Response of viviparous mutants to abscisic acid in embryo culture

Following its isolation from cotton and sycamore, abscisic acid (ABA) was shown to be present in a large number of plant species. Furthermore, ABA appears to be present in all tissues of the plant, the highest concentrations being localized in the seed and fruit pericarp. For this reason, and the fact that it inhibits germinating seedlings, ABA has been suggested to have a general function as a growth regulator which, by its interactions with other growth hormones, promotes seed dormancy. As such, it may also be involved in regulation of seed maturation processes determining seed size. A question arises concerning the failure of the "dormancy deficient" viviparous (vp) mutants of maize to respond to normal dormancy regulation. We have begun investigations to answer this question.

One line of evidence from gas chromatography indicates that no appreciable difference exists in ABA levels between wild-type and homozygous viviparous seeds (unpublished result, R. Newton).

Another line of investigation was begun using excised embryos and tissue culture techniques. Embryos were removed from surface-sterilized seeds and placed on an agar solidified medium (Murashige and Skoog, 1962) with various concentrations of ABA and no other growth regulators. Homozygous vp seeds were identified on an anthocyanin genetic background (purple aleurone) by their pleiotropic effect of pigment reduction (Robertson, 1955). Explanted embryos were then incubated at 34 C and 100 fc of illumination in a day:night regime of 12:12.

A preliminary run using only wild-type embryos incubated on log scale ABA concentrations indicated that a level of 1.0 mg% would significantly reduce germination frequency. Techniques were also perfected that allowed the removal of ABA-treated embryos to ABA-free environments (vermiculite tubes) and subsequent transfer to and maturation under greenhouse conditions. Using the 1.0 mg% ABA level, we tested homozygous and heterozygous vp embryos excised at various intervals post-pollination. Embryos of both genotypes and of the same age were placed on ABA-free medium as controls. Table 1 shows the results.

Table 1.

Around 77% of the vp/vp homozygous embryos plated on medium that inhibited wild-type growth grew at near normal rates. The vp/+ heterozygotes had about 10% germination on the same medium. Those normals that germinated never approached the growth rate of the homozygotes, indicating a real difference between the two genotypes.

These results coupled with the chromatographic analysis suggest that the viviparous phenotype is due not to a lack of ABA in the mutant seed but rather to a lack of response (dormancy) to the hormone. Current experimentation (concomitant with an expanded repetition of the above procedure) is being conducted with C-14 labeled ABA to detect the presence (or absence) of a protein receptor for ABA in these embryos.

Steve McDaniel, J. D. Smith and H. James Price


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