Third generation sequencing aims to sequence long DNA molecules. High-quality and high-molecular-weight DNA is needed to fully benefit from the potential of third-generation sequencers. The cost of sequencing continues to decrease but DNA extraction kits remain expensive. Here is a fast and cheap protocol to extract genomic DNA from Gram-negative bacteria. The original protocol has been fine-tuned for Xanthomonas ssp., which is a tricky bacteria as it secrets a lot of extracellular polysaccharides. We found that bacterial culture can impact the purity of the final DNA solution. In addition, the lysis temperature and mixing by pipetting are critical to get an efficient homogenization as the sample becomes very viscous at some steps (see 'Assessment of various parameters.xlsx' attached below).High yield Nanopore runs can be achieved with these gDNA samples (see 'Nanopore sequencing report.pdf' attached below).Acknowledgements:I would like to thank Harihar Jaishree Subrahmaniam,\u00a0Rekha Gopalan Nair, and BenjaminSchwessinger for their feedbacks and their corrections on this protocol.