Tonight I continued with the Human genome sequencing and analysis course from Nanopore Learning. I watched the video about MinKNOW Configuration for Kit 14. Bala Periaswamy from the Technical Applications Team spoke about setting up a MinKNOW run. They explained that kit 14 has improvements in the pore and motor enzyme. These changes result in higher accuracy. A single PromethION flow cell can be used for detecting variations in a human genome. The MinKNOW software automatically detects the flow cell ID. The kit 14 chemistry is compatible with MinKNOW, Flongle, and PromethION “114” flow cells. Kit 14 is available for rapid and ligation library kits. For multiplexing, rapid and native barcoding kits are available. Periaswamy noted that new kit 14 options would be available this year. For human genome analysis, ONT recommends using at least high accuracy base calling. Methylation calling is optional and encouraged if using native DNA. Fast5 files are now optional if space is an issue. Interestingly, Bam files are recommended even in the absence of alignment, as Periaswamy noted they can be used by EPI2ME workflows. Post-run analysis can be used for basecalling, alignment, and analysis. Simplex and duplex basecalling can be run post-run through the command line. They also noted that GPU-enhanced features can be used via command line. I would like to explore duplex basecalling with datasets we generate in courses.
