Abstract
Methods of measuring the activities of biomolecules and pathways are fundamental to the study and engineering of biological systems. Traditional marker genes that generate light or color are limited in their dynamic range and often require expensive equipment. Here we describe a way to simultaneously detect the identity and measure the activity of each molecular variant within a mixed population of cells using nanopore sequencing. By linking the desired activity to expression of a base editor that induces mutations in a “canvas” adjacent to the DNA encoding the molecule or pathway of interest, we show that a single long-read sequencing step can retrieve identity and activity at numerous timepoints in bacteria, potentially greatly increasing the available dynamic range. Our technique can replace flow cytometry in directed evolution and may be capable of directly mapping biomolecular fitness landscapes.
Competing Interest Statement
B.T. and K.M.E. are the authors of a provisional patent filed by MIT on the method.