An encapsulation protocol to obtain synthetic seed was standardized in Cucumis sativus L. Initiallymicroshoots were regenerated through organogenesis from epicotyl segments of in vitro grown seedlingson Murashige and Skoog’s (MS) basal medium fortified with 4.4 M 6-benzylaminopurine (BAP) and0.6 M -napthaleneacetic acid (NAA). The shoots were continuously multiplied on MS basal mediumsupplemented with 4.4 M BAP. The shoot tips from the fourth subculture were encapsulated with 3%sodium alginate and 100 mM calcium chloride. Encapsulated shoot tips were successfully stored at 4◦Cup to 8 weeks. The synthetic seeds were converted into plantlets on 0.8% agar solidified growth regulatorfree full-strength MS medium. Plantlets retrieved from the encapsulated shoot tips were hardened andestablished in field. 16 random amplified polymorphic DNA (RAPD) and 10 inter simple sequence repeat(ISSR) markers were applied to analyze the genetic stability of the plants regenerated from syntheticseeds with the mother plant. A total of 106 loci amplified and all loci were found to be monomorphicin nature indicating the homogeneity among the regenerants. The present study paves the way for theidentification and maintenance of genetically uniform C. sativus plants re-grown from synthetic seedsand hardened in the field.