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Team:Newcastle/Transformation of E. coli
From 2010.igem.org
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| - | === | + | =Transformation of ''E. coli''= |
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# Thaw a 200 µl aliquot of the desired strain of ''E. coli'' and add the transforming DNA (10 ng of vector DNA in 10 µl). Incubate for 45 min at 42°C. | # Thaw a 200 µl aliquot of the desired strain of ''E. coli'' and add the transforming DNA (10 ng of vector DNA in 10 µl). Incubate for 45 min at 42°C. | ||
# Heat-shock the cells for 120 secs, and place on ice again for 3-4 min. | # Heat-shock the cells for 120 secs, and place on ice again for 3-4 min. | ||
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# Plate out 100-200 µl/plate on LB (agar at 1.5%), containing the appropriate selection markers. | # Plate out 100-200 µl/plate on LB (agar at 1.5%), containing the appropriate selection markers. | ||
# Incubate plates overnight at 37°C. | # Incubate plates overnight at 37°C. | ||
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Revision as of 14:56, 28 July 2010
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Transformation of E. coli
Procedures
- Thaw a 200 µl aliquot of the desired strain of E. coli and add the transforming DNA (10 ng of vector DNA in 10 µl). Incubate for 45 min at 42°C.
- Heat-shock the cells for 120 secs, and place on ice again for 3-4 min.
- Add 1ml of LB broth and incubate the cells at 37°C for 1-15 hr in a water bath with gentle shaking.
- Plate out 100-200 µl/plate on LB (agar at 1.5%), containing the appropriate selection markers.
- Incubate plates overnight at 37°C.
   
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