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Nanopore RNA Sequencing Protocol
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===Hardware=== * A computer with a '''strong CPU/GPU running a Unix-based system''' (either ''Linux'' Ubuntu 18+, ''macOS'' 10.11+ El Capitan, or ''WSL''). For this guide, we will assume you are running Linux with a CUDA enabled GPU, but you should be able to follow along on ''Mac'' as well (just know that some of the paths may be different to your files, you may need to download slightly different packages, and your system may process files slower). You can also use Windows via WSL, but I will not be covering how to set that up in this guide. If you wish to do that, [https://learn.microsoft.com/en-us/windows/wsl/install install WSL on Windows as described here] first and then follow the guide as if you were on a ''Linux'' machine. * 30 GB minimum of RAM (although ideally 64+, since 30 GB will be needed ''on top of the available RAM for the OS'' to basecall with ''Guppy''). You can go lower in RAM, but I generally don’t recommend it. * A [https://nanoporetech.com/products/minion MinION Mk1B Nanopore]. * A [https://store.nanoporetech.com/us/flow-cells.html flow cell]. * An [https://store.nanoporetech.com/us/flow-cell-priming-kit.html EXP-FLP002 Flow Cell Priming Kit]. * A [https://store.nanoporetech.com/us/direct-rna-sequencing-kit.html SQK-RNA002 Direct RNA Sequencing Kit]. Additional reagents may be required for the wet lab portion for RNA extraction and preparing the RNA library, but I will cover that in more detail in the Running Samples section below.
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