Chrysanthemum morifolium Ramat. is an exciting trade plant due its ornamental importance. Our research aimed to assess plantlet regeneration efficiency by using stem nodal segments as ex plant source through direct shoot regeneration, organogenesis, and somatic embryogenesis through callusing. Various sterilization protocols were followed to raise pathogens free plantlets. Various PGR concentrations and combinations were applied to induce, direct shoot-roots regeneration from explant, organogenic, and somatic embryogenic callusing for mass scale plantlet regeneration were induced. Optimal stem nodal segments sterilization protocol (75% alcohol for 15 s + 0.1% HgCl2 for 2 mins +washing with double distilled sterilized water 3 times) gave satisfactory results. Maximum shoot regeneration response was achieved from stem nodal segments and shoot tips when Benzyl Amino Purine (BAP) was used on MS 0.5 mg/L. The no of shoots induction was higher when MS BM was fortified with BAP with 1-2 mg/L. With fortification MS 2 mg/L of BAP shoot multiplication increased but growth was much suppressed. Further shoot growth, elongation and photosynthetic tissue formation occurred on MS BM fortified with Kn 1.0 mg/L+ IAA 0.5 mg/L. All micro shoots rooted on MS supplemented with 0.5mg/L IAA+ 0.1 mg/L Kn. Rooted plantlets growth occurred on MS BM and up to 60 plantlets per segment were obtained. From stem nodal segments and shoot tips Organogenic callus was induced on half MS fortified with BAP + 2,4-D 1.0 +0.5m/L, Kn + IAA, 2+1 mg/L. However, callus was also induced on MS BM supplemented with 1-2 mg/L BAP in 50% cases. Shooting occurred from the callus after 4-6 weeks of inoculation, on BM with 1-2 mg/L of BAP. The micro cuttings rooted on BM fortified with 0.1mg/L Kn + 0.5 mg /L IAA after 4.5 to 6 weeks of inoculation. However, MS BM fortified with 0.5 mg/L BAP and .1 mg/L NAA induced embryogenic callus. A light maroon embryogenic callus was formed on MS fortified with BAP and NAA minimum quantity (0.5mg/L). Embryogenesis or plantlet regeneration from the callus was induced by fortifying the MS with 1.0 mg/L BAP. Up to 60 plantlet redevelopments from small callus mass occurred after 3-4 weeks of culture. It was found that more than 95% plantlet established under ex vitro conditions when using polythene bags etc. for acclimatization and transferring to natural conditions. Micropropagagules were morphologically better than mother plant and flourished well, showed superior quality in morphology, color of flower, shine, texture, and early maturity compared to conventional propagules. Per flask 200 plantlets regeneration rendered somatic embryogenesis the most effective way for producing mass scale propagation of virus free plantlets. Ornamental market can take benefit of mass scale propagation in Pakistan.

Index Terms: in vitro – Chrysanthemum morifolium, direct shoots-roots induction, Organogenesis; Somatic embryogenesis, pathogens free plantlets regeneration