Getting set-up

Before starting

The following protocol is intended to help our collaborators (and ourselves!) navigate through the steps involved in analysis of 16S rRNA sequencing data via QIIME2. We wrote this during and immediately following one of the QIIME2 workshops, incorporating much of the content we learned there. This page is no way meant to be a replacement for the extensive and excellent QIIME2 documentation. Check back often, as we plan to update this protocol as we refine how we use QIIME2 in the lab.

Connect to CHMI linux cluster

<aside> 📌 if you do not have an account on our cluster and would like to get one, please contact Dan Beiting. Accounts are for PennVet only


ssh [email protected]

After you’re connected to our server, activate the QIIME enivronment as follows:

source activate qiime2-2023.5

Prepare your metadata

Before you get started, there are a few basic QIIME commands that you will want to get familiar with

#take a look at all the plugins you have available to you through QIIME
qiime tools

#documentation for any QIIME function ca be accessed by appending --help at the end of any command. For example:
qiime tools --help

Metadata is information about your samples (e.g. date collected, patient age, sex, pH, etc). This information should be contained within a single spreadsheet that has samples as rows and variables as columns.

In order to use QIIME, you must have your mapping spreadsheet correctly formatted. Set this file up as a google sheet, using this example. In order to check whether your file is correctly formatted, use the Keemei plugin for google sheets. Once Keemei says your spreadsheet is correctly formatted for QIIME2, you’re ready to proceed!

Create a .tsv or .txt version of your metadata spreadsheet and transfer it to the server so that it resides in your working directory. You can then inspect further if you want using:

qiime tools inspect-metadata [filename]

Create a visualization of your metadata on the QIIME2 viewer (.qzv is a qiime zipped visualization).

qiime metadata tabulate --m-input-file sample-metadata.tsv --o-visualization tabulated-metadata.qzv

Navigate to QIIME2 viewer in browser to view this visualization

Prepare your raw data