INgen: Intracellular Genomic DNA Amplification for Downstream Applications in Sequencing and Sorting

Here, we introduce intracellular genomic amplification (INgen), a method that harnesses the cell membrane as a natural reaction chamber for DNA amplification, enabling downstream sequencing and cell sorting. Unlike traditional single-cell techniques, INgen utilizes a strand-displacing, isothermal polymerase to amplify DNA within fixed, permeabilized cells while maintaining the cell’s structural integrity. This approach overcomes challenges associated with both typical single-cell DNA sequencing and hindrances encountered when previously attempting to sequence genetic material from fixed microbial cells, allowing amplification of genomic regions up to 100 kb and sequencing of whole genomes from diverse cell types, including S. cerevisiae, B. subtilis , and E. coli . Additionally, INgen can be adapted for targeted DNA enrichment using biotinylated primers and for fluorescence-based cell sorting.