Troubleshooting Test
To determine how the frequency of transformation could best be increased, and thus reduce the impact of error, stock competent E. coli cells were transformed in the manner described above, with 1無 or 5無 of self-isolated pBluescript, or 1無 of stock pBluescript. Each was done with and without heat shock (2 minutes at 42°C, followed by 2 minutes at 0°C, before 37°C incubation), and 90無 of each sample were plated on a stock and a self-made amp+ LB agar plate.
Results
The same number of bacteria are assumed to have been exposed to transforming conditions for each condition tested.
Discussion
These results indicate that 1無 of stock pBluescript with no heat shock produces the most transformants, likely because the concentration of stock pBluescript is higher than that of the self-isolated pBluescript. More transformants are evident on stock plates than on self-made plates, probably because the stock plates were poured more thickly.38 While enough stock plates were not available for use, enough stock pBluescript was available for use in the final experiment. A greater volume of frozen competent cells was also used in the final experiment, to increase the number of transformants.
