We have developed methods for quantitative extraction and analysis of zeins from maize (Zea mays L.) flour. Extraction involved solubilization of total endosperm proteins in an alkaline buffer containing SDS and 2-mercaptoethanol with subsequent precipitation of nonzein proteins by the addition of ethanol to 70%. Analysis of these proteins by SDS-PAGE with Coomassie blue staining and by Westem blotting and ELISA assay with zein antibodies revealed that this extraction method is more quantitative than the traditional Landry-Moureaux procedure, especially for the ,-and -y-zeins. This Characterization of the maize mutant o25 by Mertz et al. (10) engendered considerable excitement among maize breeders. Its improved nutritional value, due to significantly increased percentages of lysine and tryptophan, led to the prediction that its use would become widespread in a short time. Unfortunately, poor agronomic characteristics (reduced yield, soft endosperm, and decreased disease resistance) associated with the o2 mutant have prevented its extensive commercial utilization.

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Paenibacillus azotofixans is a nitrogen-fixing bacterium often found in soil and in the rhizospheres of different grasses. In this study, two Brazilian clay soils were planted with cross-hybrid maize (BR-201) and four stages of plant growth were analyzed to characterize the P. azotofixans populations present in the rhizoplanes, rhizospheres, and non-root-associated soils (herein called nonrhizospheres). A total of 106 strains were isolated and identified as P. azotofixans with an API 50CH kit, by classical biochemical tests, and via the use of specific primers based on the 16S rRNA gene in PCRs. To compare the isolated strains, phenotypic characteristics were determined and three different probes were used in hybridization experiments: two nif probes and one probe comprising a 0.58-kb fragment cloned from the P. azotofixans C3L4 genome. These results were used to construct a dendrogram, in which two main clusters could be observed. One cluster contained exclusively strains from Várzea soil, and the other contained the majority of strains from Cerrado soil. The 60 strains from Várzea soil and the 46 strains from Cerrado soil were further analyzed with REP and BOX primers, respectively. Based on the patterns obtained, it was possible to identify 21 different groups among strains from Várzea soil and 4 different groups among strains from Cerrado soil. These different patterns were tested by multivariate analysis of variance, and differences in the populations of P. azotofixans during the four stages of plant growth were demonstrated. Moreover, strains isolated from the rhizoplanes, rhizospheres, and nonrhizospheres of maize planted in Cerrado and Várzea soils were shown to be statistically different; the diversity of P. azotofixans strains was affected by the soil type.

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A simple and cost-effective first-tier screening strategy for VIP-derived activities in B. thuringiensis collections can be developed by combining PCR and feeding bioassays. Moreover, the employed primers showed to be useful as a tool for strains differentiation at DNA level, and for characterization and isolation of Vip-like genes in tropical B. thuringiensis germplasm.

The traditional process of obtaining maize hybrids involves the generation of inbred lines through successive generations of selWng and subsequent testcrosses in order to identify the best combining ability by allelic complementation. A fast alternative to obtain inbred lines is to induce the formation of haploids followed by chromosome doubling. However, even with the aid of haploid-inducing genetic sources, this strategy has not been widely used in maize breeding programs, partly due to diYculties inherent to haploid generation and identiWcation. In order to evaluate the possibility of using dihaploids to generate homozygous maize tropical lines, we used the androgenetic haploid inducer line W23 as a female parent in crosses with the tropical single-cross hybrid BRS1010.Within the progeny of these crosses, 462 seeds were phenotypically selected as putative haploids by the purple-colored endosperm and colorless embryo conditioned by the R1-nj gene. Among these, only four individuals were conWrmed as being haploids using SSR markers, chromosome counting and Xow cytometry, showing that the phenotypic marker was not eYcient in detecting haploids in the tropical maize genotype used. All four haploids as well as some diploid plants presented reduced size, corroborating the diYculties for haploid identiWcation by phenotypic evaluation. Genetic diversity analysis revealed by SSR markers divided the haploids in two groups represented by Xint and dent maize inbred lines, which could be helpful in identifying complementary dihaploid lines. The present article demonstrates that a combination of haploid production and SSR Wngerprinting is a feasible strategy for maize hybrid development in tropical germplasm.

Hematoxylin staining is an early indicator of Aluminum (AI) toxicity effects on the apices of young, developing roots grown in nutrient solution. In this work, the potential of this technique as a reliable and reproducible phenotypic index for AI tolerance in tropical maize genotypes was assessed, with its performance systematically compared to two other parameters widely used in breeding programs -relative seminal-root length (RSRL) and net seminal-root length (NSRL). Seeding roots from contrasting genotypes for AI sensitivity stained remarkably different after 24-and 48-h and 7-day exposures to 222 J1M AI in nutrient solution, with the Al-dye complex being detected in both the outer (epidermis) and inner (cortex) portions of the roots from the sensitive cultivar. Hematoxylin staining was compared to the RSRL and NSRL parameters using 20 families from the third generation of selfing (S3) following the cross between two contrasting inbred lines that had been previously classified by the RSRL index in an independent procedure. The coloration technique showed the highest capacity to discriminate among tolerant and sensitive genotypes and displayed significant correlation coefficients to the other two indexes. Evaluation of the results from diallel crosses involving nine inbred lines proved that hematoxylin staining was also particularly adequate for identifying expressive hybrid vigor, as demonstrated by the general (GCA) and specific (SCA) combining ability estimates obtained by using the three indexes simultaneously. Hence, hematoxylin staining of Communicated by G. Wenzel