Green Haploid Plants Research Articles

Efficiency of three haplomethods in durum wheat (Triticum turgidum subsp. durum Desf.): isolated microspore culture, gynogenesis and wheat × maize crosses

This study presents the first report comparing the efficiency of microspore culture, gynogenesis and durum wheat × maize crosses for haploid plant production from three durum wheat genotypes (Razzek, Karim and Jneh Khotifa). The results showed that the best induction, calli or embryos formation and plant regeneration rates for the three genotypes were obtained with gynogenesis (47.2, 7.6, 0.8%), followed by interspecific crosses (33.1, 1.7, 0.4%) and isolated microspore culture (8.2, 0.05, 0.01%). Interestingly, all plants regenerated by gynogenesis and durum wheat × maize crosses were green whereas all plants obtained by isolated microspore culture were albino. In the haploid production system, all steps of the process are important for the three methods. The critical steps that have greatly reduced the number of regenerated haploid plants were induction, embryogenesis and regeneration for microspore culture, forming and regeneration of calli or embryo and haploid regeneration for interspecific crosses and gynogenesis. Genotypes with good capacity of induction have not necessarily a good capacity of haploid plantlets regeneration and vice-versa. However, calli or embryos formation seems to be an indicator of the haploid production. Overall, Razzek showed a good ability to produce haploids using the three methods. Each haplomethod showed a specific advantage. Although gynogenesis is the less used method for durum wheat, it has proved to be a successful approach for green haploid plant production.

Regenerasi Tanaman pada Kultur Antera Padi: Pengaruh Persilangan dan Aplikasi Putresin

The efficient use of anther culture in crop improvement depends on the success of plant regeneration techniques and the recovery of sufficient number of either green haploid plants that may be artificially or spontaneously doubled yielding homozygous diploid pure lines in first generation. Putrescine, one of growth regulator polyamines, was known as essential factor in embryogenesis on various plant tissues. The objective of this research was to study the effect of crosses and putrescine application on certain stages of culture most efficient in increasing green plant regeneration. The experiment was arranged factorially. The first factor was reciprocal crosses of Taipei 309 to Asemandi and Asemandi to Taipei 309. The second factor was application of putrescine consisted of control (M0: N6 induction medium and MS regeneration medium); M1: N6 induction medium, MS regeneration medium + 10 -3 M putrescine; M2: N6 induction medium + 10 -3 M putrescine, MS regeneration medium; and M3: N6 induction medium + 10 -3 M putrescine, MS regeneration medium + 10 -3 M putrescine. The results indicated that the addition of 10 -3 M putrescine into callus induction and plant regeneration media was the best in increasing number and percentage of responding anther, callus formation, responding calli, green plants and number of total plants. Rice anther culture efficiency, which was indicated by ratio of green plant to responding calli and percentage of green plant to number of anther innoculated, was also increased by the addition of 10 -3 M putrescine. At that best treatment maternal effect showed in number of calli producing green plantlets, number of green plantlets, and percentage of green plant to number of anther innoculated. F1 from Taipei309/Asemandi was better than F1 from Asemandi/Taipei30. Key words : Rice, anther culture, putrescine, green plant, reciprocal crosses

Regenerasi Tanaman pada Kultur Antera Beberapa Aksesi Padi Indica Toleran Aluminium

<p class="p1">Anther culture provides the quick route in obtaining pure lines in a single generation from either green haploid plant that may be artificially or spontaneously doubled. Indica rice known as recalcitrant genotype because of its difficulty in regenerating sufficient number of green plantlets among the regenerated plants through anther culture. Whilst, research on studying anther culture ability has to be done to assure the success of rice breeding through anther culture. The objective of this research was to determine regeneration ability of five accessions of indica rice tolerance to aluminum through application of putrescine in anther culture. Completely randomized design with 15 replications was used in this research. Treatments consisted of five accessions of aluminum tolerance indica rice, ie. CT6510-24-1-3, Grogol, Hawara Bunar, Krowal, and Sigundil. Callus induction medium based on N6 medium + 10<span class="s1">-3 </span>M putrescine, while regeneration medium based on MS + 10<span class="s1">-3 </span>M putrescine. The results indicated that culture ability is controlled by the genotype. From this research, Grogol, Krowal and Sigundil were selected as accessions having good rice anther culture ability, and therefore can be used as parents for developing new rice varieties tolerance to aluminum through anther culture.</p>

Development of highly tissue culture responsive lines of Lolium temulentum by anther culture

Most of the important forage and turf grasses are outbreeders, require vernalization to flower, and in some cases are polyploid. With the development of gene sequencing information in grasses, there is an urgent need for a model system to perform large-scale functional analysis of candidate genes. We propose to use Lolium temulentum L. (Darnel ryegrass) as a model system for genetic manipulation studies in forage and turf grasses because L. temulentum has the following advantages: self-fertile, short life cycle (11 weeks), diploid, easy to grow, and closely related with other major grass species. In order to improve tissue culture response of L. temulentum, two relatively responsive lines were crossed and putative hybrid seeds obtained. Analysis of the F 1 plants by SSR markers confirmed that the F 1 plants were true hybrids. Anthers were dissected from F 2 plants of the cross, 3.0% of the cultured anthers responded with the formation of calluses. Green haploid plants were recovered from 48.9% of the anther-derived calluses. Seeds were harvested from doubled haploid plants and mature embryos were used as explants for comparing tissue culture responses with other lines. Besides anther culture, F 2 seeds of the cross were subjected to a cycle of selection for callus formation and plant regeneration, and F 4 seeds were obtained from the regenerated plants. Comparison of tissue culture response of different lines revealed that the doubled haploid plants had a much higher frequency of embryogenic callus formation than that of the parental lines and the F 4 seeds. This is the first report on the generation of green haploid and doubled haploid plants in L. temulentum. The technique was successfully used for the rapid production of homozygous diploid plants that are highly tissue culture responsive. The anther culture-derived new L. temulentum lines could be valuable material for functional test of genes in grasses.

Green haploid plants from unpollinated ovary culture in tetraploid wheat (Triticum durum Defs.)

In Triticum durum Defs., gynogenesis was investigated on six genotypes, Cocorit, Isly, Jori, Oued Zenati, Sarif, issued from Morocco,and Cham1 from Syria. The experiments described here were made during two seasons (spring and summer) in which batches of 1,036 and 3,750 unpollinated ovaries were used. A 4 °C pretreatment was applied, during 7, 11 or 15 days and the A (modified Jahne's et al.,1991) and B (modifed San Nœum's, 1976) induction media were compared. The spikes were harvested when microspores were at bi- or trinucleate stage. After the 4 °C cold pretreatment, they were sterilized, and the excised ovaries underwent the following phases and sequence of successive media. The A or B ‘induction’ media, induced cell divisions in the female gametophytes, and after 4 to 9 weeks in dark conditions, the swelling ovaries burst. The growing calluses were excised and transferred to 16 h. light onto a differentiation medium (dif); there generated shoots were then placed on a development (dev), followed by a rooting medium (r). The comparison of the two experiments showed that summer was the best season, for in vitro unpollinated ovary culture; this period corresponds to a growth of mother-plants during spring, while those developing in winter gave rise to lower scores. In Experiment 1 the best level of regenerated plants per 100 ovaries were obtained for Isly with 14.3% and for Oued Zenati with 10.1%. In Experiment 2, the best values were those of Isly with 21.8%, after a 7 day cold pretreatment and use of induction medium B, and of Cocorit with 18.1%. Jori, exhibited good results, in three define situations, with 17.1%, 16%, and 9.3%.Only Cham1 and Sarif showed very low scores or no plants. Jori remaining aside, for each genotype, the best values rose up from a specific treatment combination, as a very strong interaction was expressed between genotype, induction medium and cold pretreatment duration. After this ‘first phase’ of regeneration by ovary culture 150 haploid plants were obtained. When combining these results with other attempts, a total number of 191 green plants was regenerated, 90 for Isly, 74 for Jori, 19 for Cocorit, 7 for Oued Zenati and 1 for Cham1. In a ‘second phase’, permanent regenerating callus lines were obtained through subculture on dif medium. Depending on genotype, 2 to 9 transfers (up to one year for Jori) were performed, giving 507 regenerated plants. From direct regeneration and subcultures, a total of 698 plants were produced, all green and mostly haploid. Fertile doubled haploid plants were obtained, either spontaneously or by colchicine treatment, for all these genotypes. These results are of interest for breeders, who need pure lines for genotypes evaluation and for the creation of improved homozygous varieties.

Durum wheat × maize crosses for haploid wheat production: Influence of parental genotypes and various experimental factors

AbstractTo improve haploid plant production in durum wheat, the haplomethod involving intergeneric crossing with maize followed by embryo rescue was used. The influence of parental genotypes and various experimental factors were studied. Ten cultivars of Triticum turgidum ssp. durum (female parent) were crossed with eight genotypes of Zea mays. After pollination, plant stems were either maintained in situ or cut near the base and kept in a 2,4‐dichlorophenoxyacetic acid (2,4‐D)‐sucrose solution. Ten to 18 days after pollination, embryos were excised from developed ovaries and cultured on one of MS, MS/2, or B5 media. Haploid embryos and plants were obtained (78 green haploid plants regenerated in 0 year). The wheat genotype was significant for ovary development, embryo and plant formation, whereas the maize genotype was significant only for embryo formation. Detailed results of all crosses showed the best crossing partner for each wheat genotype. Cutting the plant stems after pollination gave better results than maintaining them in situ. The optimal stage for embryo rescue was 14 days and B5 and MS/2 media were more efficient than MS for embryo culture.

Production of green haploid durum wheat plants by pollination of wheat with maize

Eight genotypes of Triticum turgidum L. ssp. durum (2n = 4x = 28) were pollinated by three varieties of maize (Zea mays L.) to obtain green haploid durum wheat plants. After pollination, plants were cut and maintained in a nutrient solution until the ovary developed, and the embryo was rescued through in vitro culture. The influence of both the wheat and the maize genotypes on ovary development and embryo and green haploid plant formation was apparent. Ovary development and embryo formation were observed in all wheat genotypes with at least one maize variety. Green plants were obtained from six of the eight cultivars tested. Sixty doubled haploid green plants were recovered after colchicine treatment.Key words: Triticum turgidum ssp. durum, Zea mays, intergeneric hybridization, embryo rescue, haploid green plants.

Analysis of anther culture response in hexaploid triticale

AbstractThe androgenic response of five varieties and 10 F1 intervarietal hybrids of 6 × triticale was analysed. The overall response involved three independent phenomena: the production of embryoids, the capability of embryoids to regenerate plants, and the production of green plants. Each of these factors appeared to be controlled by independent genetic systems. From a practical point of view, the product of these three factors, the number of green haploid plants per anther (GPA), is the most reliable variable for the evaluation of genotypes. Positive heterotic effects on GPA were observed in the hybrids, which may indicate that GPA is controlled predominantly by additive genes. No correlation was found between the number of green haploid plants produced by hybrids and that yielded by the best parental line. Therefore, it is difficult to predict the yield of green haploid plants that can be produced by a hybrid simply from the number produced by its parents.