For mass production of super male hybrid lines of Asparagus officinalis(Mary Washington 500 W' CV.), in vitropropagation system was initiated to obtain a sterile, healthy culture using shoot tip and single-node spear. Explants established on Murashige and Skoog (MS) medium supplemented with kinetin (KIN) at 1 mg l^-1, naphthalene acetic acid (NAA) at 0.2 mg l^-1and gibberellic acid (GA₃) at 0.5 mg l^-1showed 100% of survival, the highest number of thick, dark green shoots (8.5) and nodes (8.7). Multiplication of shoots was carried out using benzyl amino purine (BAP) or KIN. BAP containing media gave higher shoot multiplication rate with high vitrification ratio in a positive correlation with concentration. The vitrification ratio was declined by replacing BAP with the same concentration of KIN (2 mg l^-1). Adding the growth retardant; paclopatrazol (PP333) at 1.0 mg l^-1completely eliminated the vitrification. Direct somatic embryos from stem segment (internode) induced on MS medium supplemented with BAP at 1 mg l^-1and NAA at 0.1 mg l^-1and seedling regeneration occurred on MS medium with KIN at 1 mg l^-1and GA₃at 0.5 mg l^-1. Medium supplemented with NAA, indole-3-butyric acid (IBA), PP333 and Aspergillus nigerextract were tested for rooting. MS medium supplemented with IBA at 1.0 mg l^-1and PP333 at 0.5 mg l^-1was optimum for producing white-rooted plants (79.5%) within 54 days from old clusters of shoots, which were better, compared to young single shoots. Healthy rooted clusters from each treatment were successfully transferred to growth champers then greenhouse for acclimatization.