Ligation and gel electrophoresis

Dilute linear DNA into 10µM

Dilute T4 ligation primer into 10µM

Add 11µL of RNase-free water into a eppendorf

Add 3µL of 10X T4 Ligase buffer

Add 9µL of 10µM T4 ligation primer

Add 3µL of 10µM linear DNA

Add 4µL of T4 Ligase

Spin down after vortex

Incubate for 4h 30m 0s at 26Room temperature

Put 2% agarose gel into the electrophoresis tank.

Prepare 0.5X TAE buffer ( add 3.5mL 50X TAE buffer and 350mL ddH2O and mix them), then pour into the electrophoresis tank. Make sure the gel is soaked into the buffer completely.

Take 5µL of marker and load it in the first well.

Take 1µL of 6X loading dye and drop on the parafilm separately, pipetting dye with 5µL of sample

Load samples into each well

Run gel at a constant voltage of 100V for0h 30m 0s at room

temperature

After gel electrophoresis, put the gel into gel reading machine and report the result.