Gentiobiose Feeding in Gentian in vitro Overwintering Buds or Plantlets

Materials and Reagents

1. Gentian (Gentiana triflora) tissue culture plantlets
   
2. Sucrose
   
3. Gentiobiose (Sigma-Aldrich, catalog number: G3000-5G )
   
4. Gellan gum (Wako USA, catalog number: 075-03075 )
   
5. Liquid nitrogen
   
6. Milli Q grade water
   
7. MS vitamins (see Recipes)
   
8. Propagation medium (see Recipes)
   
9. IOWB induction medium (see Recipes)
   
10. Gentiobiose medium (see Recipes)
    

Equipment

1. Sterile magenta boxes (sterilized by autoclaving)
   
2. Surgical tape (3M, catalog number: 1530-0 )
   
3. Glass culture tubes (sterilized by autoclaving)
   
4. Silicon plugs (sterilized by autoclaving)
   
5. Sterile 15 ml plastic tubes (such as Greiner Bio-One GmbH, catalog number: 188271 )
   
6. Sterile syringe filter (Millipore, catalog number: SLGP033RS )
   
7. Sterile petri dishes (IWAKI PUMPS, catalog number: SH90-20 )
   
8. Cutoff filter (Millipore, catalog number: UFC5003BK )
   
9. Growth chambers
   
10. Clean bench
    
11. Scalpel
    
12. Forceps
    
13. Ball miller
    
14. Centrifuge
    
15. Freeze dryer
    

Procedure