Difference between revisions of "IGEM:Cambridge/2008/Notebook/Bacillus/2008/09/05"

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(Checking vectors)
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==Transformation of Bacillus==
 
  
* Result of Nanodrop
 
 
{|class="wikitable" style="text-align:center" border="1"
 
|-
 
!  Vector !! 260/280 !! ng/μL
 
|-
 
| ECE112 || 1.75 || 64.6
 
|-
 
| ECE166 || 172 || 138.6
 
|}
 
 
- Prepare medium A with tryptophan, and medium B
 
 
- add 5mL of medium A in 3 different tubes, inoculate each tube with colonies (1A1, IA751 ans IA771)
 
 
- Check OD every 20min
 
 
- Incubate 90min
 
 
- Add 0.45mL of medium B and 0.05mL of culture in Ependorf tubes
 
 
- Incubate 90min
 
 
- Transform : add 1μg of DNA (some with ECE112, some with ECE166) (so you need to nanodrop samples before!)
 
 
- Incubate 30min
 
 
- Pipette 200μL of solution, spread it on aech plate, wait 10min, and do it again
 
 
- Incubate 24hours
 
 
- A few glycerol tubes to stock cells : add 2/7 glycerol to cell tubes
 
 
 
- Transformation from glycerol stock from 30/07/2008
 
 
- Spin glycerol stocks, pipette out glycerol
 
 
- Add 0.5mL of medium B, incubate for 1 hour
 
 
- Add 10μL of ECE112 (640ng)
 
 
- Incubate 2hours
 
 
- Plate 200μL, and 10min later, still 200μL
 
 
 
==New stocks==
 
 
- Do glycerol stock of I746001 and I746101 (no sterile glycerol)
 
 
- Put IA751, IA771 in 10mL LB
 
 
- Put ECE 176 in 10mL LB + antibiotic
 
 
- Reinoculate the tube of LB from yesterday with ECE166 plate
 
 
- ECE176 replated onto Amp100 + Cm5
 
  
 
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Revision as of 03:58, 5 September 2008



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