Team:Macquarie Australia/Protocols/Ligation

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Ligation Procedure

Element Volume
Upstream Digestion part 2 µL
Downstream Digestion part2 µL
Destination Plasmid1 µL
10X T4 DNA ligase buffer2 µL
T4 DNA ligase1 µL
H2O12 µL


A reaction mixture was prepared containing, Element Volume Upstream Digestion part 2 µL Downstream Digestion part 2 µL Destination Plasmid 1 µL 10X T4 DNA ligase buffer 2 µL T4 DNA ligase 1 µL H2O 12 µL




We incubated each ligation mix at 30°C for 30 minutes, followed by heat inactivation at 80°C for 20 minutes.

4 µL of the ligation product was transformed into 100 µL of competent E. coli, the transformants were then incubated for one hour.

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