Hi everyone,
We have sequenced genome of data of a comparatively less studied species using Pacbio long read sequencer. Now after initial assembly we compared the contigs with previous version of the genome. From comparative study we came to know two newly assembled contigs came from a same chromosome, so we believe there is gap in pacbio sequencing data we have generated. Now, we also had a Illumina paired-end data (150*2) for the species. We tried to fill the gap using Pilon. but failed. So I am looking for any suggestions on how to fill the gap : (a) using bioinformatics (b) using another sequencing experiment. Or we are missing any thing prior to that.
Thanks in advance...
Hi Sourav Nayak , I think this problem is quite complex since you don't really know what might be 'causing' the gap. For example there could be a structural variant between the individuals sequenced or that different sequencing and assembly approaches have resolved a complex region differently. However, if you are certain you want to span this gap I think the best way of doing that would be to add pacbio data, ideally with the longest reads possible (although this is only practical if your organism's genome is small and you have money to do this) and hope some of these will allow your assembler to span the gap. Secondly if you really want a contiguous sequence, at least to see what might be going on in that region, the tool quickmerge: https://github.com/mahulchak/quickmerge might be of use to you (if you do not mind some kind of hybrid assembly between the existing one you have and your new assembly). Good luck! Rishi
Although PacBio is good for long reads, it has some glitches in the functional regions/genes and this can cause errors. Which in the end results in gaps in the assembly.
But initially, before answering, I would ask about the gaps? How long are the gaps and what are the percentage (missing coverage) of gaps in your genome? What is your organism? How large do you think your genome is? Do you have any good reference? Are you doing de novo assembly?
Gap filling is one of the final phases of genome assembly, especially in large genome. To do this tricky job smoothly, you may try gapFinisher- a reliable gap filling pipeline for SSPACE-LongRead scaffolder output like you do have.
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