Beáta Barnabás

Direct effect of colchicine on the microspore embryogenesis to produce dihaploid plants in wheat (Triticum aestivum L.).

SummaryIn the present experiment, different chromosome reduplication techniques were applied to microspore-originated Triticum aestivum L. (cv ‘Ciano’) haploids. In addition to the conventional treatment (whole plant exposure to colchicine solution), spontaneously redoubled haploids were also examined. As an experimental treatment, different concentrations (0.01, 0.02, 0.04%) of colchicine were added directly to the induction media. Colchicine did not affect the anther response or the plant regeneration capacity. The success and stability of genome redoubling was estimated on the basis of the fertility of the regenerated (R0) plants and their progeny (R1). Chromosome doubling produced by colchicine before the first microspore mitosis was significantly more efficient than the conventionally used techniques.

Colchicine, an efficient genome-doubling agent for maize (Zea mays L.) microspores cultured in anthero

Abstract The construction of maize genotypes with high haploid induction capacity made it possible to study the effect of colchicine on maize androgenesis in vitro. Anther cultures of three hybrids were treated with 0.02% and 0.03% colchicine for 3 days at the beginning of microspore induction. Colchicine added to the induction medium had no negative influence on the androgenic responses (anther induction, induction of structures of microspore origin and their regeneration ability) of the genotypes examined. However, significantly higher fertility was observed in plants originating from colchicine-treated microspores, especially at 0.03%. Cytological examinations showed that colchicine treatment before the first microspore division efficiently arrested mitosis and resulted in homozygous doubled-haploid microspores. Under the experimental conditions, the antimitotic drug had no later effect on the division symmetry of the microspore nucleus, and unequal divisions remained dominant. Callus formation from the induced microspores seemed to be more typical (ranging between 60–70%), but embryo frequency was increased by approximately 10%, especially at the higher colchicine concentration. These results suggest that the mechanism of colchicine action in premitotic maize microspores may differ from that previously observed in wheat.

Storage of maize (Zea mays L.) pollen at −196°C in liquid nitrogen

SummaryThe water content of pollen has a decisive influence on its storability in liquid nitrogen. Pollen with an initial high water content cannot be stored successfully at extremely low temperatures, so a certain degree of drying must be carried out before storage. Provided the viability of the pollen is not significantly reduced during drying, the pollen remains viable and fertile when kept at −196°C.

Substitution analysis of callus induction and plant regeneration from anther culture in wheat (Triticum aestivum L.).

The genetic determination of callus induction, total plant regeneration and green plant regeneration from anther culture were studied using a “Chinese Spring”/“Cheyenne” substitution series. All the three characteristics were found to be polygenically determined, but their inheritance was independent from one another. The 7A and 18 chromosomes had a considerable effect on callus induction. In the case of total plant regeneration the most influential chromosome “as the 3A while the 2D chromosome showed a definite influence on green plant regeneration. The interaction between the genetic background of the recipient plant and the substituted chromosome plays an important role in the manifestation of the studied features.

Embryo and endosperm development in wheat ( Triticum aestivum L.) kernels subjected to drought stress

The aim of the present work was to reveal the histological alterations triggered in developing wheat kernels by soil drought stress during early seed development resulting in yield losses at harvest. For this purpose, observations were made on the effect of drought stress, applied in a controlled environment from the 5th to the 9th day after pollination, on the kernel morphology, starch content and grain yield of the drought-sensitive Cappelle Desprez and drought-tolerant Plainsman V winter wheat (Triticum aestivum L.) varieties. As a consequence of water withdrawal, there was a decrease in the size of the embryos and the number of A-type starch granules deposited in the endosperm, while the development of aleurone cells and the degradation of the cell layers surrounding the ovule were significantly accelerated in both genotypes. In addition, the number of B-type starch granules per cell was significantly reduced. Drought stress affected the rate of grain filling shortened the grain-filling and ripening period and severely reduced the yield. With respect to the recovery of vegetative tissues, seed set and yield, the drought-tolerant Plainsman V responded significantly better to drought stress than Cappelle Desprez. The reduction in the size of the mature embryos was significantly greater in the sensitive genotype. Compared to Plainsman V, the endosperm cells of Cappelle Desprez accumulated significantly fewer B-type starch granules. In stressed kernels of the tolerant genotype, the accumulation of protein bodies occurred significantly earlier than in the sensitive variety.

The isolation of viable egg cells of wheat (Triticum aestivum L.)

The isolation of viable egg cells of wheat has been achieved without enzymatic maceration of the ovules. 2,4-D applied to the stigmas resulted 3 to 7 days later in soft ovule tissues which disintegrated upon mechanical manipulation. The isolated egg cells were viable even 2 h after isolation. Their morphology corresponded to that of the in situ egg cells. The mean isolation frequency was 20% (two egg cells per ten ovules).

Electro-fused isolated wheat (Triticum aestivum L.) gametes develop into multicellular structures.

The electrofusion-mediated fertilization of single egg cells of wheat with isolated individually selected wheat sperm cells was successfully carried out for the first time. On average the fusion frequency was 30% but under optimal conditions it was possible to reach as much as 55%. Two days after electric fusion 60% of the fusion products started to divide, 88.5% of them forming multicellular structures and in a few cases microcalluses. The culture of single unfertilized egg cells with or without the application of AC field and electric pulses induced no cell division. The egg cells and fusion products were cultured in a maize feeder-cell system.

In Vitro Androgenesis of Wheat from Fundamentals to Practical Application

Wheat anther cultures have a history of almost 30 years and are nowemployed efficiently in many countries of the world for the developmentof doubled haploid lines for breeding. The present paper discusses keyquestions of the elaboration and perfection of the method: cytologicalaspects of in vitro androgenesis, the conditions required for theembryogenic development of microspores and the applicability of anthercultures in the Martonvásár wheat breeding programme.

Ultrastructural studies on pollen embryogenesis in maize (Zea mays L.).

Maize anthers have been induced on modified N6 medium to produce embryoids. Different stages from the cultures were sampled and prepared for microscopical examination. The microspores at the onset of culture were in an early developmental stage, with the nucleus and numerous organelles centred in the middle, surrounded by many small vacuoles with a lipid content. The binuclear pollen grains contained small vesicles and much starch. The partially condensed vegetative nucleus indicated participation of the vegetative component in the formation of multicellular pollen grains (MPGs). Several MPGs have been observed which differed in morphology. We suggest, on the basis of these ultrastructural observations, that in maize mainly the vegetative cell contributes to the MPG which further develops directly into embryoids.

Colchicine induced embryogenesis in maize

The present study involves in vitro androgenesis of Zea mays L. using anther culture. We tested combinations of single factors and their influence on microspore induction. Embryogenic induction of microspores within anthers in in vitro conditions was the best when combination of cold treatment, TIBA (0.1 mg l−1) in media and colchicine (0.02% during first 3 days of culture) was applied, but colchicine alone can be factor, which can stimulate or initiate embryogenesis in anther culture of maize.