The protocol that Hunter was following was simple and iterative. He incubated twenty sets of the cultures for 20 generations which increased the population by a factor of a million. Then he subjected ten of the twenty cultures to a coarse “ladder” where he added varying amounts of bacitracin, an antibiotic and an additional dose of nutrient.
The antibiotic did not kill the bacteria. Rather, it interferred with the bacteria’s ability to reproduce. Consequently, it took a week to determine the cut-off level where one concentration stopped the culture and the lesser concentration did not.
Hunter then used the remaining ten cultures to “dial in” the exact concentration between the two coarser steps to determine the log 4 concentration where the antibiotic killed 9999 of every 10,000 bacteria.
After determining that concentration he increased the concentration of antibiotic in the viable cultures that had received the lower, non-inhibited dosages.
He was able to determine existence of fresh generations with polarized light. The older cells were not motile and sank to the bottom of the bottles. Hunter rarely needed to decant a bottle.
After a second incubation period Hunter chose the culture with the highest number of motile bacteria and pitched the rest.
Lather, rinse, repeat.
The backstory given by the protocol sheet informed Hunter that antibiotic resistance was being bred into the strain of bacteria so antibiotics could be added to the reaction vessels and suppress other bacteria that might contaminate the run.
Croyle was impressed by Hunter’s energy and ingenuity. He had stalled out in finding another biologist.
Croyle asked Hunter if he though he could take on more work. Hunter looked at the water bed mattress and figured he had room for at least two more cultures.
Croyle sent two more cultures and specified that one be bred for resistance to Neomycin and the other be bred for resistance to Polymyxin B.
The package also had a pre-paid with $3000 on it with the promise of another one in a week if Hunter’s work continued to show promise.