Team:Freiburg/Notebook/21 July

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green light receptor

NAME OF YOUR EXPERIMENT

Investigators:NAME

blue light receptor

NAME OF YOUR EXPERIMENT

Investigators:NAME

red light receptor

NAME OF YOUR EXPERIMENT

Investigators:NAME

Lysis cassette

NAME OF YOUR EXPERIMENT

Investigators:NAME

Precipitator

Precipitator Ruediger

inoculated cells from yesterdays plating. All 12 plates had colonies. I picked 2 from each plate. Incubation at 37°C overnight at 300rpm.

further Project plan

• if GFP Pdb works, I will pick best affinity Pbd and use it for the precipitator.

• I will also need a primer for the Precipitator with a GST tag and a TEV cleavage site[http://en.wikipedia.org/wiki/TEV_protease] inbetween.

• Need to order a GST column (GST-4B) for the ÄKTA as well as PreScission Protease GST

• check toolbox plasmid for pGEX-6P

• check affinity calculation programs Scatchand plots and Klötz (to estimate Ni-Precipitator binding)

• • data for these calculations could be generated with ultrafiltration of the protein bound to nickel, to see how much nickel goes through - can be tested chemically

• 300ml E.coli culture needed for protein purification in ÄKTA

• first there will be the GST affinity purification of the lysate of the E. coli culture -> Precipitator will be in the eluate

• second a digestion of the eluate with the Cleavage enzyme

• third another GST affinity purification -> Precipitator will be in the flowthrough