Question

You have conducted a transposon mutagenesis experiment using the same pRL27 system that was used during your lab exercise. After allowing for conjugation. you plate the conjugation mix on Luria agar and Luria + Kanamycin agar. After incubation, you count the following number of colonies on each type of plate:

Media Dilution Number of Colonies
Luria 10-5 Too Many to Count
10-6 251
10-7 28
Luria + Kanamycin 100 324
10-1 98
10-2 3

Determine the transformation efficiency. Note: You can enter you answer using long form (e.g. 1500000) or scientific notation using e notation (e.g. 1.5e6). Do not enter units.

Answer 1

The transformation efficiency for the experiment using Luria + Kanamycin agar plates with a dilution factor of 10-2 and 3 counted colonies is 3000 colonies/mL, considering the volume plated and the dilution factor.

Step-by-step explanation:

To determine the transformation efficiency, we need to consider the number of colonies that grew on the Luria + Kanamycin agar plates since these have the selective marker to show which E. coli cells were transformed by the plasmid containing kanamycin resistance from the pRL27 system used in the experiment. The dilution factor that gives an accurate count of colonies (neither too many nor too few) needs to be used for this calculation. According to the given data, the dilution factor 10-2 with 3 colonies appears to be such since the plates with higher dilution factors had too many colonies (which could have resulted in colonies merging and thus not giving an accurate count).

Using the 10-2 dilution factor, where 3 colonies were counted, and acknowledging that only 0.1 mL was plated, the transformation efficiency can be determined as follows:

• First, correct the colony count for the volume plated, which is 3 colonies × 10 (since 0.1 mL was plated) = 30 colonies/mL.
• Next, account for the dilution factor: 30 colonies/mL × 10² (the dilution factor) = 3000 colonies/mL.
• This is the number of colonies that would be present in a milliliter of the original undiluted transformation mixture.

The colony count obtained for the non-selective Luria plates is too high to provide reliable data for the calculation of transformation efficiency, so it is not considered in this calculation.