Editing Antibody Design by AI (section)

== Oligo synthesis of CDR-H3 sequences and library generation ==

We constructed a library including 9,553 sequences proposed by ensemble gradient ascent (Ens-Grad). 5,467 of these sequences were optimized from denoised seeds and thus were included in the analysis to avoid over-estimation of the performance. We also included 9,503 sequences proposed by VAE, 17,813 sequences proposed by GA-KNN, 6,286 sequences proposed by GA-CNN, and 23,606 sequences designed for other purposes and thus not described in this paper, summing up to 64,984 machine learning derived sequences. Here the number is the total number of sequences proposed by each method even though there were overlapping sequences between different methods that are only counted once when calculating the library size. We also included 39,541 control sequences consisting of 12,243 seed sequences defined as previously described and 27,298 controls which contained both negatively-enriched sequences from our panning experiment (negative control) as well as additional sequences generated to receive negative predictions from our neural network ensemble. Collectively we built a library of 104,525 CDR-H3 variable region sequences. We concatenate the sequences with the framework and restriction sites used for cloning the sequences into phages. We translated the amino acid sequences into DNA nucleotide sequence using EMBOSS 26 and the E.coli codon usage table was used. The nucleotide sequences were then used to synthesize a '''DNA oligo pool (Twist Bioscience)'''. Library inserts were generated by PCR using Phusion High Fidelity DNA polymerase (NEB Biolabs). The resulting CDR-H3 library inserts were ligated into the base template and transformed into E.coli '''TG1 DUO '''(Lucigen) aiming for a library size of 5E+08 transformants. Phages were produced using '''VSCM13 '''helper phage (Agilent) using standard previously described protocols 23.
[[Category:Antibodies]]