The main goal of any DNA extraction procedure is to ensure reliable and reproducible results in a simple, fast and inexpensive manner. When it comes to plant tissues, this goal is challenging to achieve due to the presence of a variety of metabolites that interfere with DNA during isolation and downstream analysis. In this study, we compared the efficiency of three methods for DNA extraction from plum kernels: 1) the standard CTAB Soltis method which is the most common protocol for DNA extraction from various plant tissues (seeds, young leafs, mature leafs, root); 2) CTAB-based method originally described for DNA isolation from medicinal plants with high levels of secondary metabolites; 3) and one of various commercially available kits. The usefulness of the obtained DNA was evaluated by SSR analysis with seven microsatellite markers. Although the latter two extraction protocols retrieved genomic DNA that gave positive PCR results, only DNA isolated by kit produced full SSR profile

Satureja subspicata and S. horvatii are endemic species of the Balkan Peninsula and often used in traditional medicine in Bosnia and Herzegovina to treat different health conditions. We aimed to analyze the unevaluated apoptotic, genotoxic and cytotoxic effects of two Satureja species, as well as their content of phenolics that are mainly responsible for the plant's biological activity. Apoptotic and geno/cytotoxic activities of S. subspicata and S. horvatii were investigated in vitro in human lymphocyte culture and in vivo in mice. The content of the main phenolics in plant extracts was determined by ultra-high pressure liquid chromatography-MS-MS (UHPLC–MS/MS). Genotoxic and cytotoxic activities of Satureja extracts were evaluated in vitro by applying a cytokinesis-block micronucleus cytome assay in human lymphocyte culture and in vivo applying a mice reticulocytes micronucleus assay. SALSA RT-MLPA R011-C1 apoptosis assay was used for measuring the relative expression of 44 genes associated with the regulation of the apoptotic pathways in human lymphocyte cultures treated with different concentrations of two Satureja extracts. The first analysis of phenolic compounds in S. horvatii and S. subspicata determined by an UHPLC-MS/MS method revealed high levels of rosmarinic and caffeic acids. Minor genotoxic potential was determined in relation to the tested concentrations while no cytostatic and cytotoxic effects were revealed in vitro. However, when applied in concentrations of 200 mg/kg per os, aqueous extracts of two Satureja species significantly decreased frequency of reticulocytes micronuclei in treated mice against controls. Extracts of S. subspicata and S. horvatii in concentrations of 0.2 mg/mL, regardless of solvent used, downregulated pro-apoptotic and upregulated anti-apoptotic genes, showing anti-apoptotic activity. Our results indicate that the registered anti-genotoxic and anti-apoptotic activity is most likely related to the high level of phenolic acids (particularly rosmarinic and caffeic) in the tested extracts.

The expert reports state that Bosnia and Herzegovina, despite the presence of diverse and valuable natural resources, lacks systematic, coordinated and harmonized pipeline for biomonitoring. Successful solutions to serious problems regarding environmental protection, management and research rely on the efficient use of exhaustive and unfailing information on the nature around us. However, more often than not, transitional and developing countries lack any centralized, nationally funded databases that could be used as dependable source of information in decision making process. University of Sarajevo-Institute for Genetic Engineering and Biotechnology (INGEB) developed the Regional Biodiversity Database – REBIDA with the aim to collate all known biological data on wild and domesticated natural resources of Bosnia and Herzegovina. This internet-based database represents a comprehensive, searchable and open access platform for science community, academia, governmental and non-governmental stakeholders and general public. Besides its scientific value, REBIDA will serve as an educational tool for discovering the diversity and importance of natural resources, with special emphasis on indigenous and endemic flora, fungia and fauna from the Balkans. It is the only such database in the country, consisting of three functionally connected segments: tissue database, DNA database and digital genetic database on plant, animal and human samples. To complement REBIDA, a mobile application called REBIDA SCANNER was also developed. It will be free to download for IOS and Android platforms and will enable professionals, nature enthusiasts and any other interested parties to contribute to REBIDA through data collection, field sampling and documentation of B&H wild life.

The Dinaric endemic plant species Moltkia petraea (Tratt.) Griseb. is often called a "living fossil" of ancient Tertiary flora, with great importance for Bosnia and Herzegovina’s biodiversity. Considering its narrow and limited distribution range, insufficient data on the molecular background of this species is given so far. Due to the presence of various secondary metabolites that interfere with the DNA, isolation of nucleic acids from plant cells is known to be challenging. Even in closely related species it is necessary to optimize DNA isolation protocol in order to obtain high quality PCR amplifiable DNA. We collected 91 samples from five populations in Herzegovina. Doyle and Doyle (1987) CTAB protocol was modified by adding vitamin C (ascorbic acid) to the cell lysis buffer to improve DNA yield and quality. trnL(UAA) intron and nrDNA (ITS1, ITS2) molecular markers were applied to demonstrate amplifiability of isolated DNA and elucidate the intra- and interpopulation genetic diversity. Our results suggest a successful PCR amplification for 81% of the analyzed samples. PCR-RFLP analysis of trnL(UAA) revealed that all individuals in five populations have the same haplotype based on the obtained enzymatic profile for three enzymes (TaqI, HinfI, HindII). Alignment and comparison of ITS sequences didn’t reveal any hypervariable portion that could be informative in elucidating the genetic diversity of M. petraea populations. Further studies with additional application of microsatellite loci, RAPD and AFLP methods are necessary in an attempt to get insights into the genetic diversity of M. petraea.

This study presents genetic data for nine Native American populations from northern North America. Analyses of genetic variation focus on the Pacific Northwest (PNW). Using mitochondrial, Y chromosomal, and autosomal DNA variants, we aimed to more closely address the relationships of geography and language with present genetic diversity among the regional PNW Native American populations. Patterns of genetic diversity exhibited by the three genetic systems were consistent with our hypotheses: genetic variation was more strongly explained by geographic proximity than by linguistic structure. Our findings were corroborated through a variety on analytic approaches, with the unrooted trees for the three genetic systems consistently separating inland from coastal PNW populations. Furthermore, analyses of molecular variance support the trends exhibited by the unrooted trees, with geographic partitioning of PNW populations (FCT = 19.43%, p = 0.010 ± 0.009) accounting for over twice as much of the observed genetic variation as linguistic partitioning of the same populations (FCT = 9.15%, p = 0.193 ± 0.013). These findings demonstrate a consensus with previous PNW population studies examining the relationships of genome-wide variation, mitochondrial haplogroup frequencies, and skeletal morphology with geography and language.

Raspberry cultivars are clonally propagated and therefore all plants belonging to a single cultivar represent the same genotype. Cultivar integrity of raspberry plantlings placed on the market in Bosnia and Herzegovina (B&H) is based on examining of morphological traits, which is not a reliable tool for genetic identification. In this study plantlings declared as cultivar ‘Polana’ were genotyped using seven microsatellites, in order to gain preliminary insight into the genetic integrity of raspberry plantlings marketed in B&H. Plant tissue (leaves) from 10 raspberry plants were randomly sampled from a batch of plantlings sold by major fruit nursery in Bosnia and Herzegovina. Along with these samples, four reference cultivars with confirmed identity (‘Polka’, ‘Autumn Bliss’, ‘Heritage’ and ‘Polana’) were also included in the study. Seven primer pairs amplified 31 alleles, or on average 4.4 alleles per locus. UPGMA cluster analysis, based on the Jaccard similarity coefficient, revealed that among the ten samples declared as ‘Polana’ plantlings only five were genetically identical to any of the other samples. The cluster analyses also exposed that none of the ten samples declared as ‘Polana’ seedlings were in fact identical or even closely related to the ‘Polana’ reference cultivar or any of the other reference cultivars. These findings clearly show that the genetic identity of primocane raspberry plantlings , currently sold in Bosnia and Herzegovina, needs to be tested using objective and reliable methods rather than simple morphologic observation.

UDK: 582.675.1:575(497.6) Helleborus multifidus Vis. is endemic Illyric-Adriatic species with distribution range in Italy, Slovenia, Croatia, Bosnia and Herzegovina, Montenegro and Albania. Although few studies reported different taxonomic categories for H. multifidus, this one is the first using molecular-genetic markers (trnL region and matK of chloroplast DNA and nuclear ITS1 and ITS2 region) for genetic characterization of H. multifidus presented at three localites in Bosnia and Herzegovina. The results revealed that PCR-RFLP on trnL intron was not informative for testing inter- or intrapopulation diversity. Contrary, analysis of matK, ITS1 and ITS2 sequences showed differences between populations from Trebinje region and Kupreško polje, pointing to the need to include additional analyses in order to confirm these findings.

This paper presents the first estimation of polymorphism of the Bosnian-Herzegovinian-Croatian Shepherd dog Tornjak in Bosnia and Herzegovina using 10 microsatellite loci, which are an integral part of StockMarks® for Canine Genotyping Kit (Applied Biosystems, Foster City, CA, USA). Ten microsatellite loci used in this study are appropriate for assessing the genetic diversity for this breed. Measures of genetic diversity were estimated based on allelic and genotypic calculations, observed (HO) and expected (HE) heterozygosities, deviations from Hardy-Weinberg equilibrium and polymorphism information content (PIC). The lowest genetic diversity was estimated for locus PEZ20, and the highest for PEZ6 locus. Observed and expected mean heterozygosities were 0.7261 and 0.7392, respectively. Statistically significant deviation (p<0.05) from Hardy-Weinberg equilibrium was found for PEZ1, PEZ12, PEZ3 and PEZ6 loci. The PIC values suggested that all markers (100%) are very informative (PIC > 0.5) in terms of their suitability for genetic diversity studies. When all observed parameters are taken into account (observed and expected heterozygosities, PIC, number of detective and effective alleles, number of detected and maximum possible genotypes, major allele frequency and major allele frequency index), we can conclude that PEZ6 locus shows the highest genetic diversity while PEZ3 displays the lowest. However, assuming values of observed and expected heterozygosities, as well as PIC, we consider loci PEZ20 to be the least diverse, but this locus has more effective alleles and more genotypes present than PEZ3. These preliminary results are the first genetic diversity survey of the Bosnian-Herzegovinian-Croatian Shepherd dog Tornjak in Bosnia and Herzegovina and could be useful to the dog breeders in designing and managing breeding strategies. Summarizing the information above, we can conclude that the population of the Bosnian-Herzegovinian-Croatian Shepherd dog Tornjak from B&H is not affected by substantial loss of genetic diversity. Results of our study indicate presence of reasonably high level of genetic variability and lead to a better understanding of this dog breed.