Assessment of genetic stability of Cucumis sativus L. regenerated from encapsulated shoot tips

- We have optimized the concentration of sodium alginate and calcium chloride for the production of synthetic seeds in cucumber for efficient conversion of encapsulated shoot tips into plantlets.
- First application of slow growth storage techniques in cucumber with assessing influences of different medium was developed.
- In vitro flowering were induced making the method more effective.
- We have also hardened and developed true to type plantlets re-grown from synthetic seed as assessed by molecular marker (RAPD and ISSR).

An encapsulation protocol to obtain synthetic seed was standardized in Cucumis sativus L. Initially microshoots were regenerated through organogenesis from epicotyl segments of in vitro grown seedlings on Murashige and Skoog's (MS) basal medium fortified with 4.4 μM 6-benzylaminopurine (BAP) and 0.6 μM α-napthaleneacetic acid (NAA). The shoots were continuously multiplied on MS basal medium supplemented with 4.4 μM 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 °C up to 8 weeks. The synthetic seeds were converted into plantlets on 0.8% agar solidified growth regulator free full-strength MS medium. Plantlets retrieved from the encapsulated shoot tips were hardened and established 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 synthetic seeds with the mother plant. A total of 106 loci amplified and all loci were found to be monomorphic in nature indicating the homogeneity among the regenerants. The present study paves the way for the identification and maintenance of genetically uniform C. sativus plants re-grown from synthetic seeds and hardened in the field.