Blagajev volčin (Daphne blagayana) je nizka polegla grmovnica iz družine volčinovk (Thymelaeaceae). Zaradi svoje redkosti je praktično v vseh državah, v katerih se pojavlja, zavarovan z nacionalnimi akti. Namen naloge je bil ovrednotiti varstveni status Blagajevega volčina na podlagi različnih molekularnih markerjev. Današnji areal Blagajevega volčina je fragmentiran in obsega Balkanski polotok in južne romunske Karpate. Način nastanka fragmentiranih nahajališč ni znan. Po eni od razlag naj bi Blagajev volčin v interglacialu zasedal strnjen areal in se iz njega širil proti severu in severozahodu, po drugi pa naj bi bila fragmentirana severna nahajališča posledica razkosanja v glacialu in kot taka naj bi se ohranila do danes. Način nastanka izoliranih populacij igra vlogo pri znotrajpopulacijski in medpopulacijski genski variabilnosti, kar posledično vpliva tudi na varstvo vrste. Vzorčenje Blagajevega volčina je potekalo na 21 lokacijah znotraj celotnega areala vrste v letih 2009 in 2010. Kot orodje za oceno varstvenega statusa smo uporabili kloroplastno DNA ter AFLP markerje. V filogeografsko analizo smo vključili 95 rastlin iz vseh 21 populacij. Raziskava je temeljila na analizi petih medgenskih vmesnikov kloroplastne DNA: rpl20-rps12, atpB-rbcL, trnL-F, psbA-trnH ter trnK-matK skupaj z genom matK. Rezultati kažejo na obstoj treh skupin: severne skupine, ki vključuje slovenske in italijanske populacije, južne skupine, ki vključuje populacije iz Makedonije, Črne gore in Stolca (BiH) ter osrednje skupine, ki vključuje ostale vzorčene populacije. Tako haplotipna kot nukleotidna raznovrstnost sta najvišji v osrednji, najnižji pa v severni skupini. V drugem delu raziskave smo s pomočjo dominantnih AFLP markerjev ocenili genetsko strukturo izbranih 13 populacij ter ocenili genski pretok med njimi. Rezultati kažejo na razmeroma močno genetsko diferenciacijo vrste (FST = 0,4) in na odsotnost genskega pretoka med oddaljenimi populacijami. S pomočjo programov STRUCTURE in BAPS smo populacije razvrstili v skupine, ki pa niso popolnoma usklajene s skupinami glede na kloroplastno DNA. Neusklajenost vzorcev, ki jih kažeta kloroplastna DNA in AFLP, lahko pripišemo različnim načinom razmnoževanja posameznih populacij ter različnemu načinu dedovanja kloroplastne in jedrne DNA. Na podlagi obeh markerskih sistemov lahko za Blagajev volčin predlagamo identifikacijo treh evolucijsko pomembnih enot (angl. Evolutionary significant unitsESU) in ene dodatne upravljavske enote (angl. Management unitsMU), na katere bi se moralo osredotočiti varovanje vrste v prihodnosti.Blagay\u27s daphne (Daphne blagayana) is a small decumbent bush from the family Thymelaeaceae. Due to its rarity it is protected by national legislation in almost all countries within its range. The aim of this work was to assess the conservation status of Blagay\u27s daphne (Daphne blagayana) throughout its distribution range, using chloroplast DNA (cpDNA) and AFLP markers. The present distribution of D. blagayana is fragmented and comprises the Balkan Peninsula and the southern Romanian Carpathians. However, the mechanism leading to such fragmented distribution range is not well understood. According to one interpretation, D. blagayana had a smaller distribution range during the interglacial, after which it expanded north and northwest. According to another interpretation, the fragmented areal is the result of fragmentation during the glacial and is still preserved today. The mechanisms leading to isolated populations play an important role in within-and between-population genetic variability, which in turn is relevant to species conservation.Sampling of D. blagayana was conducted at 21 locations across the entire distribution range in 2009 and 2010. cpDNA and AFLP markers were used as a tool for assessing its conservation status. The phylogeographic analysis included 95 plants from 21 populations. The study was based on the analysis of five spacer regions of chloroplast DNA: rpl20-rps12, atp-BrbcL, trnL-F, psbA-trnH and trnK%matK with the gene matK. The results suggest the existence of three clusters: the northern cluster (SLO), including Slovenian and Italian populations, the southern cluster (J), including populations from Macedonia, Montenegro and Stolac (BiH), and the central cluster (SR), including the remaining populations. Both haplotype and nucleotide diversity were highest in the central cluster and lowest in the northern cluster.In the second part of the study we assessed genetic structure and gene flow between 13 selected populations using dominant AFLP markers. The results showed a relatively strong genetic differentation (FST = 0.4) and absence of gene flow among distant populations. Using programs STRUCTURE and BAPS populations were subdivided into 2 and 7 groups, respectively. The groupings proposed by STRUCTURE and BAPS are not fully congruent with cpDNA clusters. This incongruence might be attributed to different reproduction strategies within populations, and to different modes of inheritance of chloroplast and nuclear DNA.Based on both marker systems we can suggest the identification of three evolutionarily significant units (ESU) and one additional management unit (MU), on which conservation efforts should be focused in the future
