Ethyl Methane Sulfonate Lab Report

Experiment 2
The induction of mutations in Escherichia coli by ethyl methane sulfonate and in Salmonella typhimurium by Tn10
The purpose of this experiment was to identify and isolate ethyl methane sulfonate (EMS) mutagenized colonies of Escherichia coli (lac-) which could no longer use lactose as a carbon source and to isolate Tn10 mutagenized colonies of Salmonella typhimurium which were induced auxotrophs and identify independent mutations.
Results
Mutants observed in Escherichia coli:
Control 10-5 Dilution: 0/440
Control 10-6 Dilution: 0/310
Mutagenized 10-5 Dilution: 9/339
Mutagenized 10-6 Dilution: 23/ 403 A hypothesized E.coli mutant colony (opaque and white colony) from the EMS mutagenized plate was streaked on a new MacConkey’s
Typh = 0%
Mutagenized S. Typh = 10% Mutagenesis in S. typh was successful because mutants were observed in the mutagenized culture plates and not in the control plates. This indicates that the Tn10 exposure was the cause of mutant variation. There were ten hypothesized auxotrophic colonies based on the comparison of mutagenized colonies in minimal and nutrient agar. These ten colonies were confirmed to be auxotrophic by their lack of growth in the minimal agar replica plate. The required growth factors for each auxotroph were determined based on which pool plates fostered growth for each streak. Auxotrophs 1, 2, 3, and 4 require valine as a growth factor since they grew on pool 4 and pool 7 plates only. Auxotrophs 6, 7, and 8 require proline as its groth factor since they grew on pool 5 and pool 8 plates only. Auxotrophs 5, 9, and 10 showed unexpected results. The expected result was growth in two of the pool plates, no growth in the minimal agar, and growth in the nutrient agar. Auxotroph 5 grew in six of the pool plates. This may be due to error in transferring the colony –cells from more than one auxotrophic colony may have been streaked simultaneously. Auxotroph 5 may also be a prototroph that was insufficiently transferred to the other pool plates. Auxotrophs 9 and 10 showed no growth in any pool plates, but had growth in the control plate, so inability/inadequate to transfer the colonies to the pool plates is unlikely. It may be that auxotrophs 9 and
If the auxotroph grows on one plate only (the nutrient agar plate), then that auxotroph may require some growth factor absent from all of the pool plates or some combination of growth factors not provided by any one pool plate. If it grows on one pool plate only, it may require a combination of two growth factors only available on one of the pool plates.
An auxotroph that grows on three or four plates may be a mixed culture of more than one