How to order contigs generated from PlasmidSPAdes

Question

[mandigene94]

Hello everyone,

I'm new here and I'm hoping you may have some insights into my problem (I'm sorry in advance if this is a silly question, but I'm new to bioinformatics in general).

My overall goal is to align my whole genome sequence data using plasmidSPAdes to generate plasmid contigs (I'm only interested in the plasmid sequences for later analysis) and then put those together and complete gene predictions on the plasmids.

At this point, I have used plasmidSPAdes and have a .fasta file with my contigs, as well as a .fastg file that I've opened with Bandage, which seems to give me a very rudimentary map. Maybe I am overthinking it (maybe I already have all of the data/results and just need to open it) but I'm wondering if you may have suggestions as to how to put these contigs together (or at least try to) from the .fasta files so I can see the whole map for each plasmid? I've seen some programs (like Recycler, for example) that seem to do what I want, but I don't use Python and I'm not sure if it's possible to do with another program using what I already am (I'm using a Unix Shell -MobaXterm, but as far as I know, it's just using a normal command line, not Python, like Recycler requires - again, I apologize if I'm not explaining very well). I'm willing to learn Python if all else fails, but I'm only interested in using, say, Recycler for one stage overall and am hoping there's a way to do what I need without having to figure out a new program in a new system. Any input you might have would be greatly appreciated!

Cheers!

[Sej Modha]

Have you tried Mauve contig ordering tool? I am not sure if it takes a fastg file as an input though.

[mandigene94]

I'm trying it right now, and it seems to be working - thank you very much for your help!

Answer 1

[Roye Rozov]

Hi mandigene94,

I am the developer of Recycler, and I think Recycler might help you, so hopefully I can help you understand how to run it. The good news is you don't need to learn python at all to be able to use Recycler - you just need to follow the commands as they appear on the GitHub page. Those commands assume you have some necessary packages and tools Recycler depends on, such as python itself, samtools, and BWA, but most of these you will likely need anyway for common bioinformatics tasks. To see if you have python installed on your system, type the command python. If it is installed, a command prompt will appear; if not, an error should be reported that the command isn't recognized.

The easiest way to install an up to date python (2.7 - not 3.x!) distribution and needed packages such as pysam is through a distribution called Anaconda. Once Anaconda is installed, you just run conda install <package-name> (without the brackets) to install packages you want that are available through Anaconda. Most popular ones are.

Once you've sorted out the dependencies listed on Recycler's GitHub page, you can just run the commands as they appear there. It sounds like you've already run plasmidSPAdes on your data, which means you can easily run plain spades to get the assembly graph Recycler expects. Note we haven't tested running Recycler on the graph output by plasmidSPAdes, but that is definitely an interesting option as well.

Let me know if you have further questions.

Cheers, Roye