An efficient micropropagation system for Taraxacum pieninicum using seedling explants germinated in vitro is described. Shoot tips and fragments of cotyledons, hypocotyls and roots were isolated from several-day-old seedlings. The highest response, 100% frequency with 12.3 axillary shoots/explant, was from shoot tips on medium supplemented with 0.5 mg L-1 BA and 0.05 mg L-1 NAA. In subsequent subcultures the number of shoots was significantly higher on all explants cultured on medium containing 0.25 and 0.5 mg L-1 BA, and the multiplication rate was highest (20 shoots/explant) in the 4th passage. Shoots rooted on MS and 1/2 MS medium; the highest rooting frequency was 90% and the highest number of roots 2.7/shoot. Rooted plants showed 96.2% survival in sterile soil:sand, and 100% survival in hydroponic culture. Regenerated plants flowered in the second year after acclimatization and yielded viable seeds. This protocol for obtaining complete plants through micropropagation may prove useful for conservation of the genetic resources of this and other endangered species
Using four Polish Vicia faba L. minor cultivars (Bronto, Dino, Tibo, Nadwiślański) we obtained callus from epicotyl fragments collected from 7- and 14-day-old seedlings and from cotyledonary nodes of immature seeds. Callus induction efficiency varied from 81% to 97% depending on the origin of the explant. Shoots regenerated only from the cotyledonary nodes of all tested cultivars. On average, 50% of the explants grown on MS medium containing 1.0 mg dm-3 NAA, 0.5 mg dm-3 BAP, 0.25 mg dm-3 GA3, 1.0 g dm-3 casein hydrolysate, 750 mg dm-3 inositol, 3% sucrose and 0.4% agar were able to regenerate shoots. The number of calluses regenerating shoots was highest from explants collected from fruiting nodes 6 to 9. Decapitation of donor plants increased the percentage of calluses regenerating shoots. On half-strength MS medium with 2 mg dm-3 NAA and on 1/2 MS alone, 11% of the shoots rooted; on 1/2 MS with 1 g dm-3 AC, 8.0% rooted. The regenerants were transferred to Perlite with Hoagland medium and acclimated. Ten percent of the regenerated plants survived the acclimation process, flowered and produced seeds.
In flowering plants, seeds are produced both sexually (double fertilization is required) and asexually via apomixis (meiotic reduction and egg fertilization are omitted). An apomictic-like pattern of endosperm development in planta is followed by fis mutants of sexual Arabidopsis thaliana. In our experiments in planta, autonomous endosperm (AE) developed in met1 mutants. Furthermore we obtained autonomous endosperm formation in vitro not only in unfertilized ovules of fie mutants but also in wild genotypes (Col-0, MET1/MET1, FIE/FIE) and met1 mutants. AE induction and development occurred in all genotypes on the each of the media used and in every trial. The frequency of AE was relatively high (51.2% ovaries) and genotype-dependent. AE induced in vitro represents a more advanced stage of development than AE induced in fie mutants in planta. This was manifested by a high number of nuclei surrounded by cytoplasm and organized in nuclear cytoplasmic domains (NCDs), nodule formation, division into characteristic regions, and cellularization. The high frequency of AE observed in homozygous met1 (met1/met1) mutants probably is due to accumulation of hypomethylation as an effect of the met1 mutation and the in vitro conditions. AE development was most advanced in FIE/fie mutants. We suggest that changes in the methylation of one or several genes in the DNA of Arabidopsis genotypes caused by in vitro conditions resulted in AE induction and/or further AE development.
Blastocystis is a common enteric protozoan of humans and various species of animals. Culture and microscopic examination of fecal samples is the conventional method for identifying four major forms of Blastocystis (vacuolar, granular, non-vacuolar or cystic). In this article, we compared eight liquid media for cultivation of Blastocystis spp. Study material included fecal samples from clinically healthy pigs. Significant differences in the growth of Blastocystis on individual media were observed.