I am working on 3 human genes which are very homologous but also rich in structural variation and SNPs, and therefore have obtained nanopore data from sequencing their PCR products (whereby the primers bind uniquely to each gene).
I have demultiplexed the reads to each of the four donors and then filtered the reads so I only keep the reads that both start and end exactly where the primers should have bound. Now I am trying to perform variance calling for each gene per donor but am confused about which tools to use and in which order...
I'm aware of a tool called nanopolish finds variance but do I have to use another tool to get the consensus sequence of all the reads?
I have tried to use Canu to de novo assemble the reads as well to see how it differs from the reference genome (as due to the homology I'm not sure how far I can trust it) but I cant get it to work. Perhaps this is because all of my reads are stacked on top of each other without gaps?
Any advice on which direction to take this next is much appreciated!