TABLE 4.
Parameter change | Final ODa | Period of growth (h)b | Electroporation setting |
Cuvette size (mm) | Amt (μg) of DNA | Recovery mediumc | CFUd | ||
---|---|---|---|---|---|---|---|---|---|
Voltage (kV) | μF | Ω | |||||||
Settingse | 0.483 | 1.75 | 1.5 | 25 | 600 | 1 | 1 | RG2 | 3 |
Settingsf | 0.483 | 1.75 | 2.0 | 25 | 200 | 2 | 1 | RG2 | 33 |
Settings | 0.424 | 2.9 | 2.0 | 25 | 400 | 2 | 0.7 | RG2 | 157 |
Recovery medium | 0.519 | 1.75 | 2.0 | 25 | 400 | 2 | 1 | LB2 | 960 |
Recovery medium | 0.519 | 1.75 | 2.0 | 25 | 400 | 2 | 1 | RG2 | 5 |
Fast growthg | 0.539 | 1 | 2.0 | 25 | 400 | 2 | 2.5 | LB2 | 1,900 |
μg DNA addedg | 0.617 | 1.3 | 2.0 | 25 | 400 | 2 | 0.2 | LB2 | 2,409 |
μg DNA addedg | 0.617 | 1.3 | 2.0 | 25 | 400 | 2 | 0.02 | LB2 | 5,118 |
Final OD600 after the indicated number of hours when growing cells prior to making them competent.
Number of hours cells had grown before making them competent.
RG2 is LB2 with 121 g/liter sucrose and 10 g/liter glucose (29).
CFU indicate CFU per μg DNA. Using fresh cells or cells stored at −80°C did not change CFU counts.
Settings from reference 29.
Settings are based on reference 30, the same settings as those used for screening of Geobacillus strains.
In these experiments, after overnight growth, cells were transferred into 500-ml Erlenmeyer flasks or 1-liter bottles (having a similar bottom surface) instead of into a 250-ml Erlenmeyer flask to allow for more aeration.