Genetic Stability of the Endangered Species Salix lapponum L. Regenerated In Vitro during the Reintroduction Process

Abstract

Salix lapponum L. is a boreal relict species, threatened with extinction in Poland. An 80% decrease in the number of its stands was confirmed in the last half-century, so that to prevent the loss of downy willow, attempts were made to reintroduce this species in natural habitats. Micropropagation was chosen as a first stage of its active conservation. S. lapponum shoots were collected and disinfected with NaOCl, AgNO3, or HgCl2 or with a two-step disinfection with NaOCl and then placed on MS medium with BA 1 mg·dm−3 and IBA 0.1 mg·dm−3. Regenerated shoots were cultivated with addition of BA, KIN, or 2iP, alone or in combination with auxins, to find the highest multiplication rate. Inter-simple sequence repeat (ISSR) analysis and flow cytometric analyses were conducted on in vitro regenerated plants to check their genetic stability. Disinfection was quite difficult and the use of HgCl2 was the most efficient. The highest multiplication rate was obtained in presence of KIN at 0.5 mg·dm−3 + IAA at 0.5 mg·dm−3. The analysis confirmed the genome size stability, which is in agreement with the results obtained by ISSR, revealing no somaclonal variation in plantlets and therefore allowing the use of the obtained plants for reintroduction.