Multicellular living organisms and unmodified parts thereof and – Method of introducing a polynucleotide molecule into or... – The polynucleotide alters fat – fatty oil – ester-type wax – or...
Patent
1996-07-31
1999-09-21
Robinson, Douglas W.
Multicellular living organisms and unmodified parts thereof and
Method of introducing a polynucleotide molecule into or...
The polynucleotide alters fat, fatty oil, ester-type wax, or...
4351723, 4353201, 435419, 536 241, C07H 2104, C12N 514, C12N 1582
Patent
active
059556463
DESCRIPTION:
BRIEF SUMMARY
BACKGROUND OF THE INVENTION
The present invention relates to chimeric regulatory regions useful for controlling the expression of genes in plants. These chimeric regulatory regions can be derived from the opine synthase genes of the plant pathogen Agrobacterium tumefaciens.
Agrobacterium tumefaciens is a Gram-negative soil bacterium that infects most dicotyledonous and some monocotyledonous plants. An infection by Agrobacterium tumefaciens often results in the formation of crown gall tumors on the infected plant.
During the A. tumefaciens infection process, a defined DNA segment ("T-DNA") of the large tumor-inducing ("Ti") plasmid is transferred to a susceptible plant cell and integrated into the plant nuclear genome, whereby the T-DNA genes are expressed. Some T-DNA genes encode enzymes involved in the synthesis of hormones that are active in plants. These hormones can cause tumors in infected plants. Other T-DNA genes direct the synthesis and secretion of unique amino acid and sugar derivatives, termed opines. Agrobacterium tumefaciens can utilize these opines as a carbon and sometimes a nitrogen source. See Gelvin, Plant Physiol. 92: 281-85 (1990); Gelvin, TRANSGENIC PLANTS (Academic Press 1993); Ream, Ann. Rev. Phytopathol. 27: 583-618 (1989); Zambryski, Ann. Rev. Plant Physiol. Plant Mol. Biol. 43: 465-90 (1992).
T-DNA genes contain regions that are functional in plant environments and possess similarities to plant regulatory regions. For example, most plant promoters contain cis-acting elements such as upstream activating sequences ("UAS") (often called "enhancers") that, by binding trans-acting factors, define or influence the promoter strength and tissue-specific expression pattern. Atchison, Annu. Rev. Cell Biol. 4: 127-53 (1988). The overall strength of a given promoter, as well as its pattern of expression, can be influenced by the combination and spatial orientation of cis-acting elements and the presence of the nuclear factors that interact with these elements. Dynan, Cell 58: 1-4 (1989). Although initially resident on a prokaryotic plasmid, T-DNA genes possess all of the sequence elements (promoters and UAS) required for transcription in plants. For instance, T-DNA genes contain TATA boxes that set the site of transcription initiation, and often contain upstream elements, located more than 100 bp from the transcription initiation site, that modulate the levels of transcription. See Gelvin, TRANSGENIC PLANTS (Academic Press 1993).
Two T-DNA genes that possess upstream activating sequences are the octopine synthase (ocs) and mannopine synthase (mas) genes. The ocs gene encodes a product that condenses arginine and pyruvate to form octopine. Hack and Kemp, Plant Physiol. 65: 949-55 (1980). A 16-base pair palindrome located upstream of the ocs gene is capable of activating a heterologous maize adh1 promoter in a transient expression system. Ellis et al., EMBO J. 6: 11-16 (1987); Ellis et al., EMBO J. 6: 3203-08 (1987). This palindrome is also essential for ocs promoter activity in stably transformed tobacco calli. Leisner and Gelvin, Proc. Nat'l Acad. Sci. USA 85: 2553-57 (1988); Leisner and Gelvin, Plant Cell 1: 925-36 (1989).
The mas 1' and 2' genes share a dual bidirectional promoter and a 479 bp intergenic region. These genes encode enzymes for a two-step pathway for the synthesis of mannopine. Ellis et al., Mol. Gen. Genet. 195: 466-73 (1984); Komro et al., Plant Mol. Biol. 4: 253-63 (1985). The transcription of the mas genes is divergent, and the intergenic region contains all the cis-acting elements necessary for the transcription of both genes. DiRita and Gelvin, Mol. Gen. Genet. 207: 233-41 (1987); Fox et al. Plant Mol. Biol. 20: 219-33 (1992); Leung et al., Mol. Gen. Genet. 230: 463-74 (1991); Guevara-Garcia et al., Plant J. 4: 495-505 (1993).
The ocs and mas gene promoters have been used to direct the expression of linked genes in transgenic plants. However, the application of these promoters has been restricted by weak expression levels in certain tissues of transgenic plants. DiRita
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Cui Decai
Gelvin Stanton B.
Hauptmann Randal
Ni Min
Biotechnology Research And Development Corporation
Nelson Amy J.
Purdue Research Foundation
Robinson Douglas W.
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