Genetics - cis-Transposition of Bacteriophage Mu
Jeff Lawrence (G31)
1. Make a Mu lysate from an appropriate dilysogen.
2. Grow overnight culture of recipient cells. This can be wild-type E. coli or a mus mutant of S. typhimurium. The recipient should not be expressing the MuC repressor.
- For Mud1 hops, transfect any recipient.
- For MudA hops, transfect a recipient with an amber suppressor.
3. Make two serial 10-fold dilutions of Mu lysate in T2 buffer. Combine 100 µL cells with 100 µL of each lysate dilution in a separate tube. Use 100 (neat), 10-1, and 10-2 dilutions.
4. Incubate at 30° for 60 min.
5. Plate on selective media and grow at 30°.
6. Pick transductants and streak for single colonies on green plates. Grow overnight at 30°.
Last Update: Thursday, 19-Jun-2014 11:54:45 PDT
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7. Choose light colonies and store.