| For Agrobacterium |
| MG/L medium (per liter) | 5 g tryptone (Gibco, 211705);2.5 g yeast extract (BD Biosciences, 212750);5 g NaCl (Duchefa Biochemie, S0520);5 g Mannitol (Duchefa Bichemie, M0803);0.1 g MgSO4 (Sigma, M7506);0.25 g K2HPO4 (Sigma, P3786);1.2 g Glutamic acid (Sigma, G6904) |
| · Adjust pH to 7.2 with 0.1 N NaOH, add 15 g Bacto agar (BD Biosciences, 214010), and autoclave.· After cool-down, add 1 mL of 200 mM Acetosyringone, 1 mL of Carbenicillin (100 mg/mL) and a second antibiotics depending on the vector used for transformation (ex., Kanamycin).· Pour into petri dishes, and store at 4℃ (up to 2 weeks). |
| For Brachypodium |
| Callus induction medium(CIM, per liter) | 4.43 g Linsmaier and Skoog medium (Duchefa Biochemie, L0230);30 g Sucrose (Duchefa Biochemie, S0809);0.6 mL CuSO4 stock solution (1 mg/mL);1 mL 2,4-D stock solution (2.5 mg/mL) |
| · Adjust pH to 5.8 with 0.1 N KOH, and autoclave.· For liquid CIM, store in a sterile glass bottle at 4℃ (up to 2 weeks).· For solid CIM, add 6 g Phytoagar (Duchefa Biochemie, P1003) before autoclave. After cool-down, pour into petri dishes, and store at 4℃ (up to 2 weeks). |
| Selection medium(SM, per liter) | CIM+Cefotaxime+Hygromycin(ex., depending on the selectable marker) |
| · Add 6 g Phytoagar to CIM before autoclave.· After cool-down, add 1 mL of Cefotaxime stock solution (300 mg/mL) and 1 mL of Hygromycin stock solution (40 mg/mL), stir well, pour into petri dishes, and store at 4℃ (up to 2 weeks). |
| Regeneration medium(RGM, per liter) | 4.43 g Linsmaier and Skoog medium (Duchefa Biochemie, L0230);30 g Maltose (Duchefa Biochemie, M0811) |
| · Adjust pH 5.8 with 0.1 N KOH, add 6 g Phytoagar, and autoclave.· After cool-down, add 1 mL of Kinetin stock solution (0.2 mg/mL), 1 mL of Cefotaxime stock solution (300 mg/mL), and 1 mL of Hygromycin stock solution (40 mg/mL), stir well, pour into petri dishes, and store at 4℃ (up to 2 weeks). |
| Rooting medium(RM, per liter) | 4.43 g Murashige and Skoog medium (Duchefa Biochemie, M0222);30 g Sucrose (Duchefa Biochemie, S0809) |
| · Adjust pH 5.7 with 0.1 N KOH, add 2.5 g Phytagel (Sigma, P8169), and autoclave.· After cool-down, add 1 mL of Cefotaxime stock solution (300 mg/mL), and 1 mL of Hygromycin stock solution (40 mg/mL), stir well, pour into petri dishes, and store at 4℃ (up to 2 weeks). |