11/18/98: Protocol from Sandra Diaz, bugs from Ling (in Varki Lab).

1. Thaw bugs (E. coli) on ice.

2. Aliquot 100µl cells into pre-chilled 1.5 ml tube. Put excess bugs back into the -70 freezer.

3. Add DNA (1 to 5 µl), swirl tube, incubate on ice for 20 minutes.

4. Heat shock the cells at 42°ree;C fo 40 seconds.

5. Add 1ml SOC; (then transfer to larger tube).

6. Rescue at 37°ree;C for 1 hour, shaking.

7. Spread 50µl, 100µl, (& remainder spun down if difficult ligation) on (LB+Amp ?) plates, --> 37°ree;C ON.

INFO: DH5-alpha bugs ...

Shirley Reynolds