Targetselection
Enzyme free cloning
Small scale expression screening
We have previously established a small scale expression screening, this method is implemented on a Hamilton Star. We have established protocols to purify PCR product, plasmids and proteins using magnetic beads protocols. Transformation and subsequent plating can be performed. Following transformation, growth is monitored in standard flat bottom microplates at a regular interval thereby permitting thightly controled indcution of recombinant protein production. After induction, bacterial growth is monitored until growth is completed or when rerquested incubation time is reached. Samples can either be stored in a hotel or directly lysed. Total and soluble expression is detertmined using SDS-PAGE, on samples before and after magnetic beads mediated HIS-tag purification. All steps of this protocol are performed on the Hamilton Star, without manual intervention.
Large scale purification
For Large scale purification a BioCAD Vision is used using Nickel charged Poros MC columns. This allows fast and efficient purification yielding essentially pure protein suitable for biophysical characterization.
15N HSQC screening
For all proteins that could be purified, a 15N HSQC is recorded. Examples of such NMR measurements is shown above