General:This is an DNA isolation protocol adapted to the properties of sludge samples from biogas and wastewater treatment plants. The protocol reliably retrieves metagenomic DNA of sufficient quality, length and yield. The performance has been validated on 20 different reactor plants. Observed Performance on 20 ONT runs using this protocol:It does not negatively impact the overall throughput for Nanopore Sequencing. Sequencing runs were active over 2.5 days, with one refuel step after 18 h.N50 Ranges between 4000 and 8000 bp, depending on the sample type. E.g. frozen samples have a lower N50, fresh samples a higher N50. General throughput is between 18-28 Gigabases of a single flowcell (flowcell 106 Rev D, using the LSK-109 Kit). Funding:This work was funded by the Deutsche Forschungsgemeinschaft (DFG, German Research Foundation) - BR 5692/1-1 and BR 5692/1-2