Callus induction and plant regeneration derived from hypocotyls of alfalfa (Medicago sativa L.)

Hypocotyls of 4-5d's seeding after germination was used as explants and modified SH (SH macronutrients + MS micronutrients + UM organic compound) + Casein Enzymatic Hydrolysate N-Z-Amine (CH) 2 000 mg/ l as a basal medium to study callus induction and plant regeneration of alfalfa. The induction and continued medium of callus was added 2,4-D 1.0 mg/ l+KT 0.5 mg/ l+ sucrose 30 g/ l and 2,4-D 0.05 mg/ l+BA 0.5 mg/ l+ sucrose 30 g/ l. The callus was transferred into different medium, according to Orthogonal design principle, so that an optimal culture medium was selected, the results showed that the basal medium + KT 0.4 mg/ l+ sucrose 20g/ l was the optimal medium for differentiation. And it's differentiation ratio from hypocotyls of alfalfa was 73.0%. Within 10 days after the regenerated bud was transferred into the rooting medium (1/ 2MSO), it is possible to get a plantlet with shoots and roots. The result provide a basic foundation to produce transgenic alfalfas that are resistant against salt stress by introducing Na~+/H~+ antiporter gene from synechocystis species pcc 6803.