I have sequences of fungal isolates from the same species produced with HiSeq 2500 (2x150 bp with 350 bp insert). These isolates show different levels of pathogenicity so basicly I want to compare variations in their genome. So far I tried reference-guided genome assembly by mapping each isolate back to an available reference genome and calling a consensus sequence of such isolates with SAMtools. However, I got long stretches of n in some contigs. May I have your opinion how can I solve this problem? I have already tried changing a couple of aligners (BWA and Bowtie2) without success.
Also I was thinking if it would help to do de novo assembly using spades first and map contigs back to the reference genome? I appreciate all opinions and suggestions.
Many thanks in advance and have a great day!