Hristova, Stoyanov, Gevezova & Denev 2011. Application of ISSR methods in studying Broomrape’s (Orobanchaceae) biodiversity in Bulgaria

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Hristova E., Stoyanov K., Gevezova M. & Denev I. 2011.
Application of ISSR methods in studying Broomrape's (Orobanchaceae) biodiversity in Bulgaria
Biotechnology & Biotechnological Equipment 25(1): 2248-2253

Abstract:
In Bulgaria the holoparasitic family of broomrapes (Orobanchaceae) is represented by 25 species from two genera. The local diversity of the broomrapes in Bulgaria is still grossly under-studied using modern approaches. This not only leads to unclear status of taxonomically problematic species and their local intraspecific taxa, but also hampers the assessment of their relevance as actual or potential treats for agricultural. The aim of this study was to select ISSR primers that amplify polymorphic microsatellite loci suitable both for taxonomic and population studies. One hundred ISSR primers (University of British Columbia Nucleic Acid-Protein Service Unit, UBC Primer Set #9) were tested. Specimens from five different species (Phelipanche ramosa, P. mutelii, P. purpurea, Orobanche alba and O. minor), collected from different locations were used to isolate genomic DNA. The PCR reactions were carried out in a TC-512 THERMAL CYCLER (Techne). The amplified unambiguous bands were scored to compile a presence/absence matrix. Cluster analyses were performed with PAST software. We found that 13 ISSR primers produce polymorphic bands suitable to distinguish the known sections and genera. Other 3 primers can distinguish the genera and probably higher taxonomy ranks. The obtained results open a good opportunity to study broomrape’s biodiversity in Bulgaria.

Acknowledgments
The study was supported by Bulgarian National Science fund grants IFS-B-606 module 1, grant DO-02-204, grant DTK-02/40 and SYNTHESYS projects AT-TAF-3133 and ES-TAF-4407 (European Community – Research Infrastructure Action under the FP6 "Structuring the European Research Area” Program).


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