A codon optimizer for phage display library generation.

When making phage display libraries, it's easy to run out of permutations. 1 liter of phage solution can hold about $10^{12}$ different sequences.

We can approximate how many combinations we generate with $(nr\ of\ choices)^{(nr\ of\ positions)}$. If we change the aminoacids to "any" aminoacid, we can change about 9 AA's: $20^9 \approx 10^{12}$ permutations.

If we use the degenerate codon capabilities of the manufactureres such as IDT to the fullest, we can probably make combinations that are partially rational, and allow for 6 permutations in any postion.

That allows us to check out many more combinations that are likely to work: $6^{15} \approx 10^{12}$, so about 15 variable AA's instead of 9.

This tool allows you to easily specify what AA permutations you want, in which position, and calculates the best sequence of degenerate codons.

Phagetrix is a tool to generate phage display libraries. You probably have an idea what AA's you want to replace, and what you want to replace them with. The sequence companies have a reasonable number of The file format has the AA sequence on the first line. Each following line is the AA to be changed, the digits indicating its position, and the AA options that should be generated for that position. The AA options are concatenated together with no spaces. Phagetrix will generate the best degenerate codon for each position.

Example:

VLAYMVAQVQ
A3AGVIL
Y4YFW
A7AVIL

1. The first line is the sequence you want to alter.
2. The P in position 3 should be either a P, F, Y or an A
3. The Y in position 4 should be either a Y, P, F, or an E
4. The A in position 7 should be either an A, V, I, L or an M

Output:

phagetrix ./sample.phagetrix
   1   2   3   4   5   6   7   8   9  10
   V   L   A   Y   M   V   A   Q   V   Q
 GTT CTT VBA TDK ATG GTT VYA CAG GTT CAG   degenerate codons
          56  50          67               percentage on target
  1V  1L  1V  1Y  1M  1V  1V  1Q  1V  1Q
          1L  1W          1L
          1I  1F          1I
          1G  --          1A
          1A  1L          --
          --  1C          1T
          2R  1*          1P
          1T
          1P

GTTCTTVBATDKATGGTTVYACAGGTTCAG

The lines of the output show:

1. number of the AA on the first line
2. original AA on the second line
3. codon made from degenerate basepairs
4. the percentage of product for this codon that matches what the user specified
5. how many codons code for which aminoacid. The AA below the -- line are off-target codons that were not requested.

• the last line shows the codons made from degenerate basepairs again, in a format that can easily be copied and pasted.

By default this uses the degenerate codons from IDT.

Adding a comment of this form to your input file (see the pal.phagetrix file in the example directory)

# offset = 20

allow the setting of variables - in this case, the first aminoacid will be numbered 21. This is nice for long sequences where "something in the middle" needs to be edited.

• 
• Install it on your machine pip install phagetrix to get the phagetrix command line tool. Requires python3.

Other projects that are potentially interesting, if you are into this sort of stuff.

• Make primers for highly variable genomes