"
Team:Groningen/flowcytometry
From 2011.igem.org
Flowcytometry
In this project, we wanted to measure promotor activities and their leakage, but also testing the constructs
whether they work or not. Analysis of these constructs was done with the help of the FACS for measuring
fluorescence.
Flowcytometry protocol
Inoculate 10ml M9 minimal medium + 1ml of 2% casamino acids and incubate overnight at 37 °C.
Also, incubate 2 sterile erlenmeyers containing 10ml M9 minimal medium overnight at 37 °C for pre-heating.
The next day, the pre-heated erlenmeyers containing the 10ml M9 minimal medium are inoculated with 1ml of the
overnight culture and incubated for 1h-1.5h at 37 °C.
The FACS can be prepared for measurements with the protocol provided by Maarten Mols:
File:Flowcytometryprotocol.pdf
100μl suspension of the logfase cells will be used for measurements with the FACS with 100μl dilution buffer.
Later on, when the celldensity increases too much, a dilution ratio of 1:2 and 1:3 will be used.
The PBADaraC promotor needs to be induced by arabinose.
Preparation for 10% arabinose stock:
Add 1g of arabinose to 9ml milliQ and filter sterilize the stock.
Concentrations for induction can be varied from 0 to 1%.
See for more information: http://openwetware.org/wiki/Arabinose
