Assessment of ploidy stability of the somatic embryogenesis process in Quercus suber L. using flow cytometry

Loureiro¹, G. Pinto¹, T. Lopes¹, J. Dolezel² and C. Santos¹

¹Center for Cell Biology, Department of Biology, University of Aveiro, 3810-193 Aveiro, Portugal

²CECAV – Departamento de Ciências Veterinárias, University of Trás-os-Montes e Alto Douro, Vila Real, Portugal

 

Abstract: Flow cytometry analyses were used to verify the ploidy stability of Quercus suber L. somatic embryogenesis process. Leaf explants of two adult cork oak trees (QsG0 and QsG5) of the North of Portugal were inoculated on MS medium with 2,4-D and zeatin. After 3 months, calluses with embryogenic structures were isolated and transferred to fresh MS medium without growth regulators and somatic embryo evolution was followed. Morphologically normal somatic embryos (with two cotyledons) and abnormal somatic embryos (with one or three cotyledons) were used in this assay. Flow cytometry combined with propidium iodide staining was employed to estimate DNA ploidy levels and nuclear DNA content of somatic embryos and leaves from mother plants. No significant differences (P<0.05) were detected among embryos, and between the embryos and the mother plants. Also, after conversion of these embryos, no significant morphological differences were observed among the somatic embryo-derived plants. These results and further studies using converted plantlet leaves and embryogenic callus tissue indicate that embryo cultures and converted plantlets were stable with regard to ploidy level. As no major somaclonal variation was detected our primary goal of "true-to-type" propagation of cork oak using somatic embryogenesis was assured at this level. The estimation of the 2C nuclear DNA content for this species is similar to the previously obtained value.

 

Keywords:  Flow cytometry - Ploidy stability - Nuclear DNA content - Ploidy changes  -  Quercus - Somatic embryogenesis

 

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