We are performing CRISPR in a gene from a Aspergillus niger strain and then sequencing the results via Sanger. When we blast the gene sequence against the resulting sequence we observe in the reverse read that it matches plus/minus (as it should), but also plus/plus. The dotplot looks like this:
It only happens in the reverse read of 2 out of the 30 mutants we sequenced. Anyone have any clue about what could be happening here? Looks like the sequence is slightly palindromic, but we don't know the reason. Maybe it has to do with some type of primer-dimer during PCR, or some inespecific primer binding site? It is strange that it only happens in a small subset of reverse reads. It already happened to us previously with the same strains and another primer set, but it also happens with the new primers. Thanks in advance.
