SOPs

Quantitative real-time PRC is used to compare kdpFABC expression between the E. coli strains MG1655 (wildtype)and MG1655 (kdpA4, a kdpFABC-inactive mutant after a shift to K+ limitation.

A method to compare kdpFABC expression between MG1655 (wildtype) and MG1655 (kdpA4) after a shift to K+ limitation, the RNA was extracted from samples taken at different points in time.

Pulsed-FRAP measure the diffusion constants in confined volumes in small cells like E. coli and other bacteria or cellular organelles.

A procedure to measure the levels of GSH and SLG before and after exposure to MG using formic acid. A similar expreimental set up as for potassium efflux experiments is used to which a silicon oil centrifugation step is incorporated. Samples are analysed by LC-MS-MS in order to quantify intracellular GSH and SLG levels.

The reverse transcriptase synthesizes DNA, which complements the mRNA template.

To induce kdpFABC expression, cells are cultivated in K10 minimal medium (10mM K+) were shifted into K+ limiting growth medium (K0), containing 20 μM K+ by filtration.

FRAP experiments are used for studying cytoplasmic diffusion in cells and cells membrane.

Growing Escherichia coli to express KefF by adding IPTG for purification and kinetics experiments.

Generating gene knock-ins using MJF618 expressing the defective lambdoid prophage recombination system λ red.

Generating gene knock-outs using DY330 expressing the defective lambdoid prophage recombination system λ red.

Experimental system that is designed to observe the in vivo stability and aggregation of a protein of interest.