Multicellular living organisms and unmodified parts thereof and – Plant – seedling – plant seed – or plant part – per se – Higher plant – seedling – plant seed – or plant part
Reexamination Certificate
2000-07-12
2004-09-07
Fox, David T. (Department: 1638)
Multicellular living organisms and unmodified parts thereof and
Plant, seedling, plant seed, or plant part, per se
Higher plant, seedling, plant seed, or plant part
C800S260000, C800S266000, C800S278000, C800S286000, C800S287000, C800S290000, C800S292000, C800S293000, C800S294000, C800S298000, C536S023100, C536S023200, C536S023600, C435S410000, C435S411000, C435S419000, C435S423000, C435S430000, C435S469000, C435S320100
Reexamination Certificate
active
06787687
ABSTRACT:
BACKGROUND OF THE INVENTION
1. Field of the Invention
The present invention relates generally to the RIN and MC genes. More specifically, it relates to methods and compositions for the modification of plant phenotypes with the RIN and MC genes.
2. Description of the Related Art
The ripe phenotype is the summation of biochemical and physiological changes occurring at the terminal stage of fruit development rendering the organ edible and desirable to seed dispersing animals and valuable as an agricultural commodity. These changes, although variable among species, generally include modification of cell wall ultrastructure and texture, conversion of starch to sugars, increased susceptibility to post-harvest pathogens, alterations in pigment biosynthesis/accumulation, and heightened levels of flavor and aromatic volatiles (Rhodes, 1980; Hobson and Grierson, 1993). Several of theses ripening attributes translate to decreased shelf-life and high input harvest, shipping and storage practices, particularly via changes in firmness and the overall decrease in resistance to microbial infection of ripe fruit. Currently acceptable techniques for minimizing the consequences of undesirable ripening characteristics include premature harvest, controlled atmosphere storage, pesticide application, and chemically induced ripening to synchronize the timing of maturation. Unfortunately, added production, shipping and processing expenses, in addition to reduced fruit quality, are often the consequence of these practices, challenging both the competitiveness and long term sustainability of current levels of crop production.
Although most fruit display modifications in color, texture, flavor, and pathogen susceptibility during maturation, two major classifications of ripening fruit, climacteric and non-climacteric, have been utilized to distinguish fruit on the basis of respiration and ethylene biosynthesis rates. Climacteric fruit such as tomato, cucurbits, avocado, banana, peaches, plums, and apples, are distinguished from non-climacteric fruits such as strawberry, grape and citrus, by their increased respiration and ethylene biosynthesis rates during ripening (Grierson, 1986). Ethylene has been shown to be necessary for the coordination and completion of ripening in climacteric fruit via analysis of inhibitors of ethylene biosynthesis and perception (Yang, 1985; Tucker and Brady, 1987), in transgenic plants blocked in ethylene biosynthesis (Klee et al., 1991; Oeller et al., 1991; Picton et al., 1993a), and through examination of the Never-ripe (Nr) ethylene perception mutant of tomato (Lanahan et al., 1994).
Considerable attention has been directed toward elucidating the molecular basis of ripening in the model system of tomato during recent years (reviewed in Spiers and Brady, 1991; Gray et al., 1992 and 1994; Giovannoni, 1993; Theologis 1992 and Theologis et al., 1993). The critical role of ethylene in coordinating climacteric ripening at the molecular level was first observed via analysis of ethylene inducible ripening-related gene expression (Tucker and Laties, 1984; Lincoln et al., 1987; Maunders et al., 1987; DellaPenna et al., 1989; Starrett and Laties; 1993). Several ripening genes, including ACC synthase and ACC oxidase, have been shown via antisense gene repression to have profound influences on the onset and degree of ripening (Hamilton et al., 1990; Oeller et al., 1991). Although the sum effect of this research has been a wealth of information pertaining to the regulation of ethylene biosynthesis and its role in ripening, the molecular basis of developmental cues which initiate ripening-related ethylene biosynthesis, and additional aspects of ripening not directly influenced by ethylene, remain largely unknown (Theologis et al., 1993).
Single locus mutations which attenuate or arrest the normal ripening process, and do not ripen in response to exogenous ethylene, have been identified in tomato and are likely to represent lesions in regulatory components necessary for initiation of the ripening cascade, including ethylene biosynthesis (Tigchelaar et al., 1978; Grierson, 1987; Giovannoni, 1993; Hobson and Grierson, 1993; Gray et al., 1994). One such mutation, the Nr mutation, has been identified and represents a gene responsible for ethylene perception and/or signal transduction and is a tomato homologue of the Arabidopsis Ethylene response I (EtrI) gene (Yen et al., 1995; Wilkinson et al., 1995).
Tomato has served as a model for ripening of climacteric fruit. Ripening-related genes have been isolated via differential gene expression patterns (Slater et al., 1985, Lincoln et al., 1987, Pear et al., 1989, Picton et al., 1993b) and biochemical function (DellaPenna et al., 1986; Sheehy et al., 1987; Ray et al., 1988; Biggs and Handa, 1989; Harriman and Handa, 1991; Oeller et al., 1991; Yelle et al., 1991). Promoter analysis of ripening genes has been performed via examination of promoter/reporter construct activities in transient assay systems and transgenic plants. The result has been the identification of cis-acting promoter elements which are responsible for both ethylene and non-ethylene regulated aspects of ripening (Deikiman et al., 1992; Montgomery et al., 1993). Trans-acting factors which interact with these promoters also have been identified via gel-shift and footprint experiments, although none have been isolated or cloned (Deikman and Fischer, 1988; Cordes et al., 1989; Montgomery et al., 1993).
The in vivo functions of several ripening-related genes including polygalacturonase, pectinmethylesterase, ACC synthase, ACC oxidase, and phytoene synthase have been tested via antisense gene repression and/or mutant complementation in transgenic tomatoes. For example, the cell wall pectinase, polygalacturonase, was shown to be necessary for ripening-related pectin depolymerization and pathogen susceptibility, however , the inhibition of PG expression had minimal effects on fruit softening (Smith et al., 1988, Giovannoni et al. 1989, Kramer et al., 1990). Significant reduction in rates of ethylene evolution resulting in inhibition of most ripening characteristics was observed in both ACC synthase and ACC oxidase antisense mutants (Oeller et al., 1991, Hamilton et al., 1990). Non-ripening antisense fruit were subsequently restored to normal ripening phenotype with the application of exogenous ethylene.
Further analysis of transgenic tomatoes inhibited in ethylene biosynthesis demonstrates that climacteric ripening represents a combination of both ethylene mediated and developmental control (Theologis et al., 1993). Although antisense ACC synthase tomatoes which failed to produce ethylene did not ripen, gene expression analysis demonstrated that several ripening-related genes, including polygalacturonase and E8 are expressed in the absence of ethylene. This observation confirms the presence of a developmental (or non-ethylene regulated) component of ripening. In fact, an ethylene requirement was observed for translation but not transcription of polygalacturonase mRNA, suggesting interaction between ethylene and non-ethylene components of ripening for expression of at least a subset of ripening genes (Theologis et al., 1993).
While the above studies have provided some insight into the ripening process in plants, there is still a great need in the art for novel methods and compositions for the creation of plants having enhanced phenotypes. In particular, there is a need in the art for the isolation the RIN and NOR genes. The isolation of these genes would allow the creation of novel transgenic plants altered in their fruit characteristics and/or ethylene responsiveness, and having one or more added beneficial properties.
SUMMARY OF THE INVENTION
In one aspect, the invention provides an isolated nucleic acid sequence comprising a RIN gene. In one embodiment of the invention, the RIN gene may be further defined as isolatable from the nucleic acid sequence of SEQ ID NO:6, from SEQ ID NO:5 or alternatively from SEQ ID NO:8, or any combinations of the foregoing.
In yet another aspect, the inventio
Giovannoni James
Padmanabhan Veeraragavan
Ruezinsky Diane
Tanksley Steven
Vrebalov Julie
Fox David T.
Fulbright & Jaworski LLP
Kallis Russell
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