Ligation

Materials:

• MQ water
• 1.5ml tubes
• Ligation buffer
• Ligase
• Insert
• Vector

Reaction:

Component	20µl	Final concentration
MilliQ water	up to 20µl
10 ligation buffer	2µl	1x
Vector	xµl	20-100ng
Insert	xµl	3:1 molar ratio over vector
T4 DNA ligase 5U/µl	2µl	0.5U/µl

All the reagents are added following the order listed in the table above.
After the reaction is ready mix the content of the tube and spin it down.
The tubes are incubated for at room temperature (~22°C) for 1h.

Reference: http://www.thermoscientificbio.com/uploadedFiles/Resources/fast-digestion-dna.pdf