Question: plasmid recovery from hybrid assembly
0
gravatar for piotr.majewski
8 months ago by
piotr.majewski0 wrote:

Dear All,

After hybrid Unicycler assembly, I've received 3 contigs

1 (4.35 Mb) - Alcaligenes faecalis genome, circular (via Geneious) 2 (43.5 kb) - plasmid?, circular (via Geneious) 3 (11 kb) - ?

how can I be sure that contigs 2&3 stands for plasmids? or maybe single plasmid divided into 2 contigs? I've tried to use PlasmidFinder, however its limited for Enterobacteriaceae.

ADD COMMENTlink modified 8 months ago by predeus1.3k • written 8 months ago by piotr.majewski0
2
gravatar for predeus
8 months ago by
predeus1.3k
Russia
predeus1.3k wrote:

You are right to be concerned - at least once in my practice Unicycler have generated a false circular contig, which was not a real plasmid at all.

In order to be SURE, you need to run a gel with non-fragmented DNA and with long ladder, and see bands at 44kb and 11kb.

If you see typical plasmid genes (replication machinery etc), that's another good sign.

You can also simply blast the assembled sequences and see what do they match - if they match fair chunks of known plasmids, you can be reasonably confident.

For a somewhat more loose search, try PLSDB: https://ccb-microbe.cs.uni-saarland.de/plsdb/ - they use distance metrics and can find things that are more remotely related, when compared to blast.

ADD COMMENTlink written 8 months ago by predeus1.3k
0
gravatar for kapsakcj
8 months ago by
kapsakcj30
USA/Atlanta
kapsakcj30 wrote:

You might be able to get a hint if you take the sequence of the 2nd and 3rd contigs and blast them. Perhaps they will align to other plasmid sequences.

Could also annotate the contigs with Prokka to see what genes may be present - do they seem to be plasmid related?

ADD COMMENTlink written 8 months ago by kapsakcj30
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