- Trimming primers with cutadapt
- Installing Docker and QIIME2 for galaxy
- Running QIIME2
- Analyzing paired-end data
- Using DADA2
Trimming primers with cutadapt
In Illumina-based sequencing data,
Installing Docker and QIIME2 for galaxy
- Install Docker desktop:
- Install q2galaxy (GUI for QIIME 2) using the instructions under the Docker heading:
- Start Docker Desktop
- If using an Apple computer with M1/M2 chip, navigate to Settings > Features in development and check the box for “Use Rosetta for x86/amd64 emulation on Apple Silicon”
quay.io/qiime2/q2galaxy(will take 5-10 minutes to download)
- Start the container:
docker run -d -p 8080:80 -p 8021:21 -p 8022:22 -v $HOME/q2galaxy_data/:/export/ quay.io/qiime2/q2galaxy
- It might take a while to spin up…click the container and inspect the Logs to make sure it’s working:
- In a browser, navigate to
http://localhost:8080. You should see this screen if everything is working properly:
Download Docker Desktop | Docker
Docker Desktop is available to download for free on Mac, Windows, or Linux operating systems. Get started with Docker today!
GitHub - qiime2/q2galaxy: Generate Galaxy tool descriptions automatically from QIIME 2 actions.
Generate Galaxy tool descriptions automatically from QIIME 2 actions. - GitHub - qiime2/q2galaxy: Generate Galaxy tool descriptions automatically from QIIME 2 actions.
- In a browser, navigate to https://cancer.usegalaxy.org
- Upload the data to be analyzed (probably
fastq.gzfiles) using the Get Data tool:
Analyzing paired-end data
- Convert the FASTQ data (the raw data that comes off the sequencer) to QZA (the data that QIIME2 likes working with) by using the
qiime2 tools importtool (use the search box to find it).
- For “Type of data to import:” select FeatureData [PairedEndSequence]
fastq.gzfiles that were uploaded should be automatically populated.
- Run the tool:
qiime2 vsearch join-pairsto join paired-end reads together
- Ensure you have R version 4.3.1 installed (download from CRAN if not).
- Open RStudio
- Run the following command:
if (!requireNamespace("BiocManager", quietly = TRUE)) install.packages("BiocManager") BiocManager::install("dada2", version = "3.17")
cutadapt -g TGGTGCCAGCASCCGCG -a GAAASTTXAAGRAATTGACGGA -G TCCGTCAATTYCTTXAASTTTC -A CGCGGSTGCTGGCACCA -n 2 -o 13_R1_001_ca.fastq -p 13_R2_001_ca.fastq 13_R1_001.fastq 13_R2_001.fastq
(replaced I with X)
=== Summary === Total read pairs processed: 130,947 Read 1 with adapter: 84,130 (64.2%) Read 2 with adapter: 46,162 (35.3%) Pairs written (passing filters): 130,947 (100.0%) Total basepairs processed: 54,341,308 bp Read 1: 26,855,924 bp Read 2: 27,485,384 bp Total written (filtered): 52,098,642 bp (95.9%) Read 1: 25,485,477 bp Read 2: 26,613,165 bp