*2.3. Gamma Irradiation of In Vitro Explants*

In vitro regenerated shoots were obtained, at the differentiation during multiplication stage, of approximately 2 cm in length and were irradiated at the National Institute for Nuclear Research, located in the city of Toluca, Mexico, with a Transelektro irradiator (LG1- 01, Budapest, HU) using gamma rays with 60Co. The explants were treated with doses of 0, 20, 40, 40, 60, 80 and 100 Gy, with an exposure time of 2.1, 4.2, 6.3, 8.3, and 10.4 min, respectively. Shoot irradiation was carried out in glass Petri dishes containing MS medium without growth regulators. Twelve explants were used, with four explants per petri dish. Subsequently, the irradiated explants were immediately transferred to MS multiplication medium to avoid the possible effects of denaturation of the medium components. The explants were cultured for 60 d in the multiplication medium described above, under the aforementioned light and temperature conditions. The variables to be measured were survival percentage, number of shoots per explant, shoot length and number of leaves per shoot. The lethal dose (LD50) was calculated by observing survival percentage of explants at 60 days.
