Teaching Points for Protein Expression Lab


Lab Protein Expression                  Mentor Sheet                                   3/6/2011

Teaching Points

They will work in pairs – but each student should end up with their own 50 ml tube of spun down bacteria to put in the freezer.

Encourage them to fill in their lab notebooks as they do the lab.

It is due when they are done with the lab.

Be SUPER careful of the gas burners! – they don’t appreciate how dangerous they can be.

They should never leave the burner on if they leave the area.

Get the E. coli competent cells out LAST. They have to work with these quickly.

The heat shock must be EXACTLY  42OC for 45 seconds.

For the ‘FUN’ plate – they can take a Kim Wipe or a cotton swab and swipe it on anything they think is ‘dirty’ (e.g. the lab floor, the sink, etc.) Then see what grows.

 

Be sure to explain to them what each things is for:

  • Competent cells (made to easily uptake DNA)
  • Plasmid – circular DNA with their gene of interest
  • Heat shock – opens up pores on membrane to uptake DNA (or makes more ‘fluid’)
  • LB– nutrient and energy rich media
  • Agar – makes it gel like (solid)
  • Ampicillin – plasmid has beta-lactamase gene which is the enzyme that breaks down Ampicillin
  • Log growth – the ‘happy’ phase for cells to be in when they produce the most protein
  • Lysozyme – enzyme that breaks down bacterial cell walls (peptidoglycan)

 

You don’t need to mention IPTG induction since we don’t use it in this expression.

Be sure they know that they don’t spin down their sample until it is PURPLE – which means they may have to wait til the next day (Day 4)

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