From 2009.igem.org
Wet
GVP Cluster
Transporters
We decided to make MasterMixes of our own. All but 1uL template, 2uL primers and 1uL taq for 25uL reactions.
1X MasterMix Buffer NH SO
| Component
| concentrations
| volumes
|
| dNTP
| 0.2 mM
| 8 uL
|
| MgCl
| 2.0 mM
| 80 uL
|
| taq buffer KCl
| 1X
| 100 uL
|
| MQ
|
| 652 uL
|
|
|
1X MasterMix Buffer KCl
| Component
| concentrations
| volumes
|
| dNTP
| 0.2 mM
| 8 uL
|
| MgCl
| 1.5 mM
| 60 uL
|
| taq buffer (NH)SO
| 1X
| 100 uL
|
| MQ
|
| 672 uL
|
|
|
|
MasterMix3
| Component | Mg/K
|
| MQ
| 28.5 uL
|
| F mut1
| 2 uL
|
| R mut2
| 2 uL
|
| dNTP
| 2 uL
|
| MgCl
| 3 uL
|
| taq buffer KCl
| 5 uL
|
| taq buffer (NH)SO
| 5 uL
|
| taq polymerase
| 2.5 uL
|
|
|
| PCR 3 program
| Temperature
| Time
|
| Denaturing
| 95°
| 2.00 min
|
|
| Start Cycles 25X
|
| Denaturing
| 95°
| 30 sec
|
| Annealing
| 55°
| 20 sec
|
| Elongation
| 72°
| 2.10 min
|
|
| End cycles
|
| Final elongation
| 72°
| 10 min
|
| Hold
| 4°
| Forever
|
|
|
|
Metal Accumulation
Vectors
Dry
KB extended the Simbiology model to support OpG and OpH in addition to OpN. Jasper in the mean time added some more explanation to the modelling section on the metal accumulation page.
In the end of the afternoon we tried to find more modelling software. Likely candidates are SEMPPR (Japer) and cytoscape (KB) Furthermore KB is trying to get up to date with the paper knowledge.
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