Haploid Culture And Cytology Observation In Lilium

The factors including genotype, culture medium and pretreatment on the development of microspores and genogenesis were studied. The development of microspore and the process of gynogenesis were observed during the culture time. The origin of embryo and the plants of different ploidy levels were also studied in one wild species and six cultivers of Lilium. The main results are as follows:1. Factors affecting viability and development of isolated microspore were studied. The results showed that: the buds size containing uninucleate stage microspores varied with the different genotypes. While microspores were at early and middle uninucleate stage, the anther color was light yellow and the pollen color was pale yellow. Compared with NLN and PG-96M medium, in FHG medium microspores maintained higher vitality rate in all genotypes, and microspores with two symmetrical nuclei have been observed in'Ceb Dazzle'. In addition, the frequency of abnormal meiotic of pollen mother cell was closely related with heterozygosity and microspore viability. For example, Lilium regale and'Tresor'with low abnormal meiotic rate could make viability of microspores high and declining slowly in microspore culture, whereas the variety'Robina'with highly heterozygosity had higher abnormal meiotic rate and fast declined in microspore vitality.2. The factors including genotype, culture medium, the size of flower, hormone concentrations, and culture methods on the genogenesis were studied. The results showed that: compared with CBM and MS medium, high rate of embryo production could be obtained in BDS medium. The genotype of the donor plants was found to be the most important factor for the success of gynogenesis. The highest gynogenic yield was obtained from OT cultivars, while cultivars of Lilium Oriental produced lower embryos. The relationship between gynogenic yield and the size of the flowers was close, the flowers 2-3 mm before anthesis could produce significantly more embryos than small (young) and the ones 2-3 mm after anthesis. Compared to other hormone concentrations, the 2-4,D and KT concentration at 2mg/L, the difference of the high embryo production could be obtained. Ovules, embryos and ovaries were cultured to produce more plant, but it considered that the first two methods were least efficient, only ovaries culture could induce gynogenic haploids.3. The process of embryo development was observed by cleaning-squash technique, we found that usually one embryo sac contained one embryo, some at the micropylar end, some in the middle of the ovule, and some end in the zygote. Embryo sacs with two embryos were also been observed, it be proved that embryos originated from egg cells, synergid, polar nuclei or antipodal cells.4. Ploidy level was estimated for 33 cultivars in Lilium by flow cytometry analysis, the results showed that: plants of diploid, triploid and aneuploid could be obtained from ovaries culture.