Controlled crosses of seven Scots pine (Pinus sylvestris L.) trees produced 49 families that included both reciprocals and selfings. Embryogenic cultures were initiated from immature megagametophytes and after 6 months in maintenance culture, mature somatic embryos were produced from the surviving 166 lines. The effect of parent genotypes on the cultures was evaluated at initiation of the tissue culture period, after 6 months in maintenance culture and at embryo maturation. The effect of the maternal parent was most pronounced at culture initiation. After 6 months in tissue culture, the maternal effect had decreased and the effects of both parents were significant. By the somatic embryo maturation stage, the maternal effect was still considerable but the paternal effect was no longer detectable. There was little correlation between the ranking of mothers and fathers, indicating that the maternal effect was caused by factors other than the paternal effect. No mother x father interaction was found, indicating that mothers successful at initiation and after 6 months in tissue culture, pollinated by any of the successful fathers, produced somatic lines and mature somatic embryos.

Stilbenes are abundant in the heartwood of Scots pine (Pinus sylvestris L.) and are known to have strong antifungal efficiency. In this study, Scots pine sapwood blocks were impregnated with crude heartwood extract containing the stilbenes pinosylvin (PS) and the pinosylvin monomethyl ether (PSM). Impregnated blocks were submitted to brown-rot fungi, Coniophora puteana, Gloeophyllum trabeum, and Rhodonia (Poria) placenta, and fungal growth test and decay test were performed. Both tests showed that the impregnation with a high concentration of stilbenes (60 mg g-1 dry wood) significantly suppressed the growth of fungi and slowed down the decay process of wood blocks, especially in case of G. trabeum. However, chemical analysis showed that PS and PSM were degraded by all the three types of fungi, obviously via modification to resveratrol and methylresveratrol. Rhodonia placenta displayed the fastest rate of degradation. Thus, impregnation with biodegradable stilbenes could be a viable alternative for wood preservation only in service situations with low or transient risk of decay.

The involvement of the plant growth regulator ethylene in somatic embryo maturation of Pinus sylvestris (L.) was investigated. Genes that encoded 1-aminocyclopropane-1-carboxylate synthase (ACS), the rate-limiting enzyme in the ethylene biosynthesis pathway, were isolated and characterized. Two novel complementary DNAs (cDNAs) of PsACS1 and PsACS2 that encode ACS were isolated from embryogenic cultures (ECs) along with their polymerase chain reaction (PCR) products. The two sequences shared 69% similarity at amino acid level. PsACS1 was expressed in ECs at the proliferation and maturation stages, whereas the PsACS2 transcript was expressed in embryos only at the maturation stage. These expression levels varied among the five P. sylvestris genotypes studied. Moreover, the high levels of PsACS2 transcript abundance correlated with higher numbers of embryos at stage 3 of development. Ethylene production reached the highest levels at stage 3, and subsequently dropped. Hence, expression of PsACS2 corresponded to ethylene production during embryo development and could thus serve as a genetic marker for the early maturation stage. Both gene expression and ethylene production varied among the different genotypes. Investigating expression of ACC synthase genes further enhanced our understanding of the role of ethylene in embryo development and maturation and improved the efficiency of somatic embryogenesis of P. sylvestris.

Heterogenous chitinases have been introduced in many plant species with the aim to increase the resistance of plants to fungal diseases. We studied the effects of the heterologous expression of sugar beet chitinase IV on the intensity of ectomycorrhizal (ECM) colonization and the structure of fungal communities in the field trial of 15 transgenic and 8 wild-type silver birch (Betula pendula Roth) genotypes. Fungal sequences were separated in denaturing gradient gel electrophoresis and identified by sequencing the ITS1 region to reveal the operational taxonomic units. ECM colonization was less intense in 7 out of 15 transgenic lines than in the corresponding non-transgenic control plants, but the slight decrease in overall ECM colonization in transgenic lines could not be related to sugar beet chitinase IV expression or total endochitinase activity. One transgenic line showing fairly weak sugar beet chitinase IV expression without significantly increased total endochitinase activity differed significantly from the non-transgenic controls in the structure of fungal community. Five sequences belonging to three different fungal genera (Hebeloma, Inocybe, Laccaria) were indicative of wild-type genotypes, and one sequence (Lactarius) indicated one transgenic line. In cluster analysis, the non-transgenic control grouped together with the transgenic lines indicating that genotype was a more important factor determining the structure of fungal communities than the transgenic status of the plants. With the tested birch lines, no clear evidence for the effect of the heterologous expression of sugar beet chitinase IV on ECM colonization or the structure of fungal community was found.

Tricholoma matsutake is an ectomycorrhizal fungus that forms commercially important mushrooms in coniferous forests. In this study, we explored the ability of T. matsutake to form mycorrhizae with Pinus sylvestris by inoculating emblings produced through somatic embryogenesis (SE) in an aseptic culture system. Two months after inoculation, clones with less phenolic compounds in the tissue culture phase formed mycorrhizae with T. matsutake, while clones containing more phenols did not. Effects of inoculation on embling growth varied among clones; two of the four tested showed a significant increase in biomass and two had a significant increase in root density. In addition, results suggest that clones forming well-developed mycorrhizae absorbed more Al, Fe, Na, P, and Zn after 8 weeks of inoculation. This study illustrates the value of SE materials in experimental work concerning T. matsutake as well as the role played by phenolic compounds in host plant response to infection by mycorrhizal fungi.