Koeris/Notebook/2007-1-15: Difference between revisions
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=Pseudomonas aeruginosa mer operon cloning 3= | =Pseudomonas aeruginosa mer operon cloning 3= | ||
==Preparation== | ==Preparation== | ||
What | What can I do in order to make sure the cloning works this time? | ||
*Check concentrations of the chromatin DNA | *Check concentrations of the chromatin DNA | ||
*Do a dilution series | *Do a dilution series over three orders of magnitude of the template DNA | ||
==PCR of the operon== | ==PCR set-up of the operon== | ||
#Master mix | #Master mix | ||
##Template DNA equiv. | ##Template DNA equiv. | ||
| Line 13: | Line 13: | ||
##50 deg C - 45s | ##50 deg C - 45s | ||
##72 deg C - 120s | ##72 deg C - 120s | ||
##4 deg C - indefinite | ##4 deg C - indefinite | ||
==Gel image== | ==Gel image== | ||
1% agarose TAE gel, ran @ 100V for 60min | |||
Revision as of 08:27, 15 January 2007
Pseudomonas aeruginosa mer operon cloning 3
Preparation
What can I do in order to make sure the cloning works this time?
- Check concentrations of the chromatin DNA
- Do a dilution series over three orders of magnitude of the template DNA
PCR set-up of the operon
- Master mix
- Template DNA equiv.
- ddH2O to 50uL
- Thermal profile
- 92 deg C - 5min
- 92 deg C - 30s
- 50 deg C - 45s
- 72 deg C - 120s
- 4 deg C - indefinite
Gel image
1% agarose TAE gel, ran @ 100V for 60min
