Assessments of somaclonal variation in micropropagated shoots of Cedrus: consequences of axillary bud breaking

Abstract

Reversed-phase HPLC analysis and random amplified polymorphic DNA (RAPD) markers were used to monitor DNA methylation status and genetic stability of C. atlantica and C. libani shoots generated through axillary bud proliferation. Average DNA methylation in C. atlantica or C. libani seedlings and mature 200-year-old trees of C. libani was 19.8, 19.5 and 22.3%, respectively. These global amounts showed no significant variation after the in vitro establishment of seedling-originated shoot stocks. In contrast, in vitro culture caused a significant decrease in the amount of 5-methylcytosine in genomic DNA of the tissue culture (TC) progenies of one of the adult C. libani genotypes. This DNA demethylation event accompanied an enhancement of the regrowth capacity of this genotype. Detected RAPD variation between mother plants and their TC progenies was species-related, with C. libani TC progenies being genetically more stable than those of C. atlantica. Nevertheless, similarity indices ranged from 0.97 to 1 among mother plants and their TC progenies. Furthermore, the analyses of molecular variance (AMOVA) suggest that RAPD variation among the mother plants and their TC progenies might be considered as not significant. The application of various statistical approaches, including clusterbased genetic distance methods and AMOVA, demonstrates that RAPD markers discriminate C. atlantica and C. libani appropriately.

ASSESSMENT OF GENETIC STABILITY OF MICROPROPAGATED Eucalyptus globulus Labill HYBRID CLONES BY MEANS OF FLOW CYTOMETRY AND MICROSATELLITES MARKERS

“…The findings agree with the fact that the micropropagation through axillary bud proliferation is one of the safest methods for the production of genetically identical plants to the mother plant. There are several works in literature which obtained similar results (Rani and Raina, 1998;Martins et al, 2004;Renau-Morata et al, 2005;Joshi and Dhawan, 2007;Borchetia et al, 2009;Brito et al, 2010;Nayak et al, 2011;Leva and Petruccelli, 2012).…”

Assessment of genetic stability of two micropropagated wild olive species using flow cytometry and microsatellite markers

“…The feasibility of these molecular markers, as well as RAPDs, RFLPs and AFLPs, to assess true-to-type micropropagation has long been certified, as endorsed by the increasing number of papers that have been published in recent years in several species. For instance, the absence of somaclonal variation in plant material micropropagated by stem cuttings was reported for the woody species such as Cedrus libani (Renau-Morata et al 2005) and Robinia pseudocacia (Bindiya and Kanwar 2003) using RAPDs. Also Fernandes et al (2008), using AFLP and SSRs in Quercus suber described the absence of molecular changes in embryogenic cultures.…”

DNA methylation changes in triticale due to in vitro culture plant regeneration and consecutive reproduction

“…The limitation of the system to restriction sites recognized by the endonucleases is apparent (Schrey et al 2013). Contrary to marker-based approaches, high performance liquid chromatography (HPLC) has been applied in studies of DNA methylation of plants regenerated via tissue culture (Renau-Morata et al 2005;Rival et al 2013). This technique supports general information on global DNA methylation and could be applied for the evaluation of differences among plant materials that could be related to spontaneously arising off-type plants (Johannes et al 2009;Yi et al 2010).…”