Shoot-inducing medium (SIM)

1X Gamborg’s B5 salts

20 g sucrose

0.5 g 2-[N-morpholino]ethanesulfonic acid (MES)

KOH (1 N)

7.5 g agar, tissue-culture-grade

1 ml N⁶-(2-isopentenyl)adenine (2-iP) (1000X)

500 μl indole-3-acetic acid (IAA) (2000X)

50 mg kanamycin

500 mg timentin

500 mg carbenicillin may be used in place of timentin.

For 1 liter of SIM, add Gamborg’s B5 salts, sucrose, and MES. Adjust pH to 5.7 with KOH, and add the tissue-culture-grade agar. After autoclaving, cool medium to ~60ºC and then add 2-iP (final concentration 5 mg/liter), IAA (final concentration 0.1 mg/liter), kanamycin (final concentration 50 mg/liter), and timentin or carbenicillin (final concentration 500 mg/liter). The kanamycin functions in transgene selection; the timentin/carbenicillin is used to eliminate Agrobacterium after transformation.