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Protocol 2.1 - 4 Resuspend each spot in 1 mL of LB 5...

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Kurt Lowery Shane Hodges Ed Stanley BSCI412 0102 Group 9 Protocol 2.1 2.1.1 Preparation of cells for mating 1. Use cotton swabs to collect bacteria ( P. aeruginosa and E. coli ) from plate and add to the appropriate tube with 1 mL of Lysogeny broth. 2. Combine 100 microliters of the aeruginosa tube with 100 microliters of the E. coli tube and pipet up and down to mix. 3. Spot 50 µL of the third tube (the “mating mixture” on an LB plate. Incubate the tube at 37°C for 90 minutes.
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Unformatted text preview: 4. Resuspend each spot in 1 mL of LB. 5. Prepare 10-1 , 10-2 , and 10-3 dilutions and plate 100 µL of each on an LB/Gn/Irg plate. Incubate for 24 hours. 2.1.2 Determination of CFU by Spectroscopy 1. Dilute each bacterial resuspension by 10-1 , 10-2 , and 10-3 in 1 mL of LB and add to cuvette. 2. Blank spectrophotometer using water-filled cuvette, as it has a transparency of 100% 3. Set photometer to 600 nm and determine optical density of each dilution....
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