Somatic embryogenesis of Gentiana genus III. Characterization of three-year-old embryogenic suspensions of G. pannonica originated from various seedling explants |
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Authors: | Anna Miku?a Janusz Skierski Jan J Rybczyński |
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Affiliation: | (1) Botanical Garden - Centre for Biological Diversity Conservation of the Polish Academy of Sciences, Prawdziwka 2, 02-973 Warsaw;(2) Drug Institute, Flow Cytometry Laboratory, Chełmska 30/34, 00-725 Warsaw |
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Abstract: | Experiments were carried out with three-year-old embryogenic suspension culture of Gentiana pannonica Scop. The initial explant for the suspension determinated both the embryogenic character and embryo production. Cultures
were initiated by culture of hypocotyl, cotyledon and root explants on MS (Murashige and Skoog 1962) medium supplemented with
1.0 mg·l−1 Kinetin and 0.5 mg·l−1 2,4-D, later transferred and maintained in liquid MS medium with 1.0 mg·l−1 Dicamba, 0.1 mg·l−1 NAA, 2.0 mg·l−1 BAP and 80.0 mg·l−1 AS. Regeneration medium included 0.0–1.0 mg·l−1 GA3+0.0−2.0 mg·l−1 Kin.+0.0−160 mg·l−1 AS.
In these culture conditions, the effect of the explant was found to be the most important factor. The curve of growth, growth
coefficient and % of participation of various size aggregates differed in the studied suspensions. Flow cytometry revealed
various DNA content in nuclei from praembryogenic mass depending on the explant origin. To complete embryogenesis the medium
was changed from liquid to solidified in the presence of the same plant growth regulators combination required. The most embryogenic
culture appeared hypocotyl-derived and it yielded the highest number of somatic embryos. The suspension culture originating
from root proliferated the highest number of embryogenic cell clusters but did not produce embryos for fraction 120–450 μm.
One hundred mg of suspension of the fraction that was larger than 450 μm yielded 309, 175, 123 embryos for the following suspensions:
root-, cotyledon-, hypocotyl-derived, respectively. Almost 50 % of non-deformed fully developed embryos from all studied suspensions
passed conversion into germling stage and finally plants were regenerated. |
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Keywords: | Gentiana pannonica seedling explants cell suspension long term culture somatic embryogenesis germlings flow cytometry |
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