Per base N content

Some sequencer substitute a N when it's unable to make a base call with sufficient confidence. This graph shows the percentage of N base calls at each position.

A low proportion of N at the end of the sequence is not a cause for concern. There is often more N on read 2 than on read 1, this is also normal. If the proportion becomes too high, this indicates degradation of the sequenced material for example.

Novaseq sequencer do not use N anymore. On the other hand, MiSeq and HiSeq from Illumina can call Ns.