template HpaII TAat adapter TAat adapter+template TAata dapter+template+AseI
template 200ng 200ng 0 200ng 200ng
TAatadapter 0 0 300ng 300ng 300ng
10X T4 DNA Ligase Buffer 5 5 5 5 5
T4DNA Ligase 0 0 1 1 1
MseI 0 1.5 0 1.5 1.5
AseI 0 0 0 0 2.5
Total volume 50 50 50 50 50
Reacted overnight at 37 °C, T4 ligase was inactivated at 65 °C for 20 min Reacted overnight at 37 °C, T4 ligase was inactivated at 65 °C for 20 min Reacted overnight at 37 °C, T4 ligase was inactivated at 65 °C for 20 min Reacted overnight at 37 °C, T4 ligase was inactivated at 65 °C for 20 min Reacted overnight at 37 °C, T4 ligase was inactivated at 65 °C for 20 min Reacted overnight at 37 °C, T4 ligase was inactivated at 65 °C for 20 min