mummer dot plot

Question

[David]

Hello, I have aligned two complete bacterial genomes using mummer (1 contig per genome)

I´m not sure i understand well the output.

Here is the mummer plot:

I think the plot suggest that there are two main reverse regions. Is that correct.

If so is this what is happening at the genome level. Trying to understand if this is correct (A)?

Or this (B) ?

I would say that the B should be ok.

[alex.zaccaron]

If these are complete bacterial genomes, they are likely circular. I believe you are seeing this dot plot because one the genomes has the opposite orientation (reverse complement) and is rotated compared to the reference. I don't think there is any rearrangement.

Answer 1

[David]

Hi Alex, Definetly you are right (thanks for the figure)!!!!! Let me add more info to the discussion. The two genomes have been assembled from the same input pacbio dataset (RSII). The only difference is that one was assembled 3 years ago and the other yesterday with Flye.

WOuld there be any explanation why the genome have been rotated. The first one was generated by another person but i don´t have the info. I was thinking they rotated the genome according to the ori but i cannot understand it is reverse complemented compared to my assembly. Do you see any particular reason ???

[alex.zaccaron]

Hi David. I see no reason that you would obtain a new assembly with the same orientation and starting point as the previous one. It depends on the approach followed by the assembler. Note that, technically speaking, a circular genome has no starting point. Although it is not required, for better comparisons, you can manually rotate your new assembly to make its starting point match the previous assembly. You can use show-coords from MUMmer to check how many bases you should rotate the assembly (should be equal to the size of your small aligned fragment).

I would also suggest to use dnadiffwhich is also part of MUMmer package. It reports SNPs and short INDELs, which can help identify points of variability between your assemblies.

EDIT: If you know the exact number of bases to rotate the circular genome, you can use seqkit restart as described here to rotate the fasta sequence.

[rthapa]

Hi Alex, Do we need to change the orientation and starting point of the genome assembly of bacteria before using mummer to check SNPs and INDELs?

[alex.zaccaron]

Hi rthapa. No, it shouldn't matter for mummer to call SNPs and INDELs if the two genomes have different start points.

Answer 2

[David]

Thanks Alex, these are complete genomes. A quick blast shows that the longer fragment matches plus/minus to the reference. And the small fragment also blast in plus/minus compared to the reference. However i would expect a single fragment and not two. This suggests that the small fragment is also reversed and at the wrong place compared to the reference? Not sure ?

[alex.zaccaron]

From my understanding, you are getting two fragments because one of your genomes is rotated compared to your reference. A similar pattern is obtained if you align a sequence to its rotated reverse complement (see panel C of this figure).