Somatic embryogenesis and β-glucuronidase transformation in chickpea (Cicer arietinum cv. Bivanich)

Abstract

Background: Chickpea (Cicer arietinum L.) is an important kernel legume in the world. To optimize the plant tissue culture some experiments such as direct regeneration, proliferation, rooting shoots and somatic embryogenesis were done.

Methods and results: In experiments were used direct regeneration and proliferation, various levels of plant growth regulators NAA (0 and 1.0 mg/l), BAP (0, 1, 3 and 5 mg/l) and three explants' types (epicotyl, cotyledon and embryonic axis). The results of both experiments showed that embryonic axis explant was better than other explants. The highest percentage was obtained in MS media containing 1 mg/l BAP and also 3 mg/l BAP and 0.1 mg/l NAA with an average of 72%. The highest average number of branches (4.66) was found in the proliferation of embryonic axis in MS medium containing 3 mg/l BAP. The highest rooting shoot (90%) was found in 1/2MS in B5 medium vitamins with 0.2 mg/l of IBA and 0.5 mg/l NAA. Somatic embryogenesis experiments were compared on the concentration gradient of 2,4-D in fine embryonic axis explants. The results displayed that the concentration gradient of 10 mg/l 2,4-D to 5 mg/l of 2,4-D and then to zero concentration showed the highest number of embryos.

Conclusion: The best environment for regeneration embryos was MS medium with 2.5 mg/l of 2,4-D concentration gradient to zero. In this study, the PCR reaction showed the presence of the β-glucuronidase (gus) marker gene in regenerated cotyledons for 20 min in all three strains studied.

Keywords: Embryogenesis; Embryonic axis; Genetic engineering; Growth regulators; Rooting and tissue culture.