Erickson:Tri-parental Mating: Difference between revisions
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Bria Walch (talk | contribs) (New page: Tri-parental Mating 1. Grow the donor, helper, and recipient strains of bacteria separately in 3 mL of LB Broth overnight (30° C for Y. pestis and 37°C for E. coli) 2. Combine 200 µL of...) |
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Tri-parental Mating | *Tri-parental Mating | ||
Grow the donor, helper, and recipient strains of bacteria separately in 3 mL of LB Broth overnight (30° C for Y. pestis and 37°C for E. coli) | |||
Combine 200 µL of the donor, helper, and recipient strains into one centrifuge tube | |||
Centrifuge and remove supernatant | |||
Re-suspend pellet in 25 µL broth | |||
Spot onto BHI plate (no antibiotics) | |||
Incubate overnight at 30°C | |||
Scrape a colony from plate and re-suspend in broth | |||
Dilute each solution to 10^-1 | |||
Plate 100 uL of each solution onto a BHI plate with kanamycin and irgasan | |||
Incubate overnight at 30°C | |||
Latest revision as of 15:44, 24 April 2009
- Tri-parental Mating
Grow the donor, helper, and recipient strains of bacteria separately in 3 mL of LB Broth overnight (30° C for Y. pestis and 37°C for E. coli)
Combine 200 µL of the donor, helper, and recipient strains into one centrifuge tube
Centrifuge and remove supernatant
Re-suspend pellet in 25 µL broth
Spot onto BHI plate (no antibiotics)
Incubate overnight at 30°C
Scrape a colony from plate and re-suspend in broth
Dilute each solution to 10^-1
Plate 100 uL of each solution onto a BHI plate with kanamycin and irgasan
Incubate overnight at 30°C
