Team:Groningen/fluorimetry

From 2011.igem.org

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{{HeaderGroningen2011}}  
{{HeaderGroningen2011}}  
=Fluorimetry=
=Fluorimetry=
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<br>[[Team:Groningen/flowcytometry#fluorimetry protocol| Fluorimetry protocol]]
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[[Team:Groningen/fluorimetry#fluorimetry protocol| Fluorimetry protocol]]
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<br>[[Team:Groningen/cloning#M9 minimal media| M9 minimal media]]
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<br>[[Team:Groningen/fluorimetry#M9 minimal media| M9 minimal media]]
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<br> The OD600 and fluorescence of a great number of samples can be measured the same time with the fluorimeter.
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<br> The OD600 and fluorescence of a great number of samples can be measured at the same time with a fluorimeter.
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<br> We used it for measuring PBADaraC-cI-LVA+PcI-GFP with different concentrations of arabinose as an induction  
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<br> We used it for measuring pBAD/araC-cI-LVA+pcI-GFP with different concentrations of arabinose as an induction  
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<br> signal
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<br> signal.
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<br>
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<br>  
=Fluorimetry protocol=
=Fluorimetry protocol=
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<br> Inoculate 10ml M9 medium+ antibiotics + 1ml 2% casamino acids and grow the cultures overnight at 37°C.
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Inoculate 10ml M9 medium+ antibiotics + 1ml 2% casamino acids and grow the cultures overnight at 37°C.
<br> Grow a logfase to an OD600 of 0.4.  
<br> Grow a logfase to an OD600 of 0.4.  
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<br> Dilute in a way that the OD600 is decreased to a value of 0.004/
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<br> Dilute in a way that the OD600 is decreased to a value of 0.004
<br> Add 300μl of the diluted culture into a well.
<br> Add 300μl of the diluted culture into a well.
<br> The fluorimeter will measure the OD600+ fluorescence every 15 minutes.
<br> The fluorimeter will measure the OD600+ fluorescence every 15 minutes.
 +
<br> Fluorimeter used for measurements: infinite F200 TECAN. Serial no. : 811005514.
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<br>
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<br> [[Team:Groningen/fluorimetry#fluorimetry| Back to fluorimetry]]
 +
<br>
=M9 minimal media=
=M9 minimal media=
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<br>Component Concentrations
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Component Concentrations
<br>1x M9 salts  
<br>1x M9 salts  
<br>2mM MgSO4  
<br>2mM MgSO4  
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<br>For 1L of media  
<br>For 1L of media  
<br>
<br>
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<br>500ml 2xM9 salts ∗
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<br>500ml 2xM9 salts
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<br>20ml 0.1M MgSO4 ∗
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<br>20ml 0.1M MgSO4
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<br>200μl 0.5M CaCl2 ∗
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<br>200μl 0.5M CaCl2
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<br>469.8ml sterile deionized H2O ∗ (if carbon source to be added)
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<br>469.8ml sterile deionized H2O
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<br>479.8ml sterile deionized H2O ∗ (if no carbon source to be added, useful as a control for sugar metabolism)
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<br>Combine above solutions using sterile technique. (May notice some precipitation during preparation but precipitate  
<br>Combine above solutions using sterile technique. (May notice some precipitation during preparation but precipitate  
<br>should go back into solution once volume is brought up to 1l with sterile H2O.)
<br>should go back into solution once volume is brought up to 1l with sterile H2O.)
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<br>Add antibiotic as appropriate and store at 4°C  
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<br>Add antibiotics as appropriate and store at 4°C  
<br>
<br>
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<br>∗ Can be obtained from the media room
 
<br> http://openwetware.org/wiki/Endy:M9_medium/minimal
<br> http://openwetware.org/wiki/Endy:M9_medium/minimal
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<br>
 +
<br> [[Team:Groningen/fluorimetry#fluorimetry| Back to fluorimetry]]
 +
<br>
{{FooterGroningen2011}}
{{FooterGroningen2011}}

Latest revision as of 01:43, 22 September 2011


Fluorimetry

Fluorimetry protocol
M9 minimal media
The OD600 and fluorescence of a great number of samples can be measured at the same time with a fluorimeter.
We used it for measuring pBAD/araC-cI-LVA+pcI-GFP with different concentrations of arabinose as an induction
signal.

Fluorimetry protocol

Inoculate 10ml M9 medium+ antibiotics + 1ml 2% casamino acids and grow the cultures overnight at 37°C.
Grow a logfase to an OD600 of 0.4.
Dilute in a way that the OD600 is decreased to a value of 0.004
Add 300μl of the diluted culture into a well.
The fluorimeter will measure the OD600+ fluorescence every 15 minutes.
Fluorimeter used for measurements: infinite F200 TECAN. Serial no. : 811005514.

Back to fluorimetry

M9 minimal media

Component Concentrations
1x M9 salts
2mM MgSO4
0.1mM CaCl2
0.4% carbon source (e.g. glycerol, glucose, etc.)
In sterile H2O
Protocol
For 1L of media

500ml 2xM9 salts
20ml 0.1M MgSO4
200μl 0.5M CaCl2
469.8ml sterile deionized H2O
Combine above solutions using sterile technique. (May notice some precipitation during preparation but precipitate
should go back into solution once volume is brought up to 1l with sterile H2O.)
Add antibiotics as appropriate and store at 4°C

http://openwetware.org/wiki/Endy:M9_medium/minimal

Back to fluorimetry





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