Protocol

Short Name

7  

Full Name

7  

Type

cloning  

Lab

University of Toronto  

Description

Fermentas /rxn | 10 x T4 Ligase buff. 2?l *Add 6.5?l cleaned digestion product | cut vector 1?l *Ligate: 4hrs. to O/N @ 14?, or 1 hr. @ rm. temp. | ligase (1U/?l) 0.5?l Escherichia coli DH5a hit shock 80ul of cells is combined with ligation reaction, or 'in-fusion' reaction depending on the vector | incubated on ice for 10 min | heat shocked at 42 degrees for 45 seconds | incubated on ice for 2 min | 1 hour recovery in SOC @ 37 degrees with shaking | spin down @3000 rpm, pour off all but ~50ul | resuspend and plate pellet on LB + amp plates SOC 1