Difference between revisions of "Team:Paris Bettencourt/Protocols/Chemical test competent cell"

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<b>Chemical test for competent cell</b> <br>
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<b>Chemical test for competent cell</b> <br><br>
  
  
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1) 20-30 sec at 8-10 krpm for the DNA tube (from the kit).
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1) 20-30 sec at 8-10 krpm for the DNA tube (from the kit).<br>
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2) Thow cell competent on ice. Label 2 ml eppendorf µtube for each concentration and put it on ice.
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2) Thow cell competent on ice. Label 2 ml eppendorf µtube for each concentration and put it on ice.<br>
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3) Add 1µl of DNA into each µtube.
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3) Add 1µl of DNA into each µtube.<br>
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4) Add 50µl of competent cell into each tube. Flick gently to mix. Incubate in ice for 30 minutes. Pre-heat waterbath at 42°C.
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4) Add 50µl of competent cell into each tube. Flick gently to mix. Incubate in ice for 30 minutes. Pre-heat waterbath at 42°C.<br>
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5) Heat-shock cell by placing in waterbath 1 minute. CAUTION : The top need to be close to the water level, but not immerge in.
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5) Heat-shock cell by placing in waterbath 1 minute. CAUTION : The top need to be close to the water level, but not immerge in.<br>
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6) Keep back the tube to ice for 5 minutes.
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6) Keep back the tube to ice for 5 minutes.<br>
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7) Add 200µl of SOC media, incubate 37°C for 2h. Prepare agar plate of the media that you want, and label it (3 plate for each concentration).
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7) Add 200µl of SOC media, incubate 37°C for 2h. Prepare agar plate of the media that you want, and label it (3 plate for each concentration).<br>
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8) Add 20µl of each tube n  the appropriate plate. Incubate 37°C for OVN (12-16h).
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8) Add 20µl of each tube n  the appropriate plate. Incubate 37°C for OVN (12-16h).<br>
  
  
  
 
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Latest revision as of 14:01, 3 September 2015

Chemical test for competent cell

1) 20-30 sec at 8-10 krpm for the DNA tube (from the kit).
2) Thow cell competent on ice. Label 2 ml eppendorf µtube for each concentration and put it on ice.
3) Add 1µl of DNA into each µtube.
4) Add 50µl of competent cell into each tube. Flick gently to mix. Incubate in ice for 30 minutes. Pre-heat waterbath at 42°C.
5) Heat-shock cell by placing in waterbath 1 minute. CAUTION : The top need to be close to the water level, but not immerge in.
6) Keep back the tube to ice for 5 minutes.
7) Add 200µl of SOC media, incubate 37°C for 2h. Prepare agar plate of the media that you want, and label it (3 plate for each concentration).
8) Add 20µl of each tube n the appropriate plate. Incubate 37°C for OVN (12-16h).