Chemistry: molecular biology and microbiology – Plant cell or cell line – per se ; composition thereof;...
Patent
1996-10-07
2000-02-08
Lankford, Jr., Leon B.
Chemistry: molecular biology and microbiology
Plant cell or cell line, per se ; composition thereof;...
435431, C12N 500
Patent
active
060227443
DESCRIPTION:
BRIEF SUMMARY
The invention relates to methods for the induction of desiccation tolerance in plant embryoids. It also relates to methods for the germination of embryoids which have been dessicated. Novel desiccated embryoids are also provided.
Regeneration of storable embryoids has been described for a large number of species. Redenbaugh et al. (1986) were the first to create artificial seeds by encapsulation of embryoids in alginate gel beads. However, the still hydrated artificial seeds were difficult to store, because they lacked quiescence, and the conversion rate (outgrowth into plantlets) was very low. The recent achievement of induction of tolerance to complete desiccation (Senaratna et al., 1989a) may give new opportunities to the artificial seed technology. Desiccated embryoids are better suited for use in this technology, because they are in a quiescent state comparable to that of dry seeds and they therefore have better storage properties. Gray (1990) even stated that dried grape embryoids germinated better than fresh ones.
Desiccation tolerance is defined by us as the ability of embryoids to regrow after storage at 25.degree. C. for two weeks under low moisture conditions (e.g. 10%, comparable to dry seeds).
Production of desiccation tolerant embryoids has been published for the following species: alfalfa (Anandarajah and McKersie, 1990 and 1991; Senaratna et al., 1989a,b; McKersie et al., 1989), geranium (Marsolais et al., 1991), soybean (Parrott et al., 1988), spruce (Roberts et al., 1990; Attree et al., 1991), grape (Gray, 1990) and carrot (Lecouteux et al., 1992; Iida et al., 1992). In most studies the plant hormone abscisic acid (ABA) was used in amounts equivalent to the amounts used for inducing quiescence to induce desiccation tolerance. By adding the appropriate amounts of ABA to the culture medium at the right stage of development, depending on species and genotype, embryoids could be dehydrated to moisture contents of less than 10% (on a dry weight basis) with retention of some viability. Through the addition of ABA, it was even possible to induce desiccation tolerance in callus cultures of carrot (Nitzsche, 1980) and Craterostigma plantagineum (Bartels et al., 1990). Koornneef et al. (1989) and Meurs et al. (1992) have unequivocally demonstrated the role of endogenous ABA during seed development by using recombinants of ABA deficient and ABA insensitive mutants of Arabidopsis thaliana.
Not only exogenously supplied ABA, but also several stress treatments can induce the capacity to survive dehydration (Anandarajah and McKersie, 1990 and 1991). However, stress treatments such as heat or osmotic shock through high sucrose concentrations, may raise the endogenous ABA levels (Skriver and Mundy, 1990) and might thus induce desiccation tolerance.
The capacity of the embryoids to survive dehydration is also dependent on the drying method. Only Senaratna et al. (1989) gave a well defined description of the drying method, including drying rates and final moisture content in alfalfa embryoids.
Hoekstra et al. (1989) showed that regrowth of initially viable, dry organisms is impaired because of imbibitional damage. In all the previous studies on induction of desiccation tolerance in embryoids the plant recovery rates were always less than 100%. Poor embryoid quality, caused by less then optimal protocols, or asynchronic embryoid development could be the reason for the low recoveries.
The problem therefore remains to provide desiccated storable plant embryoids which are essentially 100% capable of germination after prolonged storage.
The invention provides such embryoids.
Also a method for germination of such embryoids in a manner that essentially all the embryoids germinate is provided.
The invention provides this solution through a method for the induction of essentially complete desiccation tolerance in plant embryoids wherein the embryoids are treated with an amount of abscisic acid activity which is significantly above the amount used to induce quiescence.
The physiological function ascribed to abscisic
Hoekstra Folkert Anne
Legro Robert Jean
Tetteroo Franciscus Abraham A.
Incotec International B.V.
Lankford , Jr. Leon B.
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