construct comprising an RG polynucleotide which encodes an RG polypeptide having at least 60% sequence identity to an RG polypeptide from an RG family selected from the group consisting of: an RG1 polypeptide, an RG2 polypeptide, an RG3 polypeptide, an RG4 polypeptide, an RG5 polypeptide, and an RG7 polypeptide. In alternative embodiments, the nucleic acid construct comprises an RG polynucleotide which encodes an RG polypeptide comprising an leucine rich region (LRR), or, an RG polypeptide comprising a nucleotide binding site (NBS). The nucleic acid construct can comprise a polynucleotide which is a full length gene. In another embodiment, the nucleic acid construct encodes a fusion protein.
In one embodiment, the nucleic acid construct comprises a sequence encoding an RG1 polypeptide. The RG1 polypeptide can be encoded by a polynucleotide sequence selected from the group consisting of SEQ ID NO:1 (RG1A), SEQ ID NO:2 and SEQ ID NO:137 (RG1B), SEQ ID NO:3 (RG1C), SEQ ID NO:4 (RG1D), SEQ ID NO:5 (RG1E), SEQ ID NO:6 (RG1F), SEQ ID NO:7 (RG1G), SEQ ID NO:8 (RG1H), SEQ ID NO:9 (RG1I), and SEQ ID NO:10 (RG1J).
In another embodiment, the nucleic acid construct comprises a sequence encoding an RG2 polypeptide. The RG2 polypeptide can be encoded by a polynucleotide sequence selected from the group consisting of: SEQ ID NO:21 and SEQ ID NO:27 (RG2A); SEQ ID NO:23 and SEQ ID NO:28 (RG2B); SEQ ID NO:29 (RG2C); SEQ ID NO:30 (RG2D); SEQ ID NO:31 (RG2E); SEQ ID NO:32 (RG2F); SEQ ID NO:33 (RG2G); SEQ ID NO:34 (RG2H); SEQ ID NO:35 (RG21); SEQ ID NO:36 (RG2J); SEQ ID NO:37 (RG2K); SEQ ID NO:38 (RG2L); SEQ ID NO:39 (RG2M); SEQ ID NO:87 (RG2A); SEQ ID NO:89 (RG2B); SEQ ID NO:91 (RG2C); SEQ ID NO:93 (RG2D) and SEQ ID NO:94 (RG2D); SEQ ID NO:96 (RG2E); SEQ ID NO:98 (RG2F); SEQ ID NO:100 (RG2G); SEQ ID NO:102 (RG2H); SEQ ID NO:104 (RG21); SEQ ID NO:106 (RG2J) and SEQ ID NO:107 (RG2J); SEQ ID NO:109 and SEQ ID NO:110(RG2K); SEQ ID NO:112 (RG2L); SEQ ID NO:114 (RG2M); SEQ ID NO:116 (RG2N); SEQ ID NO:118 (RG2O); SEQ ID NO:120 (RG2P); SEQ ID NO:122 (RG2Q); SEQ ID NO:124 (RG2S); SEQ ID NO:126 (RG2T); SEQ ID NO:128 (RG2U); SEQ ID NO:130 (RG2V); and, SEQ ID NO:132 (RG2W).
In other embodiments, the nucleic acid construct comprises a RG3 sequence (SEQ ID NO:68) encoding an RG3 polypeptide (SEQ ID NO:138) (RG3). In other embodiments, the nucleic acid construct comprises an RG4 sequence (SEQ ID NO:69) encoding an RG4 polypeptide (SEQ ID NO:139) (RG4).
In other embodiments, the nucleic acid construct comprises a RG5 sequence (SEQ ID NO:134) encoding an RG5 polypeptide (SEQ ID NO:135). The RG5 polypeptide can be encoded by a polynucleotide sequence as set forth in SEQ ID NO:134.
The invention also provides for a nucleic acid construct which comprises an RG7 sequence encoding an RG7 polypeptide. The RG7 polypeptide can be encoded by a polynucleotide sequence as set forth in SEQ ID NO:136.
In further embodiments, the nucleic acid construct can further comprise a promoter operably linked to the RG polynucleotide. In alternative embodiments, the promoter can be a plant promoter; a disease resistance promoter; a lettuce promoter; a constitutive promoter; an inducible promoter; or, a tissue-specific promoter. The nucleic acid construct can comprise a promoter sequence from an RG gene linked to a heterologous polynucleotide.
The invention also provides for a transgenic plant comprising a recombinant expression cassette comprising a promoter operably linked to an RG polynucleotide. The expression cassette can comprise a plant promoter or a viral promoter; the plant promoter can be a heterologous promoter. In one embodiment, the transgenic plant is lettuce. In alternative embodiments, the transgenic plant comprises an expression cassette which includes an RG polynucleotide selected from the group consisting of SEQ ID NO:1 (RG1A); SEQ ID NO:2 and SEQ ID NO:137 (RG1B); SEQ ID NO: 3 (RG1C); SEQ ID NO:4 (RG1D); SEQ ID NO:5 (RG1E); SEQ ID NO:6 (RG1F); SEQ ID NO:7 (RG1G); SEQ ID NO:8 (RG1H); SEQ ID NO:9 (RG1I) and SEQ ID NO:10 (RG1J); SEQ ID NO:21 and SEQ ID NO:27 (RG2A); SEQ ID NO:23 and SEQ ID NO:28 (RG2B); SEQ ID NO:29 (RG2C); SEQ ID NO:30 (RG2D); SEQ ID NO:31 (RG2E); SEQ ID NO:32 (RG2F); SEQ ID NO:33 (RG2G); SEQ ID NO:34 (RG2H); SEQ ID NO:35 (RG2I); SEQ ID NO:36 (RG2J); SEQ ID NO:37 (RG2K); SEQ ID NO:38 (RG2L); SEQ ID NO:39 (RG2M); SEQ ID NO:87 (RG2A); SEQ ID NO:89 (RG2B); SEQ ID NO:91 (RG2C); SEQ ID NO:93 (RG2D) and SEQ ID NO:94 (RG2D); SEQ ID NO:96 (RG2E); SEQ ID NO:98 (RG2F); SEQ ID NO: 100 (RG2G); SEQ ID NO:102 (RG2H); SEQ ID
Meyers Blake C.
Michelmore Richard W.
Shen Kathy A.
Bui Phuong T.
The Regents of the University of California
Townsend & Townsend & Crew LLP
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