Percentage base pair optimization algorithm

1. For each base of the complement of the context containing the RBS, create three other permutations which have that base mutated. All of these permutations combined form the initial population. The baseline is defined as the full sequence containing the context before the variable region, the variable region which is the complement of the RBS-containing context, and the RBS-containing context.

2. Calculate the base pairing probabilities at the two given temperatures for each test sequence using the NuPack command pairs -T TEMP -pseudo -material rna sequencename.

3. The output of the command consists in part of a list of every base and the base(s) which it has a > 0.001 probability of base pairing with by position number. Find the base pairings where one of the bases is in the RBS-containing region and add up the probabilities corresponding to that base pairing. The nature of this NuPack command should prevent duplicates. Then, subtract the probability that these bases are unpaired.

4. Subtract the number of base pairings at 30°C from the number of base pairings at 25°C. We want to maximize the number of RBS-base pairings that disappear as the temperature increases from 25°C to 30°C (for reference, the command I use for the minimum free energy calculation is mfe -T TEMP -pseudo -material rna sequencename)

5. Using the library DEAP, select 50 individuals through the tournament selection method.