IGEM:IMPERIAL/2009/Assays Protocols/M3 Assays

Assays for Cell Death Strategy

Assay to determine cell death

Media:Cell Death Assay.pdf nbsp; Qualitative and quantitative

Restriction Enzyme Assay

Using GFP marker

Basic genetic circuit for the restriction enzyme testing construct:

Here, there are 3 possibilities for the expression of the GFP, and these are shown below:

i) Non-functional restriction enzyme co-expressed with the GFP.
ii) Basal levels of GFP expression suggesting that the promoter is not functioning properly.
iii) Initial increase in flourescence, followed by a plateauing of the flourescence, as the restriction enzymes destroy the genetic material within the cell.

Cell Death Assay

By Colony Forming Units

1) Plating dilution series on TSB Agar Plates

2) Counting of colonies after overnight incubation

3) Determining the number of colony forming units (number of cells still alive)