GENETIC-CHARACTERIZATION OF OAK SEEDLINGS, EPICORMIC, CROWN AND MICROPROPAGATED SHOOTS FROM MATURE TREES BY RAPD AND MICROSATELLITE PCR

DNA was isolated from seedlings of Quercus robur, collected from a sin gle provenance, and from epicormic, crown shoots and in vitro shoots f rom a single tree of Q. petraea using a CTAB method of extraction. DNA was obtained in sufficient quantity and purity, from 13 out of 30 see dlings, and from all isolations from epicormic and in vitro shoots (2. 5-10.0 mu g/g fresh/weight). Smearing was minimised at a primer concen tration of 0.12 mu M with Tag polymerase at 0.5 unit/reaction. Nine pr imers produced 142 bands, 28 of which were polymorphic. A similarity i ndex showed that 11 seedlings were closely related with high coefficie nts (0.85-0.90), but each could be identified from another using only 9 primers (OPA-02 and -05, OPG-04 and -05, OPE-01, -02, -03, -08, -09) . DNA was isolated from crown, epicormic and in vitro leaves originati ng from a single 150-yr old tree of Q. petraea and analysed by randoml y amplified polymorphic DNA (RAPD) and microsatellites. With each prim er, a characteristic RAPD pattern was obtained, and it was common to a ll six epicormic shoots derived from different parts of a single branc h of this tree; also to the shoots from the crown of the same tree wit h OPE1 OPA-05, OPA-08, OPA-01, OPA-02, OPA-04, OPA-05, OPG-02, OPG-10, OPE-12. Similarly, the RAPD pattern obtained from shoot cultures in v itro, derived from individual nodes of epicormic shoots produced by si x different branch segments, were uniform for each of 15 primers. This work was repeated using microsatellite PCR. Three microsatellite loci AG16, AG 1/2 and AG 1/5 were amplified by PCR. It showed a uniformity of these microsatellite loci in shoots from the crown of the tree, an d from epicormic shoots cultures derived from six different sections o f branch. (C) 1997 Elsevier Science B.V.