Field tests of corn co-expressing two new delta-endotoxins from Bacillus thuringiensis (Bt) have demonstrated protection from root damage by western corn rootworm (Diabrotica virgifera virgifera LeConte). The level of protection exceeds that provided by chemical insecticides. In the bacterium, these proteins form crystals during the sporulation phase of the growth cycle, are encoded by a single operon, and have molecular masses of 14 kDa and 44 kDa. Corn rootworm larvae fed on corn roots expressing the proteins showed histopathological symptoms in the midgut epithelium.

Chromosome doubling is critical for obtaining doubled-haploid plants from wheat (Triticum aestivum L.) anther culture. The most common doubling method applies colchicine to the plant. However, colchicine is phytotoxic and can induce a high frequency of plant death. In this experiment, anthers from two wheat genotypes ('Pavon 76' and 'Centurk') were placed on nine embryoid initiation media having three sugar sources (maltose, sucrose, and maltose + glucose) with three colchicine concentrations (0.0, 0.1, and 0.2 g • 1'). Wheat starch was used as a gelling agent. After three days, the anthers were washed and moved to fresh media without colchicine. Increasing the colchicine concentration decreased the number of embryoids produced from 77,4 embryoids/100 anthers to 29.9 embryoids/100 anthers, but did not significantly affect the frequency of plant regeneration (0.49 green plants/embryoid to 0.40 green plants/embryoid), and increased the frequency of doubled-haploid plants (19.0 doubledhaploid plants/100 green plants to 72.3 doubledhaploid plants/100 green plants). Considering the total number of doubled-haploid plants produced, low levels of colchicine added to the initiation media were very effective.

Sugars are critical components in bread wheat {Triticum aestivum L.) anther culture media for successful somatic embryo initiation and plant regeneration. In this experiment, anthers from three wheat genotypes were cultured on a modified Liang's 85D12 initiation medium with seven sugar combinations (I-sugars: galactose, mannose, maltose, fructose, sucrose, glucose, maltose -I-glucose) at 0.26 M, and 2,4-dichlorophenoxyacetic acid (2 mg/L), 1-naphthalene acetic acid (1 mg/L), and glutamine (254 mg/L). Wheat starch (5 % W/V), a potential source of sugars, was used as the medium gelling agent. No previous research has studied the effect of different sugars with wheat starch. A split-plot experimental design with 42 replications was used with genotypes as whole plots and sugar combinations as subplots. Galactose and mannose did not support embryoid initiation and were dropped from the analysis. Averaged over the three genotypes, maltose was the best sugar (105 embryoids/100 anthers), followed by glucose (47 embryoids/100 anthers) and maltose + glucose (37 embryoids/100 anthers). These three sugar combinations were superior to the standard medium sugar, sucrose (24 embryoids/100 anthers), and to fructose (12 embryoids/100 anthers). The embryoids were divided into two groups for plant regeneration. The first group was transferred to regeneration medium (Liang 85D12 salts, sugars at 0.06 M, and wheat starch at 7 % w/v as gelling agent) with the same sugar (R-sugar) used as in initiation. The second group was transferred to regeneration media with sucrose. I+R-maltose (0.35) Published as paper #10251, Journal Series, Nebraska Agric. Res. Div. and R-sucrose with I-maltose + glucose (0.48) produced the highest numbers of green plants per embryoid. In general, R-sucrose was a very effective sugar for plant regeneration. The highest frequency of albino plants per embryoid (0.15) occurred on Rglucose medium. I-(-R-maltose H-glucose and IH-Rmaltose induced the highest frequency of spontaneous doubling (27.5 %). Hence, the sugar used in initiation and regeneration media, even in the presence of wheat starch (a glucose polymer), can affect numerous aspects of tissue culture and must be considered.The successful application of bread wheat {Triticum aestivum L.) anther culture techniques to plant breeding depends upon having a high frequency of haploid green plant production, a high induction frequency of chromosome doubling, and the cytological stability of doubled haploid plants. The number of haploid green plants from anther culture depends on three different components: embryoid production from cultured anthers, plant regeneration from those embryoids, and frequency of green plants produced (HENRY and DE BUYSER 1981, SZAKACS et al. 1988). Genotype, plant environment, and culture condition, have been studied and used to improve the effectiveness of haploid plant production (OuYANG et al. 1983, TIAN and CHEN S. Copyngh. Clearance Cemer Code Statement: 01 79-954 1/94/1201-0053$02.50/0

The use of doubled haploid plants in a wheat breeding program requires an efficient haploid production system. While the techniques for producing doubled haploids from anther culture are well established, those for isolated microspores are complicated and inefficient. Four methods of isolating microspores from anthers (blending, stirring, macerating, and floating) were compared. Isolated microspores were washed and cultured in liquid medium. The effects of pre-isolation marmitol conditioning, cell density, culture dilution, and sucrose centrifugation on microspore viability were evaluated. Isolation by blending gave the highest initial microspore viability (75%). Mannitol conditioning and purification by sucrose centrifugation had a detrimental effect on initial viability. An initial microspore density of 2 x 105 microspores per ml was necessary for continued microspore viability. One hundred and nine haploid or spontaneously doubled haploid plants were regenerated from microspores isolated without mannitol conditioning using the blending method. Based on this research, blender isolation with an initial density of 2 x 105 microspores per ml is recommended for isolated microspore culture.

As part of a large-scale genomics project focused on understanding and improving the Shepody potato, we have increased the regeneration and transformation rates for this cultivar. Using combinations of auxins and trans-zeatin, leaf and stem explants were evaluated for callus induction and shoot formation. Several plant growth regulator combinations resulted in higher plant regeneration rates over a previous method. Using the best combination of auxin and cytokinin in combination with Agrobacterium-mediated transformation, we regenerated independent putative transformants from 59.5% of the total explants plated. We ran PCR on a sample of the plants to confirm transformation and 47.1% were nptII positive; giving a confirmed transformation rate of 28.0%.