Molecular multiplexing methods for genome-scale measurements.

University of Minnesota Ph.D. dissertation. June 2018. Major: Plant and Microbial Biology. Advisor: Peter Tiffin. 1 computer file (PDF); ix, 106 pages.I present the utility of unique DNA barcodes to tag distinct genotypes and subsequently link them to phenotypes. Such molecular tagging allowed us to perform multiplexed phenotype analysis of thousands of genotypes using next-generation sequencing (NGS) technologies. Four projects are discussed in this thesis - 1) 2D Tn-Seq, a massively multiplexed experimental approach to interrogate genetic interactions of a microbe at the genome scale 2) pFluxSeq, a molecular tool that will enable peptide-based 13C metabolic flux analysis (MFA) of a mixed population of microbial cells 3) Deep mutational scanning of phenotype arrays and 4) Barcode-based lineage tracking to measure CRISPR/dCas9 RNA interference efficacy in bacteria