Seven ectomycorrhizae on oak (Quercus robur L.) from a Slovenian forest stand are described, five of them for the first time in detail. Four of these species could be identified (Lactarius chrysorrheus (Fr.) Fr., L. serifluus DC. ex Fr., Xerocomus subtomentosus (L. ex Fr.) Quel., X. armeniacus (Quel.) Quel., one unidentified species was given a preliminary name ("Quercirhiza squamosa"). Two identified species, already comprehensively characterized on other tree species, are presented shortly (Laccaria amethystina (Bolt ex Hooper) Murr., Cenococcum geophilum Fr.). Whereas some of the encountered species show considerable host specifity with oak (Lactarius chrysorrheus, L. serifluus and, in a lesser degree Xerocomus armeniacus), other species have a broader host range and can even grow with conifers (Xerocomus subtomentosus, Laccaria amethystina, Cenococcum geophilum). A key for all species was established. A possible use of ectomycorrhizal species as bioindicators is discussed.

In the paper authors review the complexity of the impacts on the forest ecosystem caused by road construction and logging operations and discuss research activities in that field. Interdiciplinary approach is emphasized and research priorities are suggested. The studies are needed where structural parameters of the soils are related to the plant vitality. This will help us understand the forests response on the impact and enable us to develop the practical techniques for soil and stand protection during further actions.

We have been isolating AC/GT and AG/CT SSRs from the Norway spruce (Picea abies K.) nuclear genome. We isolated several hundreds positive clones from a small-insert genomic library and following sequence analysis we designed primers for 36 of them, 24 containing AG and 12 AC SSRs. After testing them on a panel of spruce individuals 25 of the primer pairs producted a single-locus hypervariable pattern, with the remaining ones giving either a single monomorphic product (18) or very poor amplification (19) or amplification of multiple bands (38). Segregation in accordance with a simple Mendelian model of inheritance was demonstrated for all the loci amplified with the primer pairs giving a simple variable pattern. We screened a panel of 19 spruce trees at these loci. The average number of alleles per locus was 14 and expected heterozygosity 0.80, with up to 23 alleles per locus and heterozygosities exceeding 0.94. This shows that nuclear SSRs can be very useful markers in the population genetics of trees even though the overall efficiency of the marker identification process is quite low due to the high percentage of primer pairs producting complex or "dirty" patterns. We attribute this phenomenon to the high complexity of the spruce genome. Other methods, including the construction of libraries highly enriched for SSR sequences, that we developed in order to make SSR retrival and typing easier and faster will be discussed. We recently extended the use of PCR amplified SSR markers to the chloroplast genome. We demonstrated that mononucleotide poly(A/T) stretches are frequent in the chloroplast genomes of plants and show high levels of between and within population variation, making them ideal tools for cytoplasmic population genetic overcoming the difficulties in finding within species variation that are frequently encountered when analysing the cpDNA molecule by RFLPs or PCR-RFLPs. We will present results of the analysis of mediterranean pine species populations by using a set of cpSSRs that are distributed over the whole cpDNA molecule and discuss the possible applications of such markers for studying gene flow and for paternity analysis.

Needles of 20 years old spruces from Prednji vrh, in the area affected by the Sostanj power station, and from Pokljuka (Alps region, with a relatively unpolluted environment), were analysed for total sulphur content (St). Measurements were taken in a period between April and August twice monthly and in the period from August to April every four weeks (May 1993 - June 1994). Sulphur content in one and two years old spruce needles from Prednji vrh were much higher (one year old needles 2.06-2.50 mg/g needle dry weight, 1993) than in needle samples from Pokljuka (one year old needles 1.05-1.25 mg/g needle dry weight, 1993).

The haploid chromosome set of Norway spruce consists of 12 chromosomes. Most of the chromosomes are very similar in their morphology (length, arm ratio, secondary constriction). For the CGBI it is necessery to recognize all chromosomes. Therefore we used different banding methods to solve this problem. We used the root meristem and the dividing cells of the macrogametophyte as test objects. We discussed morphology of chromosomes of these two different dividing tissues. The use of image analysis equipment made it possible to get in a relatively short time a great number of data for statistical analysis. Another method was, to use an expert-system for data analysis. Finally 3-D models of interphase nuclei were reconstructed.

Three principal approaches are used in our laboratory to analyze plant genomes: (i) The construction of high density molecular maps: We have established a refined microdissection procedure to construct chromosomal and subchromosomal libraries. Synchronized meristematic root tip tissue from which metaphase spreads can be prepared with a novel dropspread technique in combination with nl-scale PCR allowed the cloning of DNA of 10 or less chromosomes or chromosome segments. The construction of high-resolution maps from discrete genome regions can greatly facilitate genetic fingerprinting, gene isolation and QTL studies. (ii) Synteny analysis: The Aegilops-based deletion mapping system in wheat with an array of deleted chromosome parts and chromosome-specific RFLP markers has lead to the construction of a high density physical consensus map of wheat. The integration of wheat, barley and oat RFLP markers proves the colinearity between the wheat A-, B- and D-genomes, the H- genome of barley and the E-genome of Agropyron. (iii) Gene mapping in situ and chromosomes at high resolution: For the sensitivity enhancement of fluorescence in situ hybridization (FISH), the efficient preparation of plant chromosomes for high resolution scanning electron microscopy, mapping of low-copy sequences, and for comparative in situ hybridization a modified drop technique for plant protoplasts was developed. A tandemly amplified repetitive sequence element from microdissected barley chromosomes has enabled the karyotyping of single Gramineae genomes in a single step. These sequences are also useful for simultaneous double or triple hybridization experiments in an attempt to localize new sequence on specific chromosomes or chromosome segments. The physical mapping of the Sec-1 locus has been refined on the satellite of chromosome 1R of rye, and the syntenic locus on barley chromosome 1H identified. A method was developed for in situ hybrization and signal detection at high resolution using a field emission scanning electron microscope and a backscattered electron detector. Colloidal gold particles were localized on chromosome structures resembling the 30 mm fibre. Comparatively, an rDNA probe was located in the secondary constriction and highly compact adjacent regions of barley chromosomes.

Cytogenetical and biochemical investigations of common oak (Quercus robur L.) were applied in order to find suitable markers in bioindication of oak populations. In cytogenetical analyses the chromosome aberration and mitotic index were recorded. Chromosome banding, B-chromosomes, location and number of nuclear organizer regions were indicated using Giemsa C-banding method, fluorescence staining and silver impregnation. In biochemical analyses SDS-electrophoresis of soluble proteins and isoelectric focusing of isoperoxidases were used.

It was our aim to investigate whether the cytokinin status of Norway spruce seedlings (Picea abies L. Karst.) was changed when the plants were grown under nutritive stress. Cytokinins in shoots and roots from spruce seedlings, grown in various hydrocultural media with complete (control) or poor nutrient supply (stress), were determined by an indirect competitive enzyme-liked immunosorbent assay. The shoots of spruce seedlings grown in a poor acidic culture medium, to which aluminium ions (AlCl3, 0.8 mM) were added, showed up to 5-fold higher concentrations of zeatin riboside and isopentenyladenosine (iPa). When adding naphtylacetic acid to the nutrient medium also higher levels of cytokinin ribosides were measured in the shoots. In the roots however cytokinin riboside levels did not increase significantly under Al- or auxin treatment. Feeding experiments using tritiated iPA have shown that the metabolism of this cytokinin is strongly retarded in the roots of stressed seedlings. The experiments show that acid conditions and nutrient shortage together with aluminium ions in the rhizosphere increase the number of lateral roots in Norway spruce seedlings. It is assumed that the increased number of sites of cytokinin biosynthesis and the reduced metabolism of iPA result in the higher content of cytokinin ribosides in the shoots of seedlings.

With the development of molecular marker technologies to construct detailed molecular maps of crop species it has become possible to study much more precisely than before the possible roles of ABA by analysing the distribution of quantitative trait lici (QTL). This has been done with mapping populations of maize and wheat to study the importance of endogenous ABA in determining stomatal function, water-use efficiency and root growth in plants growing under water-stressed conditions. An F2 population of maize was mapped with 100 RFLP markers and F2 plants and their F3 progenies sampled for leaf and xylem ABA contents and stomatal conductance. Correlations using genotype mean data at QTL significant for ABA content showed that stomatal conductance was more likely to be regulated by xylem ABA content than leaf ABA content. The same maize F3 population was used to identify QTL for root growth was positively associated with root ABA content. Water-use efficiency, measured as 13C discrimination, was measured in a mapping population of doubled haploid lines of wheat. Leaf ABA concentrations were also measured. Comparison of QTL for the two traits showed that leaf ABA concentrations were unlikely to have had an effect on plant water-use efficiency in those field conditions. QTL analysis is a very powerful technique for testing the likely roles of endogenous ABA.

This article deals with use of several bioindication methods in forest decline studies within an international TEMPUS project carried out in Slovenia. The aim of the project was to introduce and connect different bioindication methods in order to improve methods and qualify the research staff working in this research field in Slovenia. The following bioindication methods were introduced or adapted for forest decline research in Slovenia: biochemical and physiological markers of air pollution / stress impact on forest trees (Graz Univ.: photosynthetic pigments, thiols, ascorbic acid, protective enzyme systems; Cambridge Univ.: plant hormones; Munich Univ.: mineral nutrition of forest trees); epiphytic lichens as indicators of air quality on forest sites (Graz Univ., Ljubljana Univ); determination of types of ectomycorrhizae as indicators of forest soil condition (Munich Univ., Ljubljana Univ.). The significance and practical use of bioindication methods in environmental studies is also discussed.