Oregon State University

Effects of somatic embryogenesis and genetic background on the phenotype of the shootless mutant dks8
--Hardeman, K and Rivin, CJ

In the W22 background, the mutation dks8 results in small seeds that contain no observable shoot component and yet produce a functional root meristem, and a normal appearing cotyledon. Although the dks8 mutant kernels are capable of germinating a primary root, the root is devoid of any root hairs. To determine if the dks8 mutation was due to a lack of function in the seed or the embryo itself we asked if somatic embryos also had a shootless phenotype. To undertake this experiment, we introduced the dks8 allele into the maize line H99 because it forms embryonic callus at high frequencies.

Interestingly, we found that the phenotype of the dks8 kernels in H99 was significantly altered from the previously defined dks8 phenotype. When the H99 dks8 kernels were germinated on a hormone-free growth medium, they germinated into seedlings having pale leaves and seminal roots which, like the primary root, lacked root hairs. The seedlings died after expanding 5-6 leaves. We are further characterizing the phenotype of the H99 dks8 seedlings and attempting to determine the basis for the change in phenotype in the H99 background.

To determine if the dks8 mutation was autonomous to the embryo, we derived embryogenic callus from wildtype and mutant embryos, induced them to form somatic embryos, and determined their phenotypes. The results, shown in Table 1, show that the phenotypes of the somatic embryos reflect the phenotype of the callus source: Somatic embryos derived from wildtype callus formed green shoots and roots with root hairs, while the dks8 somatic embryos were either shootless or made a pale sickly shoot and in either case the roots lacked root hairs. This result suggests that the defect in dks8 mutant development is embryo-autonomous.

Table 1. Summary of tissue culture experiment.
Mutant embryos:
Total embryos used: 144
Total forming callus: 94
Total forming organs: 42
root and shoot: 15
root only: 27
Note: all roots lacked root hairs and all shoots were pale and slow growing.

Wildtype embryos:
Total embryos used: 279
Total forming callus: 232
Total forming organs: 102
root and shoot: 100
root only: 2
Note: roots made root hairs and shoots were green and grew well. 

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