Preparation of Bacterial Cultures and Inoculation of Plants

Plants for infection with P. syringae should be grown on a short-day light cycle (8-12 hours light), so that they develop large leaves. For consistent results, it is important that the plants are watered well and do not experience any abiotic stresses. It is also necessary to control the humidity, as the extent of bacterial growth is highly dependent on humidity and temperature: 22°C and 80% relative humidity give good results. These conditions are best achieved by growing and testing the plants in a temperature- and humidity-controlled growth chamber.

Materials

Bacterial stock culture

King's B medium (per liter)

10 g of proteose peptone #2 (Difco) 1.5 g of anhydrous K2HP04 15 g of glycerol

Adjust the pH to 7.0 with HC1. After autoclaving, add 5 ml of sterile 1 m MgSO (if MgS04 is added before autoclaving, the medium becomes clouay) and any necessary antibiotics.

Syringe (1-cc without a needle)

Silwetm L-77 (0.02% v/v) (Vac-in-Stuff, available from Lehle Seeds)

Optional, see Step 3. CAUTION: K2HP04, MgS04 (see Appendix 3)

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