The optimized extraction protocol for tissue utilizes pestle homogenization and proteinase K digestion with agitation for sample lysis, followed by a protein removal step and precipitation of the extracted DNA onto the surface of large glass beads. A slightly modified extraction protocol for bacteria utilizes lysozyme for the efficient lysis of the bacterial cell wall, prior to proteinase K digestion. DNA size ranges from 50- ≥500 kb for the standard protocols and into the Mb range when the lowest agitation speeds are used for processing soft organ tissues and bacteria. Purified DNA is recovered in high yield with excellent purity, including nearly complete removal of RNA. For tissue and bacteria, the processing time is 90 min. Purity ratios are typically 1.8-1.9 (260/280) for tissue and bacteria, 2.1-2.5 (260/230) for tissue, and 2.1-2.2 (260/230) for bacteria. Purified HMW DNA is suitable for a variety of downstream applications including long-read sequencing (Oxford Nanopore Technologies® and Pacific Biosciences®), optical mapping (Bionano Genomics®), and linked-read genome assembly.