The tools fastq-dump and fasterq-dump are used to extract reads from the Sequence Read Archive and export them to (for example) fastq format. There is a hidden gotcha that you should be aware of using fastq-dump to extract data.
Category Archives: Lab blog
No python in CentOS 8
CentOS 8 does not include a command that responds to python! Here are some solutions to this!
Getting WGS data from GenBank via the SRA
Sometimes when you look at a record in RefSeq/GenBank it is a virtual record that is really a pointer to a set of records. For example, the entry for Callorhinchus milii isolate IMCB2004 points you to the WGS records AAVX02000001-AAVX02067420. Here we show how to get these records.
Accessing SRA in the Cloud
As part of the STRIDES initiative, the NIH has moved the SRA to the cloud. This includes the metadata, and the whole SRA archive. Here, I show how to set up a new instance to access the sequence read archive in the cloud. In a separate post, we’ll explore getting the metadata out of bigtable.
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New features in CentOS8
We have just made the transition of most of the servers from CentOS6 or CentOS7 to CentOS8. Most everything should be unified on CentOS8 (unless you know what you are doing).
This brings several new changes (as always) and some added benefits. This is a summary and does not reflect all the changes.
To check your servers operating system version, use this command:
cat /etc/redhat-releaseSoftware Installs
The biggest changes should allow you to install software by yourself! There are two different ways you can install easily install software if either are supported by whatever you are trying to install.
Please note, that if you do not want to do either of these, it is fine. Just let me know and I am happy to install software for you (and everyone else) to use.
Conda
A lot of bioinformatics software is now available via conda. It is installed globally, but you can not install packages globally. You can create your own environment and then use that.
The first time you use conda, you will need to create a local environment. Start with:
source /usr/local/anaconda3/bin/activateconda create --name <username>
But use your username instead of <username>!
After this has run, any time you need to use conda, you can use the command
conda activate <username>And you will get into your environment.
A simple test is to install my fastq-pair package and see if it works:
conda install -c bioconda fastq-paironce it has installed, this command should give some output
fastq-pairDocker
Another popular way of sharing software is by using docker. We don’t support docker, but we support a drop-in replacement called podman.
Anywhere you see docker, you can use podman instead. For example, we created a focus docker image for the cami challenge described here: https://hub.docker.com/r/linsalrob/cami-focus and you can install that with
podman pull linsalrob/cami-focuspip
If you are trying to run some python code and don’t have the appropriate library, you should be able to use pip install as a user to add it. For example:
pip3 install --user xmlschemathis will install the appropriate libraries into your account. Of course, if you want them globally installed, just let me know.
Deprecated software and alternatives
| Deprecated | Alternate | Used For | Alternative |
| screen | tmux | Virtual terminals. You should use this! | tmux has similar keys to screen but uses ctrl-b instead of ctrl-a to access them. eg. create a new window: “ctrl-b n” |
| cd-hit | mmseqs | Clustering sequences | cd-hit is still an option if you want, but mmseqs2 appears to be much better |
Download a genome and remove the ribosomal RNA operon
For our search SRA engine, we want to remove the ribosomal RNA operon (not just the 16S gene, the whole opeon) before we run the search, otherwise all our hits are to the rRNA genes!
Here’s who you can use PATRIC to download a genome and remove the 16S region. For the example, we’re going to use a Faecalibacterium prausnitzii genome, because, well why not!
First, we download the genome and convert the GTO to fasta
p3-gto 657322.3
rast-export-genome -i 657322.3.gto contig_fasta > 657322.3.fnaNext, we use a couple of helper scripts from the EdwardsLab Git Repo. We start by converting the gto to a tab separated file with features and their locations
python3.7 ~/EdwardsLab/patric/parse_gto.py -f 657322.3.gto -p > 657322.3.tabThen we can grep through that file for the ribosomal genes:
grep rna 657322.3.tab | grep SubunitWe only find two of the genes:
fig|657322.3.rna.5 Large Subunit Ribosomal RNA; lsuRNA; LSU rRNA FP929046 586941 - 589785 (-)
fig|657322.3.rna.6 Small Subunit Ribosomal RNA; ssuRNA; SSU rRNA FP929046 590567 - 591540 (-)Now we can trim out the sequences and keep only the non-rRNA regions. Note that here I trim a little extra off the sequences, but you may not wish to do that
python3.7 ~/EdwardsLab/manipulate_genomes/trim_fasta.py -f 657322.3.fna -e 576941 -c FP929046 > FP929046.fna
python3.7 ~/EdwardsLab/manipulate_genomes/trim_fasta.py -f 657322.3.fna -b 601540 -c FP929046 >> FP929046.fnaWe run this twice, which is suboptimal, but this is definitely not the most computationally challenging thing we will do with those sequences!
Connecting to an anvi’o server on tatabox
We use anvi’o for all sorts of ‘omics analysis, but it is a pain to run on your laptop as you can’t watch netflix and youtube, check facebook, and post to twitter at the same time (well, you can, but why would you?).
Instead, we have the latest version of anvi’o installed on tatabox, one of the machines in our HPC environment. After you have run all the anvi-commands, very often you want to launch anvi-interactive, but tatabox is safely behind a firewall.
We can make a two step connection to tatabox using port tunneling. Depending on how you do this, you will need three terminals open.
First, start anvi-interactive on tatabox, and keep that window open (or use screen or tmux which are much better alternatives).
Next, open a terminal on your computer, and use this command. Change XXXX to a port near the on that you ssh to on edwards-data, change YYYY to the port that you normally use, and change USERNAME to your USERNAME.
ssh -L 5555:localhost:XXXX -N -p YYYYUSERNAME@edwards-data.sdsu.edu
Next, open another terminal (or if you are using screen or tmux, open a new terminal emulator), and login to edwards-data.sdsu.edu using your normal account (the USERNAME from above).
On edwards-data, run this command:
ssh -L XXXX:localhost:8080 -N USERNAME@tataboxFinally, on your laptop, you should open a new browser window and paste this URL:
http://localhost:5555/You should see the anvi-interactive interface appear, and you can get to work.
SRA Metadata with JSON
We have been working on updating access to the SRA metadata. In previous posts, we used the SQLite database provided by the Meltzer lab, but for a variety of reasons, we are now using the XML provided by the NCBI.
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Press about Global Phylogeography of crAssphage
Our paper on the global phylogeography of crAssphage is published in Nature Microbiology. You can read the paper at the Nature Microbiology website or on ReadCube. The paper garnered international press attention, and here we have summarized the press coverage.
Please let Rob know if you are aware of any other reports that are not included here.
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NSF Collaborators and Other Affiliations Information
For several years NSF ran a trial where they would ask for a conflicts form in excel-type format. Recently, that has been codified into the Collaborators and Other Affiliations Information form. You can find more information about that form at the NSF Website and the NSF GPG.
We developed a simple script to help complete this form for you. It does not do all the work, but it gets you a long way there, and you can do the rest a lot easier. After all, you have a lot of other things to worry about when you are writing that grant. Read more to see how to use it.
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