ABSTRACT This supplement is requested to support R01 GM129468: Decoding the non-canonical polyubiquitin chains using chemical approaches. The goal of the parent proposal is to develop new approaches for protein polyubiquitination and subsequent identification and characterization of readers of the noncanonical ubiquitin chains in DNA damage tolerance. The success of this proposal critically depends on the ability to generate modified ubiquitin species that contain reactive linker molecules or unnatural amino acids. Using automated solid-phase peptide synthesis (SPPS) to generate the critical ubiquitin species in sufficiently large quantity will ensure the success of proposed research.