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PubMed2005

Natural variability of metabolites in maize grain: differences due to genetic background.

Understanding the impact of genetic diversity on crop biochemical composition is a prerequisite to the interpretation and potential relevance of biochemical differences experimentally observed between genotypes. This is particularly important in the context of comparative safety assessments for crops developed by new technologies such as genetic engineering. To interrogate the natural variability of biochemical composition, grain from seven maize hybrids grown at four geographically distinct sites in Europe was analyzed for levels of proximates (fat, protein, moisture, ash, and carbohydrates), fiber, amino acids, fatty acids, four vitamins, nine minerals, and secondary metabolites. Statistical evaluation of the compositional data at the p < 0.05 level compared each hybrid against every other hybrid (head-to-head) for all analytes at each site and then across all sites to understand the factors contributing to variability. Of the 4935 statistical comparisons made in this study, 40% (1986) were found to be significant. The magnitude of differences observed, as a percent, ranged between 0.84 and 149% when all individual sites and the combined sites were considered. The large number of statistically significant differences in the levels of these analytes between seven commercial hybrids emphasizes the importance of genetic background and environment as determinants of the biochemical composition of maize grain, reflects the inherent natural variability in those analytes across a representative sampling of maize hybrids, and provides a baseline of the natural range of these nutritional and antinutritional components in maize for comparative compositional assessments.

Algorithms↗

PubMed2005

Event specific qualitative and quantitative polymerase chain reaction detection of genetically modified MON863 maize based on the 5'-transgene integration sequence.

Because of the genetically modified organisms (GMOs) labeling policies issued in many countries and areas, polymerase chain reaction (PCR) methods were developed for the execution of GMO labeling policies, such as screening, gene specific, construct specific, and event specific PCR detection methods, which have become a mainstay of GMOs detection. The event specific PCR detection method is the primary trend in GMOs detection because of its high specificity based on the flanking sequence of the exogenous integrant. This genetically modified maize, MON863, contains a Cry3Bb1 coding sequence that produces a protein with enhanced insecticidal activity against the coleopteran pest, corn rootworm. In this study, the 5'-integration junction sequence between the host plant DNA and the integrated gene construct of the genetically modified maize MON863 was revealed by means of thermal asymmetric interlaced-PCR, and the specific PCR primers and TaqMan probe were designed based upon the revealed 5'-integration junction sequence; the conventional qualitative PCR and quantitative TaqMan real-time PCR detection methods employing these primers and probes were successfully developed. In conventional qualitative PCR assay, the limit of detection (LOD) was 0.1% for MON863 in 100 ng of maize genomic DNA for one reaction. In the quantitative TaqMan real-time PCR assay, the LOD and the limit of quantification were eight and 80 haploid genome copies, respectively. In addition, three mixed maize samples with known MON863 contents were detected using the established real-time PCR systems, and the ideal results indicated that the established event specific real-time PCR detection systems were reliable, sensitive, and accurate.

Animals↗

PubMed2006

Monolithic supports for the characterization of commercial maize products based on their chromatographic profile. Application of experimental design and classification techniques.

Modern analytical techniques based on the use of RP-HPLC with monolithic stationary phases and the application of experimental design and classification tools have been applied to the analysis of maize proteins. Solubilization conditions of maize proteins and separation conditions (temperature, detection wavelength, type and concentration of ion-pairing agent, and gradient) were optimized. The elution gradient was optimized by the application of experimental design techniques. The optimized method consisted of a linear binary gradient of water/acetonitrile/0.1% trifluoroacetic acid in three steps at a flow rate of 3 mL/min with a column temperature of 35 degrees C and UV detection at 280 nm. The developed method enabled the separation of maize proteins in an analysis time close to 8 min. Moreover, this is the first time that commercial maize products have been characterized by the use of multivariate classification techniques.

Chromatography, High Pressure Liquid↗

PubMed2006

Uptake of atrazine and cadmium from soil by maize (Zea mays L.) in association with the arbuscular mycorrhizal fungus Glomus etunicatum.

A greenhouse pot experiment was carried out to investigate the effect of the arbuscular mycorrhizal fungus Glomus etunicatum on the uptake of atrazine (ATR) and cadmium (Cd) from soil by maize (Zea mays L.). Mycorrhizal colonization led to an increase in the accumulation of Cd and ATR in maize roots but a decrease in the shoots. Atrazine alleviated the adverse effects of Cd on maize growth, and this was more pronounced in the inoculated plants. An increase in Cd accumulation by maize roots was observed when ATR was also present. After harvest, the residual ATR concentration in the soil decreased markedly. With mycorrhizal inoculation the amount of residual ATR decreased more in the bulk soil but less in the rhizosphere soil compared to the noninoculated controls. Cadmium application significantly decreased the ATR residual concentrations in both the rhizosphere and bulk soils irrespective of inoculation treatment.

Atrazine↗

PubMed1999

Inhibition of ACCase220 and ACCase240 isozymes from sethoxydim-resistant and -susceptible maize hybrids.

Acetyl-coenzyme A carboxylase (ACCase) isozymes were separated from cyclohexanedione-resistant and -susceptible maize. ACCase240 from resistant maize was 3.7-, >77-, and 12.8-fold more resistant to inhibition by clethodim, sethoxydim, and tralkoxydim, respectively, than ACCase240 from susceptible maize. The resistant ACCase240 preparation had 3.0-fold more protein and 14.5-fold lower specific activity than susceptible ACCase240. Resistant ACCase240 has a V(max) 5.5-fold lower than that of susceptible ACCase240, whereas apparent K(m) values were similar. ACCase220 from resistant maize was >25- and 7.2-fold more resistant to inhibition by sethoxydim and tralkoxydim, respectively, than susceptible ACCase220 but was inhibited to the same extent by clethodim. In summary, sethoxydim-resistant corn has an altered herbicide-resistant ACCase220 isozyme and increased expression of a less efficient, herbicide-resistant ACCase240 isozyme. However, to what extent alteration of both isozymes contributes to sethoxydim resistance is not clear.

Acetyl-CoA Carboxylase↗

PubMed1999

Dioscorea alata (water yam) as a replacement for maize in diets for laying hens.

One hundred and twenty-six point-of-lay birds were randomly alloted into six groups of 21 birds each and fed six diets containing 0%, 10%, 20%, 30%, 40% or 50% of Dioscorea alata meal as a replacement for maize, corresponding to 0%, 20%, 40%, 60%, 80% and 100% maize replacement. There were no significant differences (p > 0.05) in feed intake, egg weight or feed efficiency between the control diet and the test diets. However, the daily egg production per 100 birds (hen day production), differed significantly (p < 0.05) among the treatments. The birds on the control diet and diets with substitution rates of up to 80% laid significantly (p < 0.05) more eggs than those on the 100% substitution rate. This shows that D. alata can replace up to 80% of maize or constitute 40% of a laying chicken diet, provided the rations are isocaloric and isonitrogenous. For this to be achieved, more soybean and/or more fish scraps, and more palm oil are needed in the diets containing D. alata than in diets based on maize.

Animal Feed↗

PubMed1997

Molecular characterization of L2 lipoxygenase from maize embryos.

We investigated the expression and accumulation pattern of lipoxygenase isoforms throughout the maize plant life. Two forms of lipoxygenase L1 and L2 have been identified as acidic proteins of 100 kDa (pI 6.4) and 90 kDa (pI 5.5-5.7) which accumulate in dry embryos and in various organs of maize seedlings. In young embryos, only the L2 form was detected and accumulation of L2 mRNA decreased during embryo development. Identification of lipoxygenases from in vivo and in vitro synthesized proteins indicates that similar levels of both L1 and L2 forms accumulated during treatment with abscisic acid, (ABA) gibberellic acid (GA3) and jasmonic acid (JA). However, differences in the activity of both enzymes were detected. By using an antiserum directed against purified L2 we isolated and characterized a partial cDNA clone of maize embryos encoding a lipoxygenase. The deduced amino acid sequence of L2 cDNA shares 78% identity with the rice L2 protein, and 51-56% identity with lipoxygenases from the dicotyledonous plants soybean and Arabidopsis. DNA blot analysis indicated that maize contains a family of lipoxygenase genes which are presently being characterized.

Abscisic Acid↗

PubMed1998

Maize contains a Lon protease gene that can partially complement a yeast pim1-deletion mutant.

We have identified a gene in maize that encodes a product belonging to the Lon protease family. In yeast and mammals, Lon-type proteases catalyze the ATP-dependent degradation of mitochondrial matrix proteins. The maize gene, which we have designated LON1, is predicted to encode a protein with a molecular mass of 97.7 kDa. Lon1p is more similar in sequence to bacterial Lon proteases than to the yeast and human mitochondrial Lon proteases. LON1 transcripts are present in shoots of 4-day-old etiolated maize seedlings, and transcript levels decrease when these seedlings are heat-shocked. LON1 transcripts are also present at comparable levels in leaves and roots of 2-week-old greenhouse-grown seedlings. In yeast, the mitochondrial Lon-type protease, Pim1p, has been implicated in mitochondrial protein turnover, the assembly of mitochondrial enzyme complexes, and mitochondrial DNA maintenance, and it is essential for respiratory function. We show that maize Lon1p can replace the Pim1p function in yeast for maintaining mitochondrial DNA integrity, but not in the assembly of cytochrome a x a3 complexes.

ATP-Dependent Proteases↗

PubMed1998

Processing of a chimeric protein in chloroplasts is different in transgenic maize and tobacco plants.

Transgenic maize (Zea mays L.) and tobacco (Nicotiana tabacum Petit Havana SR1) plants have been generated, which overproduce a mitochondrial Nicotiana plumbaginifolia manganese superoxide dismutase (MnSOD) in chloroplasts. For this, the mature MnSOD-coding sequence was fused to a chloroplast transit peptide from a Pisum sativum ribulose-1,5-bisphosphate carboxylase (Rubisco) gene and expression of the chimeric gene was driven by the cauliflower mosaic virus (CaMV) 35S promoter. The transgenic MnSOD gene product was correctly targeted to the chloroplasts both in maize and tobacco. However, despite the use of the CaMV 35S promoter, the MnSOD was predominantly localized in the chloroplasts of the bundle sheath cells of maize. Furthermore, the transit peptide was cleaved off at a different position in maize and tobacco.

Amino Acid Sequence↗

PubMed1998

Gene expression induced by physical impedance in maize roots.

Two cDNA clones, pIIG1 and pIIG2, corresponding to mRNAs that accumulate in maize root tips subjected to 10 min of physical impedance, were isolated by differential screening of a cDNA library. The deduced proteins, based on DNA sequence analysis, have molecular masses of 13 and 23 kDa for pIIG1 and pIIG2, respectively. pIIG1 showed 97% similarity at the nucleic acid level to a maize root cortical cell delineating protein (pZRP3) and was also similar to some bimodular proteins that are developmentally or stress regulated in other plant species. In situ localization of pIIG1 showed some expression in cortical cells of control maize roots; however, after a 10 min physical impedance treatment, pIIG1 accumulation increased greatly in cortical cells and extended to include the procambial region. pIIG2 did not show sequence similarity with any identified gene of known function, but a bipartite nuclear targeting sequence occurs in its deduced amino acid sequence which indicates it may function in the nucleus. Thus, rapid accumulation of specific mRNAs occurs in maize roots in response to impedance stress, and these mRNAs may be responsible for some responses of the roots to physical impedance.

Amino Acid Sequence↗

PubMed1998

A ca. 40 kDa maize (Zea mays L.) embryo dehydrin is encoded by the dhn2 locus on chromosome 9.

Dehydrins (LEA D11 proteins) are the products of multigene families in a number of higher plants. To date, however, only one dehydrin locus, dhn1 (a major embryo and drought-induced protein of ca. 18 kDa) has been placed on chromosome 6L of the genetic linkage map of maize. The presence of a larger, ca. 40 kDa embryo protein that is also specifically detected by anti-dehydrin antibodies had been observed in some maize inbreds, including B73, suggesting that other dhn loci may exist. The ca. 22 kDa and ca. 40 kDa immunopositive proteins were purified from B73 and their amino acid compositions determined. The two proteins' amino acid compositions are typical of dehydrins, yet they differ from each other, indicating that they are distinct dhn gene products. Different size alleles for both proteins, or presence/absence in the case of the ca. 40 kDa protein, were evident from comparisons of embryo proteins of various maize inbreds. Analysis of segregating F2 progeny derived from self-pollination of F1 hybrids from four crosses (B73 x OH43, Mo17 x A632, AHO x A632, Latente x A632) revealed that alleles of the two genes assort independently. Map positions of the two dhn loci were then determined using two maize recombinant inbred line (RIL) mapping populations. The predicted map position of the gene controlling production of the ca. 22 kDa protein confirmed that this protein is the product of the dhn1 gene. The gene encoding the ca. 40 kDa dehydrin-like protein maps to a new locus on chromosome 9S near wx1, which we have named dhn2.

Amino Acid Sequence↗

PubMed1999

A transcript encoding translation initiation factor eIF-5A is stored in unfertilized egg cells of maize.

Differential screening of cDNA libraries of unfertilized egg cells and in vitro zygotes of maize resulted in the isolation of more than 50 different genes whose expression is up- or down-regulated after in vitro fertilization (IVF). Among these genes, we identified a cDNA encoding the eukaryotic translation initiation factor eIF-5A. This highly conserved factor is thought to be necessary for selective mRNA stabilization and translation. It is also the only known protein that contains the unusual amino acid hypusine which is required for biological activity. High transcript amounts are stored in the egg cell, which is, in terms of metabolism, relatively inactive. Upon fertilization transcript amounts decrease, in contrast to metabolically inactive embryos in which the transcript cannot be detected and transcript levels increase upon germination. The expression pattern during the first embryonic cell cycle is also different from that observed during the somatic cell cycle: egg cells in the G0 phase contain high transcript levels, while arrested suspension cells contain few transcripts. In the somatic cell cycle, eif-5A is strongly induced during the G1 phase and transcripts are continuously degraded during the S, G2 and M phases until new induction during the G1 phase of the next cycle. eif-5A, a member of a small gene family in maize, is expressed in most maize tissues investigated. Based on our results, we suggest that the unfertilized egg cell of maize, although relatively inactive regarding its metabolism, is prepared for selective mRNA translation that is quickly triggered after fertilization. We also suggest that the regulation of eif-5A in the first embryonic cell cycle is different from the somatic cell cycle.

Amino Acid Sequence↗

PubMed2001

Transcriptional activation of a maize calcium-dependent protein kinase gene in response to fungal elicitors and infection.

Plants respond to pathogen infection with the activation of the expression of pathogenesis-related genes, a response that involves Ca2+-regulated protein phosphorylation processes. We report here the isolation of a full-length complementary DNA encoding a calcium-dependent protein kinase (CPK) gene from maize. CPK genes occur in maize as members of a multigene family, but only one specific CPK gene, the ZmCPK10 gene here described, is transcriptionally activated in response to both fungal infection and treatment with fungal elicitors. Activation of the ZmCPK10 gene is extremely rapid. ZmCPK10 transcripts could be detected 5 min after elicitation and reached maximum levels at 30 min after treatment. Afterwards, there was a decline in the level of ZmCPK10 transcripts followed by a basal level of accumulation which is maintained over the time period of elicitor treatment. The activation of this kinase is accompanied by an increase in the level of PRms mRNA, the PRms being a pathogenesis-related protein from maize whose expression is induced in maize tissues in response to fungal infection and treatment with fungal elicitors. In situ mRNA hybridization analysis revealed a remarkable cell-type specific pattern of expression of ZmCPK10 during growth and development of the elicitor-treated or fungus-infected seedling. Moreover, the ZmCPK10 gene is expressed only in those specific cell types in which the PRms gene is also expressed. The involvement of ZmCPK10 in the elicitor-induced signal transduction pathway leading to the activation of PRms gene expression is discussed.

Amino Acid Sequence↗

PubMed1997

Handling and aflatoxin contamination of white maize in Costa Rica.

Projects funded by International Development Research Centre (IDRC) of Canada and the European Commission have enabled the examination of more than 3000 samples of maize collected from all regions of Costa Rica at different stages, from the growing crop through storage to final sale, and at different water contents. Contamination with Aspergillus flavus was frequent and about 80% of samples contained more than 20 ng aflatoxins g(-1) grain. Average contamination with aflatoxins in the Brunca Region was > 274 ng g(-1) while that in other regions was < 70 ng g(-1). Except in Brunca region, where it averaged 376 ng g(-1), contamination of grain from commercial sources was slightly less than of that from farms (< or = 15 ng g(-1)). It appeared that samples kept on the cob after harvest contained almost no aflatoxin while shelled samples were frequently highly contaminated. Experiments were therefore done in Brunca and Huetar Atlantic Regions, utilising 34 experimental maize crops to study in detail the development of A. flavus and aflatoxin from before harvest, through postharvest treatment before drying and through storage for six months. A. flavus was isolated more frequently from maize shelled immediately after harvest than from that kept on the cob until it could be dried, and from more samples from the Brunca Region than from the Huetar Atlantic Region. Samples harvested with > or = 18% water content often contains > 70% of grains infected with A. flavus but sometimes there were few grains infected. As found in the initial survey, more aflatoxin contamination developed in shelled maize than in that handled on the cob during the period from harvesting to drying, especially if the delay was more than 5 days, and more in Brunca than in Huetar. Shelled grain contained 400-800 ng aflatoxin g(-1) in Brunca but < 100 ng g(-1) in Huetar while grain kept on the cob contained < 30 ng g(-1), even with > 18% water content. Incidence of Fusarium spp. exceeded 50% except where A. flavus colonized more than 80% of grains.

Aflatoxins↗

PubMed1999

Occurrence of fumonisin B1 in maize and poultry feeds in Haryana, India.

A total of 100 maize and 50 poultry feed samples collected in 1998 at random from nine and eight districts of Haryana, respectively, were analysed for fumonisin B1. The samples were collected from poultry farms, feed manufacturers and markets. Ninety one (91%) maize samples and forty two (84%) poultry feed samples were found to contain fumonisin B1. Fumonisin B1 contamination in the maize samples ranged from 0.1-87.0 ppm. Whereas the poultry feed samples contained fumonisin B1 in the range of 0.02-28.0 ppm. It indicated widespread prevalence of fumonisin B1 in maize and poultry feeds in different areas of Haryana.

Animal Feed↗

PubMed1997

Chemical and sensory evaluation of vegetable milks from African yam bean Sphenostylis stenocarpa (Hochst ex A Rich) Harms and maize (Zea mays L.).

Vegetable milks were developed from fermented and unfermented African yam bean (AYB) flours and their maize blends. AYB was cleaned, dehulled, milled and fermented for 24 hours by the natural microflora present in the legume flour. Maize was fermented for 48 hours. A ratio of 70:30 (protein basis) of AYB: maize was used to formulate the blends. Vegetable milks were prepared from the AYB flours and their maize blends. Standard assay techniques were used to evaluate the milks for proximate, mineral, ascorbate and antinutrient composition. The protein contents of the milks (1.47-2.06 percent) was comparable to soymilk (2.04 percent) and bambara groundnut milk (2.00 percent). The milks contained appreciable quantities of carbohydrate and minerals tested. The milk blends had traces of ascorbate and contained higher phosphorus than the milks from the AYB flours. The fermented milk blend had higher protein, ash and sugar levels and lower phytate and stachyose levels compared to non-fermented blend. Raffinose was reduced to trace levels in the fermented milks. The milks were appetizing. The fermented milk blend was more acceptable than others and was preferred in terms of flavor and color. It had greater advantages over the other vegetable milks evaluated in terms of zinc, phosphorus and stachyose levels.

Fabaceae↗

PubMed1998

Processing of maize germ oil cake into edible food grade meal and evaluation of its protein quality.

The commercial oil cake produced during expeller pressing of maize germ, was extracted with n-hexane and 80 percent ethanol followed by seiving to remove undesirable materials. In defatted maize germ oil cake (DMGOC): protein, starch, fat, crude fiber (CF) and ash were respectively 24.69, 36.55, 5.68, 7.56 and 3.90 percent and they decrease after processing except ash, which increased slightly. It contains better quality protein having only 3 percent zein and 47 percent albumin. Its amino acids like lysine and tryptophan and biological value (BV) were higher than that of whole maize grain, and was comparable with that of the amino acid requirement of preschool children and casein diets both. Its digestible energy (DE) was lower compared with whole maize grain as well as the casein diets. After processing albumin, globulin and zein decreased whereas glutelin and the residual fraction increased. Not much differences were observed in chemical composition and different amino acids, in vitro protein digestibility (IVPD), true digestibility (TD), BV and DE improved after processing.

Amino Acids↗

PubMed2001

Selection and orientation of adjacent genes influences DAM-mediated male sterility in transformed maize.

Anther-targeted expression of E. coli DNA (Adenosine-N6-)-Methyltransferase (DAM) in maize was tested as a means to produce male-sterile plants. A high frequency of male-sterile plants with reduced anther size was observed when DAM was regulated by the maize anther-specific promoter 5126 (5126:DAM) and placed upstream of the herbicide resistance gene, pat, regulated by the cauliflower mosaic virus (CaMV) 35S promoter (35S:PAT). In contrast, placement of 5126:DAM upstream of a pat gene regulated by either the maize ubiquitin (UBI:PAT) or rice actin (rACTIN:PAT) promoters resulted in male-fertile plants. Based on these observed differences, DAM-mediated sterility was used as a phenotypic marker to assess the contribution of factors affecting gene expression such as orientation of the transcription units, choice of regulatory sequences mediating expression of adjacent genes, and effects of varying the anther-specific promoter regulating DAM. Constructs that place a portion of the CaMV 35S promoter, including the native AS-1 sequences, between 5126:DAM and UBI:PAT yielded a high frequency of male-sterile plants with reduced anther size. Significant differences in the frequency of male-sterile events and the associated anther size were also observed when the position of 35S:PAT was changed relative to 5126:DAM. These data provide evidence that gene expression in transformed maize plants can be impacted by simply altering the order, orientation or regulatory sequences of adjacent genes.

Acetyltransferases↗