Early checking of genetic stability of cork oak somatic embryos by AFLP analysis

Early checking of the genetic stability of tissue culture-derived plants is necessary to obtain all the potential benefits of clonal forestry. Previous work in Quercus suber L. using random amplified polymorphic DNA (RAPD) markers showed no genetic variation among somatic embryos within embryogenic lines of zygotic origin. Genetic fingerprinting based on amplified fragment length polymorphisms (AFLP) allows the direct analysis of variation at the entire DNA level by generating more reproducible markers than RAPDs. To confirm the absence of genetic variation within six embryogenic lines of zygotic origin, six primer pairs were selected out of 48 combinations of primers for revealing up to 512 AFLP markers, 301 of them (58.8%) being polymorphic. The mean number of markers per genotype was 375. Again differences were recorded among embryogenic lines, even between those that arose from half-sib zygotic embryos, but no variation was observed among somatic embryos within embryogenic lines. To check variation in embryogenic lines raised from nonembryonic tissues, somatic embryogenesis was induced in expanding leaves collected from sprouts originating in three mature cork oak. DNA was extracted from leaves and from somatic embryos derived from each tree, and three primer pairs showed 165, 110, and 108 markers per genotype/tree, respectively. In one tree, AFLP patterns generated from leaves and somatic embryos were identical, but variation was detected in samples from the other two trees. Although the level of genetic variation detected in these lines (5.6% and 7.3% of polymorphism, respectively) is lower than that recorded for half sibs of cork oak (25%-31%), its influence on phenotypic variation needs further assessment.