Team:XJTLU-CHINA/Protocols Electroporation

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

Electroporation for L. lactis

Materials:

Procedure:

  1. 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.
  2. Use the Electroporator with following adjustments:
    2500 V (12.5 KV/cm)
    25 μF
    200 Ω - Pulse (normal reading is 4.5-5 msec)
  3. Add 1 ml G/L-M17B + 20 mM MgCl2 + 2 mM CaCl2.
  4. Keep the cuvette for 5 min on ice and incubate 1-1.5 h at 30 °C.
  5. 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:

  1. 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.
  2. 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

Location

Rm 363, Science Building
Xi'an Jiaotong-Liverpool University
111 Ren'ai Road, Suzhou, China
215123

Get in touch

emali

igem@xjtlu.edu.cn

XJTLU-CHINA iGEM 2017