Early selection in a breeding program has been performed applying PCR (Polymerase Chain Reaction) paternity tests to micropropagated plants derived from in vitro germinated embryos. Paternity tests allowed us to screen out non-hybrid clones during the initial propagation phase and thus reducing the incidence of accidental pollinations. In this work the efficiency of DNA extraction for paternity tests from foliar tissues of plants grown under different environments and culture media is studied. In vitro plant leaves have been affected by both the environment and the culture media composition, modifying the ratio DW/FW (dry weight/fresh weight) and the DNA extraction performance. The highest FW per cm-2 and the highest DW/FW ratio were found in field plants, however, shaded plants grown in a greenhouse yielded the highest extracted DNA concentration. In vitro plants cultured on BAPcontaining medium showed lower extracted DNA concentration than ex vitro plants, but higher than that of plants cultured on ZEA-containing medium that showed the lowest FW per cm-2. These results are discussed in terms of the optimization of the DNA extraction for PCR analysis and of the effects of both the environment and the cytokinin type used in vitro on plant leaf tissues.