Trinucleotide microsatellites in Norway spruce ( Picea abies): their features and the development of molecular markers.

Trinucleotide microsatellites have proven to be the markers of choice in human genetic analysis because they are easier to genotype than dinucleotides. Their development can be more time-consuming due to their lower abundance in the genome. We isolated trinucleotide microsatellites in Norway spruce ( Picea abies K.) using an enrichment procedure for the genomic-library construction. Here we report on the characterisation of 85 ATC microsatellite-containing clones, from which 39 markers were developed. Many of the clones showed the occurrence of tandem repeats of higher order than the trinucleotide ones, often resembling minisatellite repeats. The sequencing of a sample of the alleles at one of the loci revealed size homoplasy due to base substitutions within the microsatellite region. The presence of ATC motifs within repetitive sequence families was observed. We found a significant relationship between the level of polymorphism and the length of the microsatellite. The levels of variability for ATC trinucleotide markers were lower than those for dinucleotides, both when tested on all loci in a set of six individuals and on a subset of loci in four natural populations. This difference is most likely attributable to lower mutation rates for trinucleotide than for dinucleotide loci. The availability of markers with different mutation rates allows one to select the proper marker set to investigate population processes on different time scales.

Towards second-generation STS (sequence-tagged sites) linkage maps in conifers: a genetic map of Norway spruce (Picea abies K.).

Genetic linkage maps have been produced for a wide range of organisms during the last decade, thanks to the increasing availability of molecular markers. The use of microsatellites (or Simple Sequence Repeats, SSRs) as genetic markers has led to the construction of "second-generation" genetic maps for humans, mouse and other organisms of major importance. We constructed a second-generation single-tree genetic linkage map of Norway spruce (Picea abies K.) using a panel of 72 haploid megagametophytes with a total of 447 segregating bands [366 Amplified Fragment Length Polymorphisms (AFLPs), 20 Selective Amplification of Microsatellite Polymorphic Loci (SAMPLs) and 61 SSRs, each single band being treated initially as a dominant marker]. Four hundred and thirteen markers were mapped in 29 linkage groups (including triplets and doublets) covering a genetic length of 2198.3 cM, which represents 77.4% of the estimated genome length of Picea abies (approximately 2839 cM). The map is still far from coalescing into the expected 12 chromosomal linkage groups of Norway spruce (2n = 2x = 24). A possible explanation for this comes from the observed non-random distribution of markers in the framework map. Thirty-eight SSR marker loci could be mapped onto 19 linkage groups. This set of highly informative Sequence Tagged Sites (STSs) can be used in many aspects of genetic analysis of forest trees, such as marker-assisted selection, QTL mapping, positional cloning, gene flow analysis, mating system analysis and genetic diversity studies.