Alignment of short sequences allowing gaps and wildcards

Question

[filippo.martignano]

Hi everybody!

I have sequenced some DNA fragments (with nanopore) that had been previously barcoded with a barcode like this:

AATACGACTCACTATAGNNNNNNNNNNTATCCTCANNNNNNNNNNCTATAGTGTCACCTAAA

so basically it is made by 3 different barcodes separated by 10 random nucleotides:

Barcode1 - 10Ns - Barcode2 - 10Ns - Barcode3

I'm looking for an alignment tool that takes in account the presence of the Ns (wildcards) in order to detect the reads that incorporate this barcode. Also, since I used nanopore for sequencing, the reads produced are full of fake indels, so the aligner should also allow the presence of little gaps (1-3 bp), or it should be possible to adjust the gap penalty.

Thank you very much in advance!

Answer 1

[shelkmike]

You may want to look at the "Extraction of UMI reference sequences" paragraph in the paper "Enabling high-accuracy long-read amplicon sequences using unique molecular identifiers with Nanopore or PacBio sequencing" (https://www.biorxiv.org/content/10.1101/645903v3.full ). Authors had a similar task and dealt with it by using cutadapt.