Droplet-Vitrification Protocol for Cryopreservation of Ginger (Zingiber officinale) Shoot Tips

Ginger (Zingiber officinale), a globally grown and economically valuable plant, has inadequate research on germplasm cryopreservation, and droplet-vitrification is yet to be applied. The present study established an efficient droplet-vitrification protocol for Z. officinale ‘Yunnan Xiaohuangjiang’. The droplet-vitrification procedure was as follows: excise 1.5–2.0 mm shoot tips with 3–4 leaf primordia from five-week-old cultures, preculture on MS medium with 0.25 M sucrose for 1 d, treat with MS liquid medium with 2 M glycerol and 0.4 M sucrose for 20 min, dehydrate with PVS2 plus 0.1 M ascorbic acid at 0 °C for 20 min, plunge into LN for 1 h, thaw in MS liquid medium with 1.2 M sucrose for 20 min, post-culture on shoot recovery medium (MS with 0.1 g/L GA3) in the dark for 3 d. Histological and ultrastructural analyses revealed that PVS + ascorbic acid-treated shoot tips exhibited numerous living cells with small vacuoles in the apical dome, leaf primordia, and basal parts. Genetic stability results showed that the plantlets regenerated from cryopreserved shoot tips had no genetic variation. This is the first report on ginger cryopreservation via droplet-vitrification, providing technical support for ginger germplasm cryopreservation and virus elimination cryotherapy in ginger.