The grey wolf (Canis lupus) is an iconic large carnivore that has increasingly been recognized as an apex predator with intrinsic value and a keystone species. However, wolves have also long represented a primary source of human-carnivore conflict, which has led to long-term persecution of wolves, resulting in a significant decrease in their numbers, genetic diversity and gene flow between populations. For more effective protection and management of wolf populations in Europe, robust scientific evidence is crucial. This review serves as an analytical summary of the main findings from wolf population genetic studies in Europe, covering major studies from the 'pre-genomic era' and the first insights of the 'genomics era'. We analyse, summarize and discuss findings derived from analyses of three compartments of the mammalian genome with different inheritance modes: maternal (mitochondrial DNA), paternal (Y chromosome) and biparental [autosomal microsatellites and single nucleotide polymorphisms (SNPs)]. To describe large-scale trends and patterns of genetic variation in European wolf populations, we conducted a meta-analysis based on the results of previous microsatellite studies and also included new data, covering all 19 European countries for which wolf genetic information is available: Norway, Sweden, Finland, Estonia, Latvia, Lithuania, Poland, Czech Republic, Slovakia, Germany, Belarus, Russia, Italy, Croatia, Bulgaria, Bosnia and Herzegovina, Greece, Spain and Portugal. We compared different indices of genetic diversity in wolf populations and found a significant spatial trend in heterozygosity across Europe from south-west (lowest genetic diversity) to north-east (highest). The range of spatial autocorrelation calculated on the basis of three characteristics of genetic diversity was 650-850 km, suggesting that the genetic diversity of a given wolf population can be influenced by populations up to 850 km away. As an important outcome of this synthesis, we discuss the most pressing issues threatening wolf populations in Europe, highlight important gaps in current knowledge, suggest solutions to overcome these limitations, and provide recommendations for science-based wolf conservation and management at regional and Europe-wide scales.

Traditionally, simple sequence repeat (SSR)markers have been developed from libraries of genomic DNA. However, the large, repetitive nature of conifer genomes makes development of robust, single-copy SSR markers from genomic DNA difficult. Expressed sequence tags (ESTs), or sequences of messenger RNA, offer the opportunity to exploit single, low-copy, conserved sequence motifs for SSR development. From a 20,275-unigene spruce EST set, we identified 44 candidate EST-SSR markers. Of these, 25 amplified and were polymorphic in white, Sitka, and black spruce; 20 amplified in all 3 spruce species tested; the remaining five amplified in all except one species. In addition, 101 previously described spruce SSRs (mostly developed from genomic DNA), were tested. Of these, 17 amplified across white,Sitka, and black spruce. The 25 EST-SSRs had approximately 9% less heterozygosity than the 17 genomic-derived SSRs (mean H=0.65 vs 0.72), but appeared to have less null alleles, as evidenced by much lower apparent inbreeding (mean F=0.046 vs 0.126). These robust SSRs are of particular use in comparative studies,and as the EST-SSRs are within the expressed portion of the genome, they are more likely to be associated with a particular gene of interest, improving their utility for quantitative trait loci mapping and allowing detection of selective sweeps at specific genes.

In the first large study of conifer expressed sequence tag-simple sequence repeats (EST-SSRs), two large conifer EST databases were characterized for ESTSSRs. One database was from "interior spruce" (white and Engelmann spruce in Southern British Columbia) and Sitka spruce, while the other was from loblolly pine. We found 475 and 629 unique EST-SSRs in loblolly pine and spruce, respectively. 3′ ESTs contained 14% more SSRs than 5′ EST reads in loblolly pine and 41% more in spruce. Conifer EST-SSRs differed conspicuously from angiosperm ESTSSRs in several aspects. EST-SSRs were considerably less frequent in conifers (one EST-SSR every ∼50 kb) than in angiosperms (one EST-SSR every ∼20 kb). Dinucleotide repeats were the most abundant repeat class in conifers, while in angiosperms, trinucleotides were most common. Finally, the AT motif was the dominant motif recovered in both conifer species, whereas AG was the most common dinucleotide repeat in angiosperms. Also, as these ESTSSRs in conifers could be developed into useful genetic markers, our work demonstrates the value of large-scale EST sequencing projects for in-silico approaches for marker development.

Since their discovery in the 1980s microsatellite or simple sequence repeat (SSR) markers have been widely used in many species to generate relatively dense genetic maps or framework maps on which to anchor more abundant, but anonymous, markers such as amplified fragment length polymorphisms (AFLPs). They are typically highly polymorphic, robust, and often portable, particularly among different mapping populations or crosses and often to related species. They have been useful in species where low levels of genetic diversity limit the use of other markers. Cultivated cotton (Gossypium hirsutum L.) has a history of genetic bottlenecks that have considerably reduced its diversity, with the consequence that most molecular marker genetic linkage studies are done with inter-specific crosses. In this study we evaluated the potential for SSR markers to be used in marker-assisted selection (MAS) breeding in cotton by quantifying the level of polymorphism detected with a set of commercially available SSR markers between and within a collection of cotton cultivars being used in our breeding programs. Although the majority of these markers are polymorphic between the 2 tetraploid species of cotton, G. barbadense and G. hirsutum, they are not highly polymorphic (~5%) either among or within G. hirsutum cultivars. However, 6 of the 8 cultivars studied were found to be segregating for alleles of these SSR markers. This suggests that where polymorphisms exist, heterozygosity within cultivars is maintained by the breeding strategies adopted by many modern cotton breeders. Although SSRs clearly have utility in genetic studies using inter-specific crosses or in the introgression of wild germplasm, they will be more difficult to use for standard cotton breeding until greater numbers are available. The utility of some markers may be reduced in some breeding populations where heterozygosity remains in the parental material.