2,4-D induction of somaclonal variations in in vitro grown date palm (Phoenix dactylifera L. cv Barhee)

The present study is a part of a program designed at improving the date palm, Phoenix dactylifera L. cv. Barhee, through induced somaclonal variation. In this work, caulogenic cultures were subcultured on MS media supplemented with 0, 1, 5, 10, 20 and 40 mg L−1 2,4-D in order to induce genetic and epigenetic variations. The highest doses of 2,4-D were found to induce severe negative effects on in vitro cultures, although some tissues were able to survive and to produce calli with high morphogenetic capacities. Our analysis showed some significant effect of 2,4-D on several physiological parameters. Indeed, chlorophyll and growth rates were found to drastically decrease while proline content increased from 535 to 2973 nmol g−1 FW when 40 mg L−1 2,4-D were used. In vitro cultures showed several signs of oxidative stress, such as high levels of hydrogen peroxide and malondialdehyde; likewise, the specific activity of several antioxidant enzyme was found to increase. Plant regeneration from in vitro cultures treated with 2,4-D was obtained after subculturing explants onto PGR-free media. The ISSR analysis of 2,4-D-treated material showed that this plant growth regulator (PGR) induced measurable genetic variations. The global DNA methylation rates (GMR) as estimated through the HPLC analysis of nucleosides also confirmed the presence of epigenetic changes caused by 2,4-D as GMRs increased from 13.8 to 18.93%.