Multiple pulse electroporation of lactic acid bacteria Lactococcus lactis and Lactobacillus casei

Genetic manipulation of lactic acid bacteria is often difficult due to the inability to transform them with high efficiency. Multi-pulse electroporation offers a simple approach to increase transformation efficiencies. Using cells grown with 1% glycine and pretreated with lithium acetate and dithiothreitol, multi-pulse electroporation (five pulses of 12.5 kV cm^-1) of Lactococcus lactis JB704 cells resulted in a transformation efficiency of up to 1.2 × 10⁶ colony forming units (CFU) μg^-1 pGK13, an 8-fold increase in the transformation efficiency compared to single pulse electroporation. Other cell growth and pretreatment conditions with JB704 resulted in lower transformation efficiencies but had 4-fold to 27-fold higher transformation efficiencies with the five pulse electroporations. With similarly grown and pretreated Lactobacillus casei 32G cells, multi-pulse electroporation (five pulses of 7.5 kV cm^-1) resulted in a mean transformation efficiency of 7.3 × 10³ CFU μg^-1 pTRKH2, a 4-fold increase in the transformation efficiency compared to single pulse electroporation.