Component	Ligation	Control
3ZF GNH/ANN vector	600 ng	200 ng
2ZF GNH/ANN insert	900 ng	-------
5× T4 DNA Ligase Buffer	48 μL	16 μL
T4 DNA Ligase	12 μL	4 μL
Nuclease-free water	to 240 μL	to 80 μL
Total	240 μL	80 μL