Bacterial Transformation Lab
Bacterial Transformation Lab
Introduction: In this experiment we transformed a strain of E. Coli bacteria without antibiotic resistance with plasmid DNA. This plasmid produces a fluorescent green glow under black light due to the gfp(green fluorescent protein) as well as antibiotic resistance. E. Coli cells will be plated on an agar medium, some with and some without the antibiotic ampicillin. Only bacterial cells that contain the plasmid will survive the ampicillin and produce the green glow. This experiment allowed us to observe the process of bacterial transformation. I believe that only a small percentage of the cells will transform and the gfp plasmid will be most apparent in the agar plate containing both the plasmid and ampicillin. …show more content…
Coli colonies into each of the tubes using a sterile toothpick
Add .10ml of the gfp plasmid to the tube labeled “DNA+”
Incubate the tubes on ice for 15 minutes place both tubes into a 42 degree hot water bath for 90 seconds allowing the heat shock to let the plasmid DNA enter the cells
Immediately return both tubes to the ice bucket and incubate for2 minutes
Using a sterile pipette, add .25ml of luria broth to each tube and mix
Incubate cells in a 37 degree water bath for 30 minutes to recover
Using a sterile pipette, transfer .25ml of the “DNA-” mixture to the plate labeled LB- and LB/amp- Spread cells over the entirety of each plate then cover both plates
Use a sterile 1ml pipette to transfer .25ml of the “+DNA” to the middle of the LB+ and LB/amp+ …show more content…
As expected, neither of the gfp negative plates showed any trace of glowing and the ampicillin positive plate showed no bacterial growth at all. The gfp positive plate showed large amounts of bacterial growth, but as stated in the hypothesis, the regular bacteria greatly outgrew the gfp positive bacteria and no glow was observed. The gfp/ampicillin positive plate showed almost no growth at all. We had expected to see only gfp glowing bacteria, but so little bacteria had grown before the ampicillin had killed the normal bacteria, it was almost impossible to see even the faintest
Introduction: In this experiment we transformed a strain of E. Coli bacteria without antibiotic resistance with plasmid DNA. This plasmid produces a fluorescent green glow under black light due to the gfp(green fluorescent protein) as well as antibiotic resistance. E. Coli cells will be plated on an agar medium, some with and some without the antibiotic ampicillin. Only bacterial cells that contain the plasmid will survive the ampicillin and produce the green glow. This experiment allowed us to observe the process of bacterial transformation. I believe that only a small percentage of the cells will transform and the gfp plasmid will be most apparent in the agar plate containing both the plasmid and ampicillin. …show more content…
Coli colonies into each of the tubes using a sterile toothpick
Add .10ml of the gfp plasmid to the tube labeled “DNA+”
Incubate the tubes on ice for 15 minutes place both tubes into a 42 degree hot water bath for 90 seconds allowing the heat shock to let the plasmid DNA enter the cells
Immediately return both tubes to the ice bucket and incubate for2 minutes
Using a sterile pipette, add .25ml of luria broth to each tube and mix
Incubate cells in a 37 degree water bath for 30 minutes to recover
Using a sterile pipette, transfer .25ml of the “DNA-” mixture to the plate labeled LB- and LB/amp- Spread cells over the entirety of each plate then cover both plates
Use a sterile 1ml pipette to transfer .25ml of the “+DNA” to the middle of the LB+ and LB/amp+ …show more content…
As expected, neither of the gfp negative plates showed any trace of glowing and the ampicillin positive plate showed no bacterial growth at all. The gfp positive plate showed large amounts of bacterial growth, but as stated in the hypothesis, the regular bacteria greatly outgrew the gfp positive bacteria and no glow was observed. The gfp/ampicillin positive plate showed almost no growth at all. We had expected to see only gfp glowing bacteria, but so little bacteria had grown before the ampicillin had killed the normal bacteria, it was almost impossible to see even the faintest