Degenerate DropSynth for Simultaneous Assembly of Diverse Gene Libraries and Local Designed Mutants

Here we introduce an approach which enables the creation of large gene libraries where each library comprises a multitude of diverse, specifically designed genes, each with a corresponding set of programmatically designed fusion points or linkers. Degenerate DropSynth is a multiplex gene synthesis technique which allows for the assembly of up to eight distinct variants for each of the 1,536 parent genes in a single reaction. We assemble four libraries of chimeric sensor histidine kinases and demonstrate the assembly of genes up to 1 kbp in length.