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Chromosomal abnormalities increased with time on the relatively high concentrations (2 .0 mg/l or more) of 2,4-D . Prolonged exposures of cell cultures to high concentrations of 2,4-D significantly increased the percent polyploid mitoses and were also associated with increased abnormalities of the spindle apparatus. Cultures maintained in the presence of NAA had a large number of cells with structural alterations of the chromosomes . The best embryogenic callus production (those maintained on 0 .1 and 1 .0 mg/1 2 .4-D) occurred in cultures with least detectable genetic variation . Plants could not be regenerated from cultures that had been on the maintenance media for seven months or more . The loss of regeneration potential was uniform across the concentrations and auxins tested, and only correlated with time . This loss could not be associated directly with detectable genetic variation .

The effect of the auxin phenylacetic acid (PAA) on wheat anther and on barley anther/microspore culture was investigated. With PAA the induction response was not usually significantly different from controls but a significantly higher number of green plants were produced in wheat anther and barley microspore culture. For wheat anther culture 100 mg/L PAA was beneficial. For barley microspore culture the optimum levels were from 1 to 100 mg/L, depending on genotype. In barley anther culture there were no improvements using PAA. In wheat anther culture, 145 green plants/100 anthers were obtained with cultivar Veery'S', while the average response from twelve F1 hybrids in the breeding program was 332 green plants/100 anthers. At least 1000 green plants were obtained using isolated microspores from 100 anthers in barley cv. Igri. With cv. Bruce, regeneration occurred only when 100 mg/L PAA was used. The influence of PAA appears at the embryogenic phase of the culture system. The possible mechanisms by which PAA may improve regeneration are discussed.

This report describes rapid regeneration of green plants from microspores of the barley cultivar Igri. Use of 0.3 M mannitol during maceration and isolation was essential for response from mechanically isolated microspores of barley cv. Igri grown under our conditions. A shed microspore culture system proved to be simple and gave a fast response; plants were obtained as early as 25 days after the material was taken from the donor plant. A 28-day cold-pretreatment of spikes can also be replaced with a 3-4 day pretreatment of anthers in mannitol. Shed microspores from 100 anthers produced an average of 292 plants with 91% of them green. Approximately 80% of the regenerated plants were spontaneously doubled-haploids.

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