Ilex aquifolium L. is an endemic species in Algeria's Tellian and Saharan Atlas mountains in the country's center and east, and it is on the verge of extinction owing to a variety of reasons. To help preserve it, we devised an in vitro micropropagation technique. The cuttings with axillary buds were cultured on different media supplemented with different combinations and concentrations of plant growth regulators. The best results were obtained on Woody Plants Medium (WPM) supplemented with 6-benzylaminopurine (BAP: 1.5 mg/LL, 3 mg/L) and naphthaleneacetic acid (NAA: 1.5 mg/LL) in combination with BAP (1.5 mg/ L); BAP (1.5 mg/ L) with Gibberellins (GA3: 0.5 mg/ L). The best shoot proliferation rates were obtained on WPM supplemented with 1.5 mg/LL (BAP) and BAP: 1.5 mg/L; GA3: 0.5 mg/L; or BAP: 1.5 mg/L + ANA: 1.5 mg/L. The highest rooting rate (100%) was obtained with shoots grown on WPM medium supplemented with NAA (1.5 mg/LL). Absence of routing formation was observed in media containing indolacetic acid (IAA: 1 mg/ L) alone or combined with BAP: 0.5 mg/ L) The best-rooted shoots must be acclimatized to a substrate consisting of a mixture of peat and sand (1:1 v/vv), the seedlings have been acclimatized successfully.