I am trying to make my bacteria auto-fluorescent by incorporating luxABCDE gene into gram-positive bacteria. However, as I was thinking about this, I came across a question. Could I still use the heatshock step to transform the gene into gram positive bacteria? If not, what's the best protocol to do the transformation?
Thanks!
From my experience, Lactobaillus transformation works well using the following protocol：
'An overnight culture of the transformed L. reuteri strains was used to inoculate 15 mL MRS broth containing 2% glycine and 0.5 M sucrose, and incubated at 37°C. The absorbance of the culture at 600 nm reached 0.2 after 3 h, after which the cells were separated by centrifugation and then washed twice with 5 mL cold distilled water followed by a 10 min incubation in 50 mM EDTA solution (pH 8.0). After washing with cold distilled water and 0.3 M sucrose, the cells were re-suspended in 100 μL of 0.3 M sucrose solution as electroporation buffer. Then the suspension was transferred into a disposable cuvette (Bio-Rad Laboratories) and subjected to an electric pulse using a MicroPulser (Bio-Rad) under the following conditions: Capacitance = 25 μF; Resistance = 400 ohm; Voltage = 2.0 kV. '
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