The Role of Ecotype, Explant and Plant Growth Regulators on Cell Suspension Culture of Ferulago angulata (Schlecht.) Boiss.

The main habitat of Chavil, Ferulago angulata, in Iran is Zagros area. This plant has a rejuvenating effect and is used to treat digestive diseases and intestinal worms. Because the different explants show different amounts of callogenesis under the effect of different growth regulators, selection of an optimal explant and suitable plant growth regulators combination has a significant effect on the production of callus and their suspension culture. There is no reports on Ferulago angulata callogenesis and its cell suspension culture. Therefore, this study was designed and implemented to optimize callus production and cell suspension culture in this important medicinal plant.

Seeds of Chavil were collected from four different habitats in Kohgilooyeh and BoyerAhmad Province in Southwest of Iran include Abenahr, Guayoune, Vezg and Sisakhat. Seedlings obtained from embryo culture were used to prepare the explants. Various explants (leaf, root and stem) were cultured on MS medium supplemented with different concentrations (0, 0.5, 1 and 2 mgl-1) of NAA and BAP. Callus traits were evaluated and from the best culture medium, the best explants and the best PGRs composition for callogenesis of each ecotype were used to cell suspension culture. In order to study the growth rate of cells in suspension culture and plotting the curve of cells growth, two cell density indices and packed cell volume index were evaluated. To determine the cell density index, every 3 days, 10 ml of cell suspension were transferred to the graded falcon and centrifuged at 5000 g for 5 minutes, and the percentage of sediment cells was calculated as the total volume. To determine the packed cell volume index, also every 3 days, 10 ml of culture medium containing cells were transferred to the graded falcon and stored for 30 minutes to precipitate cells and cell masses. Finally, the cell volume was recorded and was calculated as percentage of the total collected medium.

According to the callogenesis percentage, the best ecotype is Abenahr and best explant is leaf explant. The highest level of NAA is 2 mgl-1, and the best level of BAP is 2 mgl-1, which causes 100 callogenesis percentage. The best medium for cell suspension culture is MS medium containing 2 mgl-1 NAA and 0.5 mgl-1 BAP for callus was obtained from leaf explant of Abenahr ecotype. Along with these plant growth regulators, 2,4-D was used in combination with BAP to form suspension culture. The results also showed that 2 mgl-1 2,4-D plus 0.5 mgl-1 BAP were useful in producing suspensions. The difference between 2,4-D +BAP and NAA + BAP combinations more cell volumes were observed, and cell suspension was created at a faster rate and in less time, which is an advantage in research work. Growth rate of cell suspension originated from the leaf explant was higher than root explant. In terms of culturing cell suspension, the Abenahr ecotype was favorable compared to other ecotypes. During cell suppression culture of Cyperus aromaticus by applying different levels of NAA, cell growth was increased up to 3 weeks after application, and then decreased. By applying 2,4-D, cell growth also increased until the third week, and after the third week, cell growth declined, which was very low growth rate compared with the NAA. In cell suspension culture of sugar beet, using 2,4-D was much more effective than NAA on all explants. In the present study, 2,4-D was also more effective than NAA for cell suspension culture of Chavil.

In general, the Abenahr was the best ecotype among of investigated. The explants in both callus culture and the suspension culture, and the best combination of plant growth regulator in both culture was 2 mgl-1 NAA plus 0.5 mgl-1 BAP.