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Transforming bacteria: Difference between revisions
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Mycoplasma (talk | contribs) (Created page with "Competent cells: 1) Innoculated a 5ml LB culture in 15ml tube with a DH5α colony. Grown overnight in 37°C incubator. Not shaking (might be better if it was). 2) Transferred ...") |
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Competent cells: | ===Competent cells:=== | ||
1) Innoculated a 5ml LB culture in 15ml tube with a DH5α colony. Grown overnight in 37°C incubator. Not shaking (might be better if it was). | 1) Innoculated a 5ml LB culture in 15ml tube with a DH5α colony. Grown overnight in 37°C incubator. Not shaking (might be better if it was). | ||
2) Transferred the 5ml O/N culture into an 'autoclaved' conical flask containing 100ml LB broth and back into the 37°C incubator for about an hour. Then onto hot plate/,magnetic stirrer with a water bath on top to buffer the temperature a bit, it seemed easiest to keep the temperature stable at around 34 to 36°C, not ideal but it worked this time. | 2) Transferred the 5ml O/N culture into an 'autoclaved' conical flask containing 100ml LB broth and back into the 37°C incubator for about an hour. Then onto hot plate/,magnetic stirrer with a water bath on top to buffer the temperature a bit, it seemed easiest to keep the temperature stable at around 34 to 36°C, not ideal but it worked this time. | ||
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11) On this attempt there was no dry ice so snap freezing wasn't possible. Aliquots transferred directly to -20°C freezer!! | 11) On this attempt there was no dry ice so snap freezing wasn't possible. Aliquots transferred directly to -20°C freezer!! | ||
Transformation: | ===Transformation:=== | ||
In this case transformation followed directly after competent cell protocol using aliquots directly from step 10 above. It remains to be seen whether the other aliquots are competent or survived the (non-snap) freezing process. | In this case transformation followed directly after competent cell protocol using aliquots directly from step 10 above. It remains to be seen whether the other aliquots are competent or survived the (non-snap) freezing process. | ||
