Difference between revisions of "Team:Aix-Marseille/Experiments/Protocols"

(Preparation of competent bacteria cells)
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=== Preparation of competent bacteria cells ===
 
=== Preparation of competent bacteria cells ===
  
1. Grow a culture of bacteria in LB medium until it reaches OD600 = 0.5.
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#Grow a culture of bacteria in LB medium until it reaches OD600 = 0.5.
  
2. Centrifuge cells for 10 minutes at 3500g at 4°C
+
#Centrifuge cells for 10 minutes at 3500g at 4°C
  
3. Resuspend the pellet slowly in 80mL of Tfb1 buffer.
+
#Resuspend the pellet slowly in 80mL of Tfb1 buffer.
  
4. Centrifuge 5 min at 3500g at 4°C.
+
#Centrifuge 5 min at 3500g at 4°C.
  
5. Resuspend the pellet in 8 mL of Tbf2 buffer.
+
#Resuspend the pellet in 8 mL of Tbf2 buffer.
  
6. Incubate 15 min in ice.
+
#Incubate 15 min in ice.
  
7. Aliquot 200µL of cell suspension in sterile eppendorf tubes and conserve it at -80°C.
+
#Aliquot 200µL of cell suspension in sterile eppendorf tubes and conserve it at -80°C.

Revision as of 13:12, 10 June 2016

Preparation of competent bacteria cells

  1. Grow a culture of bacteria in LB medium until it reaches OD600 = 0.5.
  1. Centrifuge cells for 10 minutes at 3500g at 4°C
  1. Resuspend the pellet slowly in 80mL of Tfb1 buffer.
  1. Centrifuge 5 min at 3500g at 4°C.
  1. Resuspend the pellet in 8 mL of Tbf2 buffer.
  1. Incubate 15 min in ice.
  1. Aliquot 200µL of cell suspension in sterile eppendorf tubes and conserve it at -80°C.