Induksi keragaman genetik melalui iradiasi sinar gamma pada kalus embriogenik hasil kultur protoplas jeruk siam

Abstract

Tangerine cv. Siam has sweet flavor, easily peeled skin, soft and juicy flesh. However, it has relatively many seeds (15-20 seeds per fruit), so it can not be competed with citrus from other countries. Fruit quality improvement of citrus has been the subject of citrus breeding program. The first step of breeding program is to increase variability, in order to efficient the selection process. Callus derived protoplast which sub cultured several years has a level of variability. The objective of this research was to increase variation of Tangerine cv. Siam through gamma irradiation on callus derived protoplast. Callus was exposed to gamma irradiation at 0, 10, 20, 30, 40, 50, 60, 70, 80, 90 and 100 Gray. Observation on the growth of callus four weeks after irradiation showed variation on morphology and weight of callus. At low doses (10-50 Gray) callus growth were not inhibited, but at high doses (60-100 Gray) callus growth were inhibited. The result of radio sensitivity dose analyzed by Curve Expert 1.4 software was 53.25 Gray. Callus regeneration ability (somatic embryo maturation and germination) were very diverse between irradiated and non-irradiated callus. Gamma irradiation affects the formation of somatic embryos. After four weeks on MW (Morel-Wetmore) medium containing 0.5 mg/l ABA, 50 Gray callus produced more somatic embryos than other doses. After four weeks on MW medium containing 0.5 mg/l GA3, only 75.9% somatic embryos from 50 Gray callus could germinate, less than other doses. All somatic embryos from non-irradiated callus could germinate. The germination of somatic embryos produced 72 regenerated plantlets. Dendogram based on morphological observations of 0 Gray regenerated plantlets showed 40% variability, while of 50 Gray were 47% and that of 60 Gray were 46%. Ten regenerated plantlets were chosen based on its variability and growth (P-2, P-8, 50-4, 50-6, 50-15, 50-24, 60-8, 60-10, 60-11 and 60-23). Dendogram based on morphological observation between 10 regenerated plantlets showed 30% variation. Molecular analysis of the 10 regenerated plantlets using three ISSR primers (ISSR-1, ISSR-2 and ISSR-4) produced 17 bands with nine polymorphic bands (52.94%). Dendogram based on molecular analysis between 10 regenerated plantlets and wild type shoots showed 22% genetic variation. In vitro and ex vitro grafting between regenerated plantlets and JC rootstock could accelerate optimal growth of plantlets. Application of ex vitro grafting was more efficient than in vitro grafting. Ex vitro grafting did not need acclimatization stage, where as in vitro grafting still need acclimatization stage before transfer to soil.