"
Team:Groningen/Notebook/25 September 2009
From 2009.igem.org
(Difference between revisions)
(New page: {{Team:Groningen/Notebook/Day/Header}} ==Wet== ===GVP Cluster=== ===Transporters=== ===Metal Accumulation=== *'''Harvest ''E.coli'' cells and As(III) uptake experiment''' **Take OD sa...) |
(→Metal Accumulation) |
||
| (One intermediate revision not shown) | |||
| Line 15: | Line 15: | ||
**Wash 2x with TB74S buffer. | **Wash 2x with TB74S buffer. | ||
**Resuspend upto OD=10. | **Resuspend upto OD=10. | ||
| - | **Dry 1ml sample for 2hrs @ 65°C. | + | **Dry 1ml sample for 2hrs @ 65°C. |
**Do uptake experiments with the different samples (1gr cells @ 22mg/ml concentration) | **Do uptake experiments with the different samples (1gr cells @ 22mg/ml concentration) | ||
***At As(III) concentrations for 1hr: 0, 10, 20, 50 and 100uM As(III). | ***At As(III) concentrations for 1hr: 0, 10, 20, 50 and 100uM As(III). | ||
| Line 23: | Line 23: | ||
**Dry for 2days @ 65°C. | **Dry for 2days @ 65°C. | ||
| - | + | :: '''The determined dry weight was 5x lower than expected, so the sample which should have contained at least 1gr dry cells only contained ~100-200mg dry cells.''' | |
===Vectors=== | ===Vectors=== | ||
Latest revision as of 20:11, 30 September 2009
[http://2009.igem.org/Team:Groningen http://2009.igem.org/wiki/images/f/f1/Igemhomelogo.png]
|
|---|
Wet
GVP Cluster
Transporters
Metal Accumulation
- Harvest E.coli cells and As(III) uptake experiment
- Take OD sample to deterimine OD600.
- Harvest E.coli WT (pSB1A2 + pLac), E.coli + pArsR-RFP and E.coli + pLac-fMT by
spinning down @ 6000rpm for 20min in Sorvall centrifuge.
- Wash 2x with TB74S buffer.
- Resuspend upto OD=10.
- Dry 1ml sample for 2hrs @ 65°C.
- Do uptake experiments with the different samples (1gr cells @ 22mg/ml concentration)
- At As(III) concentrations for 1hr: 0, 10, 20, 50 and 100uM As(III).
- And at time points with 10uM As(III) at: 0, 10, 20, 40 and (60)min.
- For fMT only the concentration samples were taken!
- Spin down, wash with TB74S and resuspend in 10ml ddH2O.
- Dry for 2days @ 65°C.
- The determined dry weight was 5x lower than expected, so the sample which should have contained at least 1gr dry cells only contained ~100-200mg dry cells.
Vectors
Dry
|
|
|
| |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
|
|
|
| |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
