Three psychrotrophic LAB species,Leuconostoc gelidumsubsp.gasicomitatumLMG 18811,Lactococcus pisciumMKFS47, andPaucilactobacillus oligofermentansDSM 15707, were used for the experiment. A frozen glycerol stock solution (1.8 ml) of a bacterium was inoculated into 9 ml of de Man-Rogosa-Sharpe mediu...
pH 7.5, 0.1 M KCl, 20 μM IseG, 80 μM reconstituted MBP-IseH, 1 mM SAM, 100 μM Ti(III) citrate and 5% glycerol was incubated at RT for 10 min in the glovebox. A control
After repeating this procedure three times, a stock solution of the third generation of bacteria was prepared in 50 mL of TSB with the addition of 10%(v/v) glycerol (Sigma-Aldrich) and stored at −80 °C. This stock solution was used as the starting point for all experiments. ...
Bacteria from a -80 °C glycerol stock were streaked on tryptic soy agar (TSA) or lysogeny broth (LB) agar with 200 μg/mL kanamycin, and grown for 24 h at 37 °C. Subsequently, a single colony was selected, and 4 mL of tryptic soy broth (TSB) or LB with 200 μg/mL kanamycin...
The cells were resuspended in 10% glycerol and stored at -80 °C in 100 µl aliquots. For electroporation, 4 µl (~ 250–300 ng) of AMp08 plasmid DNA was mixed with the recipient strain by gentle vortexing and kept on ice for 30 min. The cell-DNA mixture was then loaded ...
The cells were then harvested by centrifugation at 8000 g at 4uC for 10 min and washed four times with ice-cold 10% glycerol. The cells were resuspended in 10% glycerol at a 1,000-fold greater concentration compared to that of the original culture volume. The cell sus...
For all arsenic-resistant isolates, glycerol stocks were prepared with a 25% final glycerol concentration and stored at −80 °C. 2.5. Arsenic Transformation Assay An arsenic transformation assay was performed using a silver nitrate test [28]. Different kinds of agar plates (Luria Bertani (LB)...
The fermentation inoculum was prepared by pre-culturing the LAB consortium from glycerol stock culture in an MRS broth overnight. The bacteria were reinoculated in MRS broth, cultivated, and harvested at OD600 of 2.0 by centrifugation at 3000× g for 20 min at 4 °C (Sigma 3–16PK centrifu...
If using a glycerol stock, the integrity of the plasmid can change because most cell strains for expression are not RecA and EndA-. Use freshly transformed cells. The protein is in the insoluble section; check cell lysate...
Glycerol stocks were made from the transformed cells containing the correct antibiotic resistance. All cultures were started from a freshly streaked glycerol stock on LB agar plates with the appropriate combination of antibiotics and 0.08% glucose. Pre-cultures were inoculated with 5–20 colonies and ...