The Construction of dsDNA-Based AND-Gate (DBAG) Peptide Library in Mammalian Cells

The construction of DNA-encoded peptide libraries in mammalian cells is an evolving field that combines the advantages of mammalian cell expression systems with the vast diversity of DNA-encoded libraries. This enables high-throughput screening and identification of functional peptides in a mammalian cell context, aiding in the elucidation of disease-related proteins and the development of novel peptide drugs. We have developed a strategy for constructing dsDNA-based AND-gate (DBAG), which utilizes dsDNAs with NNK degenerate codons at the ends to implement AND-gate genetic circuits and generate peptide libraries in mammalian cells. We provide a detailed description of the library preparation, transfection, and analysis methods.