Green Plant Regeneration Research Articles

Nuclear and Cytoplasmic Control of Another Culture Response in Wheat: I. Analyses of Alloplasmic Lines

These studies were initiated to obtain definitive evidence for the role of cytoplasm genetic variation in anther culture and to test for nuclear ✕ cytoplasmic gene interaction on the anther culture responses in wheat (Triticum aestivum L.). Both cytoplasmic and nuclear genetic variations are potentially exploitable for practical purposes through recombination, breeding and selection. The roles of cytoplasmic and nuclear genetic factors in the control of anther culture responses were investigated by comparing the culture responses of alloplasmic lines in three spring wheat cultivars (Selkirk, Siete Cerros 66, and Penjamo 62) and their nucleus donors. Significant increases or decreases caused by alien cytoplasms were observed for callus induction, plant regeneration (albino and green plants), or green plant regeneration. In addition, significant nucleus ✕ cytoplasm genetic interactions were found for all three components of anther culture responses in two subsets of alloplasmic lines having common cytoplasms but different nuclear genotypes. Each of the components of anther culturability in wheat may be controlled independently by cytoplasmic and nuclear genes as well as by nucleus ✕ cytoplasm gene interactions.

Nuclear and Cytoplasmic Control of Anther Culture Response in Wheat: III. Common Wheat Crosses

Anther culture represents a potentially important technology for production of wheat doubled haploids for plant breeding and genetics research; however, the yields of haploids from culture must be increased to achieve competitive efficiencies for the method. Genetic differences for culturability are now well known, but the relative roles of nuclear and cytoplasmic genetic factors are still unclear. The purpose of this study was to determine the significance of cytoplasm genetic variation for anther culture responses among common T. aestivum wheats. Plants were grown in a greenhouse and potato‐4 and 190‐2 media were used as callus induction and plant regeneration media, respectively. Large genotype differences in an ther culture response were observed. Among the 18 pairs of reciprocal crosses studied, 9 showed reciprocal differences for green plant regeneration, 4 showed significant differences for callus induction, and 3 were different for plant regeneration. The occurrence of reciprocal differences for all components of anther culture response, especially for green plant regeneration, indicates the the cytoplasm of microspores or maternally inherited factors interact with nuclear genes to control the response of wheat genotypes to anther culture. The results provide substantial new supporting evidence that nuclear, cytoplasm, and nuclear ✕ cytoplasm genetic interactions control the responses of common wheats to anther culture.

Anther callus induction and green plant regeneration at high frequencies from an interspecific rice hybrid Oryza sativa Linn. x O. rufipogon Griff

Callus induction and green plant regeneration at high frequencies from an interspefic hybrid, Oryza sativa L. x O. rufipogon Griff. has been achieved by simply coordinating the growth regulators in the induction medium. The study was conducted with two different basal media (Potato-2 and N6) and seven different combinations of growth regulators 2,4-D, NAA and kinetin. Synergistic effects of the two auxins in enhanced anther response to callus induction and subsequent green plant regeneration were observed in both media. The highest frequency of callus induction was obtained on Potato-2 medium supplemented with 1 mg/12,4-D, 2 mg/l NAA and 1 mg/l kinetin. The same combination of growth regulators which yielded higher frequencies of callus induction also induced higher mean number of calli per anther. Although the calli formed on N6 medium showed high regenerability, there was a concomitant increase in the number of albinos among the regenerants. The auxins in the induction media had considerable influence on the regeneration capacity of the calli. The regeneration frequencies were higher from calli formed in the presence of both auxins in the induction media. The levels of growth regulator combinations seem to influence the green plant regeneration especially for calli induced on Potato-2 medium. Among the pollen grain derived plants the majority were either haploids or double haploids and very few were chromosomal variants.

Low-Dose Gamma-Irradiation Promotes Wheat Anther Culture Response

This study examined the anther culture response of wheat to different doses of gamma radiation, and the interaction of radiation dose with a cold pretreatment of ears stored prior to culturing. The cultivars Grebe and Kite were chosen on the basis of their anther culture response, Grebe being highly responsive and Kite being non-responsive. Spikes of the two cultivars were exposed to various levels of gamma radiation (60Co source) ranging from 0 to 10 Gy (0.53 Gy min-1) before anthers were plated on an agarose-solidified medium. For Grebe, doses of 1, 3 and 5 Gy resulted in more embryoids, higher green plant regeneration, and a greater number of spontaneously doubled haploids (DH), than in the non-irradiated control. The response in Kite was similar but less pronounced. The higher doses of radiation (7 and 10 Gy), however, were detrimental in both cultivars and at 10 Gy no embryoids or regenerants were produced. Anthers subjected to a cold pretreatment prior to irradiation responded significantly less than those cultured fresh. This study indicated that low-dose gamma irradiation of fresh explants can significantly improve regeneration from anther cultures in wheat and may stimulate a low frequency of regeneration in an otherwise non-responsive cultivar.

Effects of amino acids on callus differentiation in barley anther culture

Supplements of 8 mgl-1 L-Alanine (L-Ala), L-Asparagine (L-Asn), L-Glutamine (L-Gln) and L-Alanine + L-Asparagine to modified MS media increased the frequency of differentiation and the percentage of green plant regeneration in barley callus. It also decreased the ratio of albino to green plants. The actions of amino acids were related to distinct isozyme electrophoretic patterns, the activity of peroxidase and the mino acids content of the callus. The possible mechanisms of action of amino acids on callus differentiation and green plant regeneration are briefly discussed.

Investigations on the Anther Culturability of Four German Spring Wheat Cultivars and the Influence of Light on Regeneration of Green vs. Albino Plants

AbstractIn anther culture experiments with four German spring wheat cultivars (‘Turbo’, ‘Kadett’, ‘Planet’ and ‘Nandu’), ‘Nandu’ showed the best results. The rate of responding anthers with ‘Nandu’ was 0.8 % on medium N6 and 3.7% on potato‐2. This was similar to the Chinese cultivar ‘Orofen’. Higher yields of embryoids could be obtained with medium potato‐2 than with N6.Experiments with medium potato‐2 showed that anther response, production of embryos and regeneration increased with decreasing agarose concentration.Regeneration of green plantlets was improved by differentiation in the dark whereas differentiation under light yielded more than 90 % albino plants.

Improved plant regeneration from shed microspore culture in barley (Hordeum vulgare L.) cv. igri

This report describes rapid regeneration of green plants from microspores of the barley cultivar Igri. Use of 0.3 M mannitol during maceration and isolation was essential for response from mechanically isolated microspores of barley cv. Igri grown under our conditions. A shed microspore culture system proved to be simple and gave a fast response; plants were obtained as early as 25 days after the material was taken from the donor plant. A 28-day cold-pretreatment of spikes can also be replaced with a 3-4 day pretreatment of anthers in mannitol. Shed microspores from 100 anthers produced an average of 292 plants with 91% of them green. Approximately 80% of the regenerated plants were spontaneously doubled-haploids.

Effect of genotype and medium on wheat (Triticum aestivum L.) anther culture

Four winter wheat (Triticum aestivum L.) and two spring wheat cultivars were evaluated in anther culture on three to four different media for their ability to initiate callus and green plants. Five media were used in the experiment: stored-potato medium with Ficoll 400, fresh-potato medium with Ficoll 400, fresh-potato medium with agar, fresh-potato liquid medium without agar or Ficoll 400, and a one tep 85D12-3 medium. Greatly different frequencies of calli and/or green plants were obtained from different cultivars and media. The callus initiation frequency varied from 2.7% for Arapahoe to 52% for Pavon, both on the stored potato medium with Ficoll 400. The frequency of green plant regeneration ranged from 0% for Arapahoe and Siouxland on the stored-potato medium with Ficoll 400 and 0% for Redland and Arapahoe in the fresh-potato medium with Ficoll 400 to 12% for Chris in the 85D12-3 medium (one-step procedure). Chris and Centurk 78, previously reported as having high levels of response, had significantly higher (P < 0.05) frequencies of green plant regeneration on the 851312-3 medium than the other cultivars. An unexpected observation is that ‘wet’ MSC− medium enhanced callus regeneration more than a drier MSC− medium.

Embryogenesis and plant regeneration from microspores of both ‘Indica’ and ‘Japonica’ rice ( Oryza sativa)

Microspore embryogenesis of rice (both indica and japonica types) was induced in culture yielding more than 200 green plants. Direct embryo germination and more frequently plant regeneration with multiple tillers have been obtained through secondary embryogenesis. Cold pre-treatment and the use of modified MS medium containing reduced nitrogen (glutamine and casein hydrolysate) and 10% (w/v) Ficoll might have favored microspore embryogenesis and green plant regeneration. Response of microspores in regeneration, embryogenesis and albinism varies depending on the constituents of the media and genotypes used.

THE INFLUENCE OF VARIOUS CULTURAL FACTORS ON ANTHER CULTURE OF FOUR CULTIVARS OF SPRING WHEAT (Triticum aestivum L.)

Four spring wheat cultivars (Katepwa, Columbus, Chris and Israel 676) were compared for androgenic ability. Temperature, medium, agar concentration and cold pretreatment were examined to find the optimal combinations for anther culture of Katepwa and Columbus in particular. At an incubation temperature of 30 °C, anthers of Chris were significantly more responsive than the other genotypes. In three temperature treatments (26, 28 and 30 °C) the potato extract medium (P1) was superior to the synthetic medium (N6) (Chu 1978) for callus induction and green plant regeneration. Over all genotypes, liquid media provided greater anther response than either 0.8% or 1.8% agar. At 28 °C, cultured anthers showed no significant increase in callus formation after being subjected to a cold pretreatment. A cold pretreatment prior to culture at 30 °C elicited a significant decrease in callus formation as compared with no cold pretreatment. However, maximum callus induction and regeneration frequencies for Katepwa and Columbus were obtained at 28 °C in the P1 liquid medium with cold pretreatment. An incubation temperature of 26 °C was inferior to 28° and 30 °C for callus induction and regeneration.Key words: Anther culture, wheat, genotype, incubation temperature, media, agar concentration

Diallel Analysis of Callus Induction and Green‐Plant Regeneration in Rice Anther Culture

Low efficiencies of callus induction and green‐plant regeneration have limited the application of anther culture in breeding programs. This study was conducted to determine the inheritance of callus induction and plant regeneration in rice (Oryza sativa L.) anther culture. Sixteen genotypes derived from a four‐parent diallel mating set of two Japonica cultivars (Taipei 309 and Taipei 177) and two indica cultivars (IR36 and Basmati 370) were evaluated for their callus induction and plant regeneration abilities. For callus induction, the effects of genotype and genotype ✕ medium interaction were significant (P &lt; 0.05). Similarly, genotype and abscisic acid (ABA) level were found to be significant (P &lt; 0.05) for green‐plant production. The japonica cultivars generally had a higher percent callus induction and plant regeneration, and number of calli producing green plants than did the indica cultivars. Similar modes of inheritance were observed for both characters. Combining ability analysis demonstrated the predominance of additive gene effects in the control of both characters with the japonica cultivars having higher combining ability for green‐plant regeneration. Reciprocal effects were not significant (P &gt; 0.05), suggesting the absence of maternal control on the characters. Further analysis showed gene action to be partially dominant with high response being highly recessive and controlled by a few genes. There were no indications of inter‐allelic interaction.

Components of Response in Barley Anther Culture

AbstractAnther culture response with 17 widely‐grown varieties and one model variety of barley was studied with one replication from field‐grown donor plants and one replication from a growth‐chamber. Plants were regenerated from all 18 varieties and green plants were obtained from 16 of them. On average, 1.6 green plants were obtained per 100 cultured anthers from all the material. Estimated variance components for the formation of embryos/callus from the anthers were dominated by the effects of the genotypes and interactions between plant material and environments which together accounted for 60.1 and 17.0 % of the total variation respectively, while environments were nonsignificant for this character. Plant regeneration from embryos/callus were not significantly influenced by either genotype or environments. Components of variance for green plant formation were dominated by the effects of the genotypes, accounting for 73.2 % of the total variation, and a smaller effect from environments accounting for 11.2% of the total variation. Main effects from genotypes on the percentage of green regenerants divided 7 varieties into two distinct groups, indicating that major genetic factors were involved. The genetic basis for green plant regeneration seems different from that governing embryo formation. The results are discussed with respect to the possible prediction of anther culture response for new barley hybrids, as a means for directing the use of barley anther culture towards material that responds well.

Effects of donor plant environment and light during incubation on anther cultures of some spring wheat (Triticum aestivum L.) cultivars

Four different growth environments (field, two phytotron greenhouses and one growth chamber) were compared, using two genotypes of spring wheat, one recalcitrant and one responsive. Field-grown plants gave inferior results. Large improvements could be made by improving the conditions, embryoid frequencies in the two genotypes reaching 77.1% and 183.9% per 100 anthers, respectively. High light intensity during the induction phase strongly suppressed induction in both genotypes, but stimulated regeneration of green plants in the recalcitrant genotype, which had the lowest regeneration ability. Weak, diffuse light did not inhibit induction while the positive effect on regeneration was maintained. Also, another recalcitrant genotype was grown in the field, together with two F1-hybrids (recalcitrant x recalcitrant and recalcitrant x responsive). Evidence for a three-factor system was obtained.

Varietal differences in green plant regeneration ability of wheat pollen embryoids.

The present study was undertaken in order to clarify the effect of the culture media and the genotypes on the regeneration of green plantlets from pollen embryoids by anther culture of Japanese wheat cultivars. Pollen embryoids of 4 cultivars produced on Potato-2 medium were transferred onto two kinds of regeneration media (Table 1). On both media, embryoids grew into calli and then regenerated green or albino shoots and/or roots (Fig. 1 b and c). In rare cases, embryoids germinated (Fig. 1 a). The frequencies of green plantlets regenerated from embryoids differed with their genotypes even in the regeneration medium from 2.8% in Nanbukomugi to 33.5% in Chinese Spring. Moreover, the ability of embryoids to regenerate green plantlets was not affected by anther culture media, Potato-2 and C1 medium (Table 2). The effects of abscisic acid (ABA) on the embryoid formation and green plantlet regeneration were studied in anther culture of 7 cultivars (Table 3). ABA at a low concentration (0.1 and 0.5mg/l) had a stimulatory effect on the embryo formation in some cultivars which were genotypes of a rather low ability in embryoid formation, while ABA had a suppressive effect in other cultivars. However, ABA did not show a stimulatory effect on the regeneration of green plantlets in all cultivars tested. In conclusion, the ability of the green plantlet regeneration depended on genotypes regardless of the anther culture and regeneration media tested in this study: Norin 12 and Chinese Spring showed relatively high frequency of the green plantlet regeneration, while Hachimankornugi, Kitakarnikornugi and Norin 61 did not regenerate green plantlets. On chromosome constitutions of the regenerated green plants almost 659 were haploid (2n=21) and about 35% were diploid (2n=42) (Table 4, Fig. 2).

Increase of green plant regeneration efficiency by callus selection in Puccinellia limosa (Schur.) Holmbg.

Three main types of callus have been selected from seeds of salt marsh grass(Puccinellia limosa (Schur.) Holmbg.) subcultured on Murashige and Skoog medium supplemented with 2,4-dichlorophenoxyacetic acid and kinetin. Callus type I differentiated only occasionally. Callus type II produced roots but no shoots under all tested culture conditions. Both green (47 %) and albino plants have been obtained from the embryogenic callus type III. Callus type III was divided into two subtypes (greening and non-greening) according to the presence or absence of green spots. Separated greening embryogenic callus gave up to 87 % green plants, whereas non-greening callus produced only 4 %.

Diallel analysis of barley anther culture response

The genetics of barley microspore development in culture was examined by means of diallel analysis. The frequency of microspore derived green and albino plant production was shown to be under genetic control. This genotypic limitation to microspore development will limit the application of anther culture techniques to barley breeding programmes. However, significant additive genetic effects were detected for the characters measured and indicate that the frequency of green plant regeneration may be improved by the hybridization of suitable parents. Significant reciprocal differences were also detected and indicate that the direction of the cross is important in determining microspore development. An embryogenic route to green plantlet formation was observed in a number of genotypes in the diallel experiment. The implications of these findings for barley improvement and genetics are discussed.Key words: doubled haploids, barley, anther culture, microspore, embryoid.

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.

Somatic Embryogenesis and Plant Regeneration in Tissue Culture of Echinochloa glabrescens Munro ex Hook. F.

Summary Embryogenic calli which were compact, globular and white in appearance were initiated from young inflorescences and young leaf segments of Echinochloa glabrescens cultured on MS nutrient medium supplemented with 2,4-D and BAP. The induction frequency of embryogenic calli was more than 91% in young inflorescences and 41% in leaf segments. The embryogenic nature was maintained after more than seven months of subculture. The embryoids gave rise to plants on MS regeneration medium. High frequency of green plant regeneration was obtained from both explant cultures of E. glabrescens . However in leaf cultures, albino plants were also observed. Normal green plants grew to maturity, flowered and set seeds. Scanning electron microscope studies revealed that the white, compact, globular callus contained embryoids in various stages of development. Typical bipolar embryoids with scutella, coleoptiles and coleorhizae were observed.

Anther Culture of Basmati 370 At IRRI. A. Gamma Ray-Induced Green Plant Regeneration

Anther Culture of Basmati 370 At IRRI. A. Gamma Ray-Induced Green Plant Regeneration

Anther culture in rice: IV. The effect of abscisic acid on plant regeneration

The effect of abscisic acid (ABA) on plant regeneration in anther-derived calli of rice Oryza sativa L. varieties Taipei 177, Taipei 309 and Fujisaka 5 was studied.ABA at concentrations up to 4×10(-6) M stimulated fresh weight increase in Taipei 309 and Fujisaka 5 while higher concentrations effected corresponding weight decreases in the three varieties tested. Relatively high ABA concentrations resulted in decreased callus size and production of more compact and whitish calli. ABA increased the frequency of calli producing green plants in Taipei 309 and the average green plant regeneration in all varieties tested.