New Researches in Biotechnology - Facultatea de Biotehnologii ...

Proceeding of the 4 rd International Symposium“NEW RESEARCH IN BIOTECHNOLOGY” USAMV Bucharest, Romania, 2011IN VITRO CULTURES OF BALKAN AND BULGARIAN ENDEMICCENTAUREA SPECIESROSSEN GORGOROV *,1 , MARINA STANILOVA * , SVETLANA BANCHEVA ** Institute of Biodiversity and Ecosystem Research, Bulgarian Academy of Sciences23, Acad. G. Bonchev Str., 1113 Sofia, Bulgaria1 Corresponding author: E-mail: rossen6540@yahoo.comAbstract. Centaurea caliacrae and C. finazzeri are Balkan endemic species and C. davidovii isendemic for Bulgaria. In this study, in vitro cultures of the three species were initiated from seeds asa means of ex situ conservation. Due to the species’ conservation status, the available plant materialwas limited, especially with C. davidovii. An average of 90% of the seeds of C. caliacrae germinatedon MS nutrient medium in the first 20 days of cultivation regardless of phytohormone treatment(giberellic acid). Eleven MS-based media were used with different combinations and concentrationsof cytokinins and auxins. Green callus formation and occasional organogenesis were the mostfrequent response to plant growth regulators. A few adventitious shoots were formed by cultivatedstems, especially on the medium supplemented with 0,5 g/l benzylaminopurine. On the MS medium, C.caliacrae plants grew well, rooted and were successfully transferred to ex vitro conditions with asurvival rate of 84%. A single C. davidovii seedling on phytohormone-free MS medium formedadventitious stems and a callus-like tissue at the base and originated a clone, suitable for furtherexperiments with uniform plant material.On the medium supplemented with 3,5 mg/l thidiazuron and1,4 mg/l benzylaminopurine, leaf explants formed new leaf clusters from callus which continuedgrowing when transferred to the MS medium.Keywords: knapweed, ex situ conservation, plant growth regulators1. INTRODUCTIONThe genus Centaurea s. l. is represented in the Bulgarian flora by more than 70 species andis among the plant groups whose center of speciation includes the territory of Bulgaria. It isalso the genus richest in endemics in the Bulgarian flora (ca. 30%). In the new edition ofthe Red Data Book of Bulgaria (plants) (2011), 14 species are included (ca. 23%). Some ofthe Centaurea species are rare and are represented by one or few populations. This suggestsmeasures for their conservation to be taken, both in situ and ex situ, especially consideringthe reproductive problems those species encounter. Their seeds have a generally lowgermination rate and are very often damaged by insects before being dispersed. In thisstudy, in vitro culture of three endemic species was initiated as a method for ex situconservation – Centaurea caliacrae Prodán, Centaurea davidovii Urumov and Centaureafinazzeri Adamovič. C. caliacrae is a Balkan endemic, occurring along the Black sea coast.C. davidovii is a Bulgarian endemic, occurring only in Stara planina mountain, particularlyin the Central Balkan national park. Its populations are decreasing in size (Bancheva &Gorgorov 2010). C. finazzeri is also a Balkan endemic, represented in Bulgaria by only onepopulation in the Valley of Struma River (Northern part). In addition to in situ conservationmeasures, such as establishment of protected territories and habitat conservation, ex situmethods are also advisable, including in vitro cultivation and micropropagation of the133