Silver: Ligation: Difference between revisions
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#*2 µL tube #2 (DNA dilution buffer) | #*2 µL tube #2 (DNA dilution buffer) | ||
#*distilled water to final volume of 10 µL total volume | #*distilled water to final volume of 10 µL total volume | ||
#Add 10 µL of freshly mixed tube #1 (T4 DNA ligase buffer) | #Add 10 µL of freshly mixed tube #1 (2x T4 DNA ligase buffer) or 2uL of 10x T4 DNA ligase buffer | ||
#Add 1 µL tube #3 (T4 DNA ligase). | #Add 1 µL tube #3 (T4 DNA ligase). | ||
#Mix well, incubate for 15 min. at room temperature. | #Mix well, incubate for 15 min. at room temperature. | ||
#Proceed to [[Silver: Bacterial_Transformation|transformation]]. Store excess ligation mixture at -20 °C. | #Proceed to [[Silver: Bacterial_Transformation|transformation]]. Store excess ligation mixture at -20 °C. | ||
Revision as of 07:02, 30 June 2010
Use Roche's Rapid DNA Ligation Kit, stored at -20 °C.
Keep the total mass of DNA below 200 ng.
- Mix:
- Add 10 µL of freshly mixed tube #1 (2x T4 DNA ligase buffer) or 2uL of 10x T4 DNA ligase buffer
- Add 1 µL tube #3 (T4 DNA ligase).
- Mix well, incubate for 15 min. at room temperature.
- Proceed to transformation. Store excess ligation mixture at -20 °C.
