The percent work aimed to establish an efficient in vitro propagation and conservation protocol for Brunfelsia pauciflora plant using encapsulation techniques that allow the genetic stability of conserved material. The effect of plant growth regulators types (BAP, Kin, 2ip, and NAA) and concentrations at 0.0, 1.0, and 2.0 mg/l were stud-ied during the establishment phase in comparison with a hormonal-free medium. In vitro proliferated shootlets were encapsulated using various levels of sodium alginate at 1, 2, and 3% (W/V) and calcium chloride at 2.5,5 and 10 %. The regrowth ability of encapsulated explants was evaluated after conservation for 3, 5, and 7 weeks and compared with non-encapsulated buds. The findings of the current study proved that during the establish-ment phase, applying BAP at 2.0 mg/l gave the best results during the three successive subcultures. The best treatment 2ip (2.0mg/l) + BAP (2.0mg/l) + NAA (2.0mg/l) that was considered ideal propagation protocol for shooting and rooting of Brunfelsia. Successful plant recovery from encapsulated explants using low concentra-tions of sodium alginate (1%) and calcium chloride at 2.5% proved that the explants could to survive at the high-est percent (100%) during conservation periods 3, 5, and 7weeks. Increasing calcium chloride to 5% with the same concentration of sodium alginate (1%) caused a gradually reduced percent of survival to 53.33% with an increasing conservations period (from 3 to 7weeks). It is the first paper on the application of the syndetic seed technique on Brunfelsia for the shortest period of conservation and successive renovation of shootlets whose genetic stabilization was evaluated by analysis of RAPD that proved that the genetic similarity ranged from 56.1 to 80%. These results were confirmed by anatomical sections made in Brunfelsia leaf blade.