Epistasis in maize (Zea mays L.)

Summary Three flint and three dent maize (Zea mays L.) inbred lines, their possible F₁ crosses, F₂ and backcross progenies, and all possible three-way crosses were evaluated in a three-year experiment for yield, ear moisture, and plant height. The purpose was to estimate genetic parameters in European breeding materials from (i) generation means analysis, (ii) diallel analysis of generation means, and (iii) analysis of F₁ and three-way cross hybrids. Method (i) was based on the F-metric model and methods (ii) and (iii) on the Eberhart-Gardner (1966) genetic model; both models extended for heterotic maternal effects.Differences among generation means for yield and plant height were mainly attributable to dominance effects. Epistatic effects were significantly different from zero in a few crosses and considerably reduced heterosis in both traits. Additive x additive and domiance x dominance effects for yield were consistently positive and negative, respectively. Significant maternal effects were established to the advantage of generations with a heterozygous seed parent. In the diallel analysis, mean squares for dominance effects were greater than for additive effects for yield and plant height but smaller for ear moisture. Though significant for yield and plant height, epistatic variation was small compared to additive and dominance variation. Estimates of additive x additive epistasis for yield were significantly negative in 11 of 15 crosses, suggesting that advantageous gene combinations in the lines had been disrupted by recombination in the segregating generations. The analysis of hybrids supported the above findings regarding the analysis of variance. However, the estimates of additive x additive epistasis for yield were considerably smaller and only minimally correlated with those from the diallel analysis. Use of noninbred materials as opposed to materials with different levels of inbreeding is considered the main reason for the discrepancies in the results.     

Histology of somatic embryogenesis in cultured immature embryos of maize (Zea mays L.)

Summary The developmental histology of somatic embryo (=embryoid) formation in cultured immature embryos of hybrid maize cultivars (Zea mays L.) is described. Embryos cultured on media containing 2% sucrose formed distinct globular embryoids. These embryoids arose either directly by divisions confined to the epidermal and the subepidermal cells at the coleorhizal end of the scutellum or from a soft and friable embryogenic callus produced by them. On media containing 6% sucrose divisions were initiated in the cells adjacent to the procambium of the cultured embryos. Subsequently, zones of meristematic cells also were observed in the region of the node and in the basal portion of the scutellum. Mature, well organized somatic embryos as well as a compact nodular type of embryogenic callus were produced as a result of localized meristematic activity along the tip of the scutellum toward the coleorhiza. Some embryos formed only the compact type of callus, and shoot primordia were organized later in the surface layers of this callus.Abbreviations CH casein hydrolysate - MS Murashige and Skoog's nutrient medium - 2,4-D 2,4-dichlorophenoxyacetic acid     

Effect of gibberellic acid on the ion ratios in a dwarf maize mutant (Zea mays L. d1)

The effect of gibberellic acid (GA₃) on ion ratios in a dwarf maize mutant (Zea mays L. d₁) exhibiting normal growth after hormone treatment has been investigated by electron microprobe analysis. Gibberellic-acid treatment increased the ion content in chloroplasts and vacuoles whereas no change of the ion content was found in the cytoplasm. The relation of these observations to the action of the hormone is discussed.Abbreviation GA₃ Gibberellic acid     

Genotypic effects on the amino acid relationships in maize (Zea mays L.) pollen and style

Summary The free amino acid content of the pollen grains and the style from three single cross hybrids (Wf9 x H55, Ky49 x Ky27, K64 x K55) and two inbred lines (Oh43, H55) was determined. No tyrosine was detected in the pollen grains of any genotype. Significant differences between pollen genotypes were found for aspartic acid, threonine, serine, lysine and histidine with no effect resulting from the vigor of the pollen source. No proline was found in the style of any genotype. Significant differences between style genotypes were obtained for threonine, serine, glutamic acid, leucine, tyrosine, ethanolanine, aminobutyric acid and histidine with a relationship between vigor of the style genotype present for tyrosine, ethanolanine and aminobutyric acid. The relationship between the pollen and style level for each amino acid as influenced by genotype was analyzed. A significant negative correlation was found only for threonine. Apparently, a complementary relationship between the pollen and style exists for some amino acids. Proline and tyrosine are available only from the pollen and style respectively. Threonine levels are balanced with varying contributions from both pollen and style depending on the genotype.     

Dehydrogenase and urease activities of maize (Zea mays L.) field soils

Summary Dehydrogenase and urease activities, bacterial and fungal populations and physicochemical characteristics of maize (Zea mays L.) field soils have been studied for one crop cycle. A comparison has been made among soils of three different agricultural systemsviz permanent agriculture on plain lands in valleys, recently introduced terrace land agriculture and age old ‘slash and burn’ type of shifting agriculture on slopes. Results demonstrate that the enzyme activities, microbial population as well as most of the physico-chemical characteristics of soils followed the trend permanent agriculture on plain lands>terrace land agriculture>‘slash and burn’ type of shifting agriculture. Moisture and nutrient levels and topography of the lands were found to be major factors responsible for the trend.     

Transformation of maize (Zea mays L.) protoplasts and regeneration of haploid transgenic plants

Transgenic haploid maize (Zea mays L.) plants were obtained from protoplasts isolated from microspore-derived cell suspension cultures. Protoplasts were electroporated in the presence of plasmid DNA containing the gus A and npt II genes encoding ß-glucuronidase (GUS) and neomycin phosphotransferase II (NPT II), respectively. Transformed calli were selected and continuously maintained on kanamycin containing medium. Stable transformation was confirmed by enzyme assays and DNA. analysis. Stably transformed tissue was transferred to regeneration medium and several plants were obtained. Most plants showed NPT II activity, and some also showed GUS activity. Chromosome examinations performed on representative plants showed that they were haploid. As expected, these plants were infertile.     

Arylsulphatase activity of corn (Zea mays L.) seedling roots

The crude lysosomal fraction of corn seedling root tips contains an arylsulphatase (E.C. 3.1.6.1) which hydrolysed p-nitrophenyl sulphate at pH 8.0 but had no activity towards p-nitrocatechol sulphate. The Km value for p-nitrophenyl sulphate was 1.24 mM. The hydrolysis of p-nitrophenyl sulphate was linear up to 2 h and the rate was proportional to the amount of enzyme added. The enzyme was strongly inhibited by cyanide, fluoride and phosphate ions and did not resemble the arylsulphatases of bacterial and animal origin.     

Gynogenetic haploid plants analysis for agronomic and enzymatic markers in maize (Zea mays L.)

Summary Experiments were conducted to investigate whether selection occurs during the processes involved in the production of doubled haploids. Haploid plants produced from two hybrids, each heterozygous for isozyme markers, were subjected to genetic analysis. The distributions of doubled haploid lines and pedigree lines derived from the hybrid C123 x Oh7 were compared with regard to agronomic character. The results suggest that the populations of haploid plants obtained by in vivo gynogenesis represent a random gametic array. Thus, in order to introduce haploid plants into breeding programmes in maize, maternal haploidy seems to be a very attractive method.     

The microtubular cytoskeleton in cells of cold-treated roots of maize (Zea mays L.) shows tissue-specific responses

Summary Microtubules (MTs) in cells of various tissues at different distances from the apex of the maize root exhibited different sensitivities to cold (5 °C), as judged by MT reorientation and tendency to depolymerization. Their responses seem to be related to their initial intracellular arrangements. Generally, MTs in cells which were ceasing elongation were the least sensitive during the early stages (6–24 h) of cold treatment, but during the later stages (5–7 d) MTs in most of these cells eventually depolymerized. Pericycle cells showed a unique cold response. Here the MTs were conspicuously cold-labile and quickly depolymerized near the root-tip. However, after 1 d many pericycle cells in more proximal regions had repolymerized their MTs as dense, randomly organized arrays. These persisted for the remainder of the cold treatment. A similar resistance to longterm chilling, by means of MT repolymerization, was found in cells of the root cap, quiescent centre and cells of the distal part of the former meristem. MT repolymerization in the cold may enable the apex to resume growth when more favourable (warmer) conditions return.Abbreviations DAPI 4,6-diamidino-2-phenylindole - DMSO dimethylsulfoxide - EGTA ethylene glycol-bis(-aminoethylether)-N,N,N,N-tetraacetic acid - FITC fluorescein isothiocyanate - IgG immunoglobulin G - MT microtubule - PEG polyethylene glycol - PIPES piperazine-N,N-bis(diethanesulfonic acid)     

Genetic control of maternal haploidy in maize (Zea mays L.) and selection of haploid inducing lines

Summary The effect of genotype on maternal haploid plant production in maize was studied. The frequency of gynogenetic plants when Stock 6 was used as pollinator varied according to the female parent genotype. No simple relation was observed between genotypic aptitudes for gynogenetic and androgenetic development, which occured after pollination of W23 plant carrying the indeterminate gametophyte gene. Furthermore, the population NS, a favorably responsive genotype to anther culture, does not exhibit exceptional ability for in vivo gynogenesis. The effect of inbreeding and the influence of maternal haploid origin suggest that specific genes control maternal haploid initiation and development. However, gynogenetic development is not limited to a particular genotype. The frequency of maternal haploids may be increased by using specific pollen parents. Attempts were made to select for a high haploidyinducing trait and the present study reports the successful development of lines that can be utilized as pollen parents to induce haploids for experimental purposes and breeding programmes. When an inbred line WS14, derived from the cross W23 x Stock 6, was used as pollen parent, 2%–5% maternal haploids were obtained according to the female parent genotype. A high haploidy-inducing potential is a heritable trait and may be controlled by a limited number of genes. Genetic determination of the haploidy-inducing character was examined in relation to the efficiency of the selecting method and the mechanisms involved in the origin of maternal haploids.     

Duplicated chromosome segments in maize (Zea mays L.): further evidence from hexokinase isozymes

Summary The genetic control of hexokinase isozymes (ATP: d-hexose-6-phosphotransferase, E.C. 2.7.7.1, HEX) in maize (Zea mays L.) was studied by starch gel electrophoresis. Genetic analysis of a large number of inbred lines and crosses indicates that the major isozymes observed are encoded by two nuclear loci, designated Hex1 and Hex2. Five active allozymes and one null variant are associated with Hex1, while Hex2 has nine active alleles in addition to a null variant. Alleles at both loci govern the presence of single bands, with no intragenic or intergenic heteromers visible, suggesting that maize HEX's are active as monomers. Organelle preparations demonstrate that the products of both loci are cytosolic. All alleles, including the nulls, segregate normally in crosses. Vigorous and fertile plants were synthesized that were homozygous for null alleles at both loci, suggesting that other hexosephosphorylating enzymes exist in maize that are undetected with our assay conditions. Linkage analyses and crosses with B-A translocation stocks place Hex1 on the short arm of chromosome 3, 27 centimorgans from Pgd2 (phosphogluconate dehydrogenase) and Hex2 on the long arm of chromosome 6, approximately 45 centimorgans from Pgd1. It is suggested that the parallel linkages among these two pairs of duplicated genes reflects an evolutionary history involving chromosome segment duplication or polyploidy.Paper No. 10170 of the Journal Series of the North Carolina Agricultural Research Service, Raleigh, NC     

In vitro development of young maize ears

The effects of sucrose, cytokinin and auxin on in vitro growth and development of immature maize ears (female inflorescences) isolated from the first and second nodes of a prolific and a nonprolific hybrid were evaluated. The explants were cultured on an agar-solidified Murashige and Skoog medium and tested at four sucrose concentrations, five cytokinin (6-benzylamino purine and zeatin) concentrations, and six zeatin/indole-3-acetic acid (IAA) combinations. Sucrose at 60 g l^–1 resulted in the best ear growth and development in both hybrids and both node positions. Ear lengths from the first and second nodes of the prolific hybrid were further increased by 100 and 62% respectively, by the addition of zeatin (1 mg l^–1) + IAA (1.5 mg l^–1), whereas the ears of the nonprolific hybrid increased by 45 and 33%, respectively. Ear development was also increased by these treatments (114, 166, 57 and 57%, respectively), compared with the control without hormones. When growing in vivo, the second ears of the prolific hybrid were able to reach anthesis (stigma exposition) while in the nonprolific they could not develop beyond the style elongation stage.     

Mutations in corn (Zea mays L.) conferring resistance to imidazolinone herbicides

Summary Three corn (Zea mays L.) lines resistant to imidazolinone herbicides were developed by in vitro selection and plant regeneration. For all three lines, resistance is inherited as a single semidominant allele. The resistance alleles from resistant lines XA17, XI12, and QJ22 have been crossed into the inbred line B73, and in each case homozygotes are tolerant of commercial use rates of imidazolinone herbicides. All resistant selections have herbicide-resistant forms of acetohydroxyacid synthase (AHAS), the known site of action of imidazolinone herbicides. The herbicide-resistant phenotypes displayed at the whole plant level correlate directly with herbicide insensitivity of the AHAS activities of the selections. The AHAS activities from all three selections have normal feedback regulation by valine and leucine, and plants containing the mutations display a normal phenotype.     

Behaviour of chromosomes in anaphase cells in embryogenic callus cultures of maize (Zea mays L.)

Mitotic anaphase cells of highly friable and embryogenic calluses which had been induced from immature embryos of two inbred lines of maize that have contrasting levels of heterochromatic knobs were analysed for the presence of abnormalities 3, 6, 9 and 12 months after the initiation of culture. A total of 500 typical anaphases was scored at each time point, and various aberrations, such as delay in the separation of sister chromatides, chromosome bridges (single, double and multiple) and chromosome fragments, were revealed to occur extensively in the cultures of both genotypes. Preparations after C-banding revealed that primary breakages often occurred inside knobs or at junction regions between the euchromatin and the heterochromatin of the knobs. Figures characterized by the delayed separation of sister chromatids, which originated preferentially at the knob level and was considered to be an initial event in the development of breakages, were observed at constant frequencies throughout the experiment. Increasing numbers of aberrant cells were detected with time, mainly due to the accumulation of cells with chromosome bridges and fragments. Several mitotic figures suggested the occurrence of breakagefusion-bridge cycles that were initiated by broken chromosomes. The overall frequencies of aberrant cells were similar for both genotypes, despite the differences in knob composition. However, callus cultures induced from the genotype having the higher level of knobs had more aberrant cells with abnormalities that involved several chromosomes, such as multiple bridges and multiple fragments.     

In vitro pollination as a model for studying fertilization in maize (Zea mays L.)

Summary In vitro pollination was conducted using excised segments of maize female spikelets to determine the effects of age and silk length on fertilization efficiency and developmental pattern. Ovary development after 15 days resulted in: (1) normal kernels, (2) abnormal kernels and (3) enlarged ovaries; the percentages of each class varied with age. Evidence of double fertilization was observed in both normal and abnormal kernels. In vitro fertilization was traced using silk excision and autoradiography with ³²P-radiolabelled pollen and occurred between 4 and 7 h after the pollination of 4.5-cm-long silks. This study supports the validity of the in vitro pollination method for studying fertilization and emphasizes the importance of using a developmentally sensitive index (silk length) for establishing female developmental stage.     

Sterols of male and female compound inflorescences of Zea mays L.

A comparison has been made between the sterols of male and female inflorescences and of pollen from Zea mays. The female inflorescence was shown to contain cholesterol, 24-methylcholesterol, 24-ethyl-5,22-cholestadien-3-ol, 24-ethylcholesterol and (28Z)-24-ethylidenecholesterol. Themale inflorescence contained the same five compounds together with 24-methylenecholesterol. Pollen contained 24-methylenecholesterol as its main sterol together with lesser amounts of cholesterol, 24-ethylcholesterol, (28Z)-24-ethylidenecholesterol, 24-methylene-5-cholest-7-en-3-ol and 4-methyl-24-methylene-5-cholest-7-en-3-ol.     

Zinc deficiency and pollen fertility in maize (Zea mays)

Zinc deficiency decreased pollen viability in maize (Zea mays L. cv. G2) grown in sand culture. On restoring normal zinc supply to zinc-deficient plants before the pollen mother cell stage of anther development, the vegetative yield of plants and pollen fertility could be recovered to a large extent, but the recovery treatment was not effective when given after the release of microspores from the tetrads. If zinc deficiency was induced prior to microsporogenesis it did not significantly affect vegetative yield and ovule fertility, but decreased the fertility of pollen grains, even of those which visibly appeared normal. If the deficiency was induced after the release of microspores from the tetrads, not only vegetative yield and ovule fertility but pollen fertility also remained unaffected.     

Role of sulphur availability on cadmium-induced changes of nitrogen and sulphur metabolism in maize (Zea mays L.) leaves

The interactions between sulphur nutrition and Cd exposure were investigated in maize (Zea mays L.) plants. Plants were grown for 12 days in nutrient solution with or without sulphate. Half of the plants of each treatment were then supplied with 100 microM Cd. Leaves were collected 0, 1, 2, 3, 4 and 5 days from the beginning of Cd application and used for chemical analysis and enzyme assays. Cd exposure produced symptoms of toxicity (leaf chlorosis, growth reduction) and induced a noticeable accumulation of non-protein SH compounds. As phytochelatins are glutamate- and cysteine-rich peptides, the effect of cadmium on some enzyme activities involved in N and S metabolism of maize leaves was studied in relation to the plant sulphur supply. In vivo Cd application to S-sufficient plants resulted in a drop of all measured enzyme activities. On the other hand, S-deficient plants showed a decrease in nitrate reductase (NR; EC 1.6.6.1) and glutamine synthetase (GS; EC 6.3.1.2) activity, and an increase in NAD-dependent glutamate dehydrogenase (GDH; EC 1.4.1.2) and phosphoenolpyruvate carboxylase (PEPc; EC 4.1.1.31) activity as a result of the Cd treatment. Furthermore, in the same plants ATP sulphurylase (ATPs; EC 2.7.7.4) and O-acetylserine sulphydrylase (OASs; EC 4.2.99.8) showed a particular pattern as both enzymes exhibited a transient maximum value of activity after 4 days from the beginning of Cd exposure. Results provide evidence that the increase of ATPs, OASs, GDH and PEPc activities, observed exclusively in S-deficient Cd-treated plants, may be part of the defence mechanism based on the production of phytochelatins.     

A comparison between minirhizotron and monolith sampling methods for measuring root growth of maize (Zea mays L.)

Transparent plastic minirhizotron tubes have been used to evaluate spatial and temporal growth activities of plant root systems. Root number was estimated from video recordings of roots intersecting minirhizotron tubes and of washed roots extracted from monoliths of the same soil profiles at the physiological maturity stage of a maize (Zea mays L.) crop. Root length was measured by the line intercept (LI) and computer image processing (CIP) methods from the monolith samples.There was a slight significant correlation (r=0.28, p<0.005) between the number of roots measured by minirhizotron and root lengths measured by the LI method, however, no correlation was found with the CIP method. Using a single regression line, root number was underestimated by the minirhizotron method at depths between 0–7.6 cm. A correlation was found between root length estimated by LI and CIP. The slope of estimated RLD was significant with depth for these two methods. Root length density (RLD) measured by CIP showed a more erratic decline with distance from the plant row and soil surface than the LI method.