High-throughput functional variant screens via in vivo production of single-stranded DNA
A new technoique overcomes existing limitations of CRISPR:
Unlike existing techniques depending on CRISPR-Cas–directed genomic breaks for genome editing, this strategy instead uses single-stranded DNA produced by a retron element for recombineering. This enables libraries of millions of elements to be constructed and offers relaxed design constraints which permit natural DNA or random variation to be used as inputs.
A group of researchers created what they call the "Retron Library Recombineering" (RLR) technique, which could allow scientists to run millions of genetic experiments at the same time. This tool, described in a recent paper in PNAS, employs retrons, which are bacterial DNA segments that undergo reverse transcription to generate single-stranded DNA fragments (ssDNA). RLR produces up to millions of mutations concurrently in bacterial cells and "barcodes" mutant cells, enabling the whole pool to be screened at once. This way large quantities of data can be quickly produced and analyzed.
