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Team:Groningen/Protocols
From 2010.igem.org
(Difference between revisions)
(New page: CaCl2 Competent Cells '''Day 1:''' -Inoculate 5 ml LB( TY) medium with ''E.coli'' from glycerol stock -Grow overnight at 37 oC '''Day 2:''' -Inoculate 20 ml LB medium with 200 ...) |
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'''Day 1:''' | '''Day 1:''' | ||
| - | -Inoculate 5 ml LB( TY) medium with ''E.coli'' from glycerol stock | + | -Inoculate 5 ml [[LB( TY) medium]] with ''E.coli'' from glycerol stock |
-Grow overnight at 37 oC | -Grow overnight at 37 oC | ||
| Line 15: | Line 15: | ||
-Spin down 5 minute at 4000 rpm at 4 oC | -Spin down 5 minute at 4000 rpm at 4 oC | ||
| - | -Resuspend in 10 ml chilled 0,1 M | + | -Resuspend in 10 ml chilled 0,1 M CaCl2 (from here, keep on ice! ) |
-Inoculate on ice for 20 minutes | -Inoculate on ice for 20 minutes | ||
Revision as of 09:53, 24 August 2010
Day 1:
-Inoculate 5 ml LB( TY) medium with E.coli from glycerol stock
-Grow overnight at 37 oC
Day 2:
-Inoculate 20 ml LB medium with 200 μl overnight culture
-Grow at 37 oC until OD600 = 0,3-0,5(+/- 2 hours)
-Spin down 5 minute at 4000 rpm at 4 oC
-Resuspend in 10 ml chilled 0,1 M CaCl2 (from here, keep on ice! )
-Inoculate on ice for 20 minutes
-Spin down as before
-Remove supernatant
-Resuspend in 2 ml 0,1 M CaCl2 /10% glycerol
-Divide 100 μl aliquots
-Store competent cells in - 80 oC
