Electroporation for <i>L. lactis</i>

Materials:

Procedure:

Place 40 μl competent cells in a pre-chilled electroporation cuvette with 1 μl DNA (reconstituted in TE-buffer) and keep the cuvette on ice.
Use the Electroporator with following adjustments:
2500 V (12.5 KV/cm)
25 μF
200 Ω - Pulse (normal reading is 4.5-5 msec)
Add 1 ml G/L-M17B + 20 mM MgCl₂ + 2 mM CaCl₂.
Keep the cuvette for 5 min on ice and incubate 1-1.5 h at 30 °C.
Plate 10 µl, 100 µl, 900 µl on M17agar with glucose or lactose and antibiotics (depends on plasmid) - Incubate 1-2 days at 30 °C.

Tips:

Lactococcus lactis grows very slowly in G/L-SGM17B. Leaving out the sucrose is possible but can lower the transformation efficiency. The medium for cell recovery must contain MgCl₂ and CaCl₂.
The sterilization temperature can be set at 108 degrees Celsius in case of carbonization.

Applications in our project: Transformation of BBa_K2309005, BBa_K2309004 and BBa_K2309028 (independent in pNZ8148)

Collaborators and Supporters

Team:XJTLU-CHINA/Protocols dot blot