Božena Vooková

Some features of somatic embryo maturation of algerian fir

SummaryMaturation of Abies numidica De Lann. somatic embryos was tested on media with abseisic acid and various maltose and polyethylene glycol-4000 concentrations. The effect of basal medium and subculture period on maturation was also examined. The maturation of somatic embryos was promoted by polyethylene glycol-4000, at 7.5–10%. Three to 6% maltose significantly enhanced the yield of mature embryos. The most effective somatic embryo maturation occurred when embryogenictissue was transferred to maturation medium after 14–21 d cultivation on proliferation medium. The ability for A. numidica cultures to form cotyledonary somatic embryos was assessed over a period of 3 yr. Plantlets germinated on half-strength SH (Schenk and Hildebrandt) medium with charcoal and survived transfer to the soil.

Somatic Embryogenesis in Abies Alba × Abies Alba and Abies Alba × Abies Nordmanniana Hybrids

Maturation and germination of somatic embryos of hybrids A. alba × A. alba and A. alba × A. nordmanniana were followed by protein analysis of single embryogenic -suspensor masses (ESM) and analysis of storage protein accumulation during somatic embryo development. Very important step was one week pre-cultivation of ESM on medium with polyethylene glycol (PEG) and abscisic acid (ABA). Low osmotic potential of maturation medium and addition of ABA supported development of somatic embryo. Also partial drying of somatic embryo during following three weeks was needed for its normal development. In spite of morphologically fully developed, the somatic embryos were not physiologically ready for germination at least in terms of storage protein accumulation.

Storage protein dynamics in zygotic and somatic embryos of white fir

Composition and accumulation patterns of storage proteins in female gametophyte and embryos of the white fir (Abies concolor) were investigated during embryogenesis and germination of mature seeds using SDS-PAGE and immunological approach. Altogether 9 major and minor protein components with molecular masses of 14, 16, 22, 24, 27, 30, 35, 38, and 43 kDa were detected in female gametophytes and 9 protein bands in the embryos with the molecular sizes of 14, 16, 22, 24, 25, 27, 34, 38, and 43 kDa. The species seems to deviate in this respect from other representatives of Pinaceae. A conspicuous increase of storage protein synthesis was observed at the stage of fully cellularized female gametophytes and at the cotyledonary stage of embryo development. There exists a high degree of similarity between storage protein profiles of white fir zygotic and somatic embryos. Successive stages of somatic embryogenesis exhibited a high degree of similarity of storage proteins except for cotyledonary stage when a noticeable increase in storage protein synthesis was registered. Conversely, during germination of somatic embryos, an overwhelming majority of storage proteins was depleted.

Biochemical differences between normal callus and embryogenic suspensor mass of silver fir

Normal callus and embryogenic suspensor mass (ESM) were induced from the same immature embryo of Abies alba Mill. These two tissues were found to differ in their isoenzyme patterns of peroxidase, glutamate dehydrogenaseand non-specific esterase and in their requirement for myo-inositol inculture medium.

Improved plantlet regeneration from open-pollinated families of Abies alba trees of Dobroč primeval forest and adjoing managed stand via somatic embryogenesis.

Possibility to improve plantlet regeneration from Abies alba Mill. open-pollinated families of 4 trees in Dobroč primeval and 3 trees in managed forest was studied. Immature zygotic embryos were cultured in order to obtain initiation of embryogenic tissue. Totally, three from the families of the managed forest (57%) and two from the primeval families (50%) responded to initiation condition. Initiation frequencies among families ranged in managed forest: 4.5–56.2%, primeval: 5.4–16.8%. Maturation ability was shown by 77.3% of the primeval cell lines, 36.4% cell lines produced cotyledonary somatic embryos. In managed forest, in 62.5% of the cell lines embryo maturation was observed. Cotyledonary embryos developed only in 15% of cell lines. Regenerants were obtained from 9 cell lines of primeval and from 6 cell lines of managed forest. Biochemically, the mature somatic embryos were characterized by the variation in soluble and protein profiles. The corresponding profiles of insoluble proteins exhibited uniform pattern. The variation was characteristic for somatic embryos of individual cell lines rather than for the primeval and managed stands. Enzymatically, no indications were obtained supporting higher metabolic potential of somatic embryos derived from zygotic embryos of silver fir primeval stand than in somatic embryos originating from the trees of managed stand.

Secondary Somatic Embryogenesis in Abies numidica

The induction of secondary somatic embryogenesis in Abies numidica De Lann. was achieved. Precotyledonary, cotyledonary, and desiccated cotyledonary embryos were used as explants. Cotyledonary embryos before desiccation were the most suitable. The most beneficial was induction medium Schenk and Hildebrandt (SH) with 1 mg dm−3 thidiazuron and 1000 mg dm−3 myo-inositol. Initiation frequency was from 1 to 34 %. Maturation of somatic embryos was achieved on modified Murashige and Skoog medium supplemented with 40 g dm−3 maltose, 100 g dm−3 polyethylene glycol-4000 and 10 mg dm−3 abscisic acid. Mature somatic embryos after three weeks of desiccation germinated on SH medium with 10 g dm−3 charcoal and 10 g dm−3 sucrose.

Protocols for Somatic Embryogenesis of Hybrid Firs

Silver fir (Abies alba Mill.) is an important forest tree species in Slovakian dendroflora. In recent decades a significant silver fir dieback has been observed, resulting decrease of silver fir proportion in the forest of Slovakia from 9% in 1953 to less than 5% at present (Jasik et al., 1999). Views concerning the nature of this phenomenon are in principle reducible to two theories, one of which emphasizes the ecological aspect of fir forest death, while the other interprets this fact genetically (Kormu ak, 1986). Unfavorable ecological conditions (acid rains, accumulation of toxic metals in plant body) cause physiological weakening of organism which is accompanied by attack of parasitic or saprophytic bacteria and as a consequence desiccation of crowns may appear in trees. The genetic background of silver fir decline is caused by high degree of genetic uniformity has been suggested by Larsen (1989). Artificial hybridization of silver fir with allochthonous representatives of genus Abies is one of the promising and effective ways of enriching fir gene pool and hence increasing of its resistance and growth potential (Kormu ak, 1986; Greguss, 1984; Greguss and Paule, 1989). The hybridisation programme for increasing the resistance and growth potential of Abies alba is preferentially oriented at A. nordmanniana, A. cephalonica, A. numidica (Kormu ak, 1985; Kormu ak, 1986) In recent years the hybridization experiments on Slovakian Abies alba with different introduced species resulted in obtaining of hybrid embryos. We have attempted to multiply this hybrid material through somatic embryogenesis (Gajdosova et al., 1995; Salajova et al., 1996; Vookova et al., 1998; Salajova and Salaj, 2001; Vookova and Kormu ak, 2003). The aim of this chapter is to give protocols on initiation, maturation and regeneration of hybrid fir embryos obtained in hybridization experiments.

In vitro growth response of wood‐rotting fungi to IAA

Mycelial growth of some wood‐rotting fungi was studied on a solid modified medium MS (Murashige and Skoog, 1962) with indole‐3‐acetic acid at concentrations of 10‐6 to 10‐3 M. The IAA concentrations of 10‐6 M and 10‐5 M inhibited mycelial growth of the fungus Phaeolus schweinitzii, Laetiporus sulphureus and Pleurotus ostreatus while the same concentrations stimulated mycelial growth of the fungus Stereum rugosum. The IAA concentrations of 10‐6 M stimulated mycelial growth in Piptoporus betulinus and temporarily stimulated mycelial growth in Heterobasidion annosum. The IAA concentration of 10‐4 M appeared critical for wood‐rotting fungi. The IAA concentration of 10‐3 M inhibited mycelial growth in all the fungi under study.

BIOCHEMICAL VARIATION BETWEEN NON-EMBRYOGENIC AND EMBRYOGENIC CALLI OF SILVER FIR

Comparative study on SDS-protein profiles and isoenzyme composition of non-embryogenic and embryogenic calli in two callus lines of silver fir (Abies alba Mill.) revealed the presence of abundant polypeptide fractions and an increased number of isoperoxidases in non-embryogenic calli. Non-specific esterase, on the other hand, exhibited an opposite tendency, while glutamate dehydrogenase was the only enzyme system consisting uniformly of one isoenzyme band in both types of calli investigated.

In vitro study of defense reactions of pine embryo and megagametophyte

Defense reactions of embryo and megagametophyte (endosperm) of European black pine (Pinus nigra Arn.) were studied by in vitro technique. As a tester, basidiomycete Phaeolus schweinitzii (Fr.) Pat. was used. Both, defense reaction of embryo and very strong defense reaction of megagametophyte were found. Some substances which may be involved in defense reactions are discussed.