[niels_olson]

There are regions with sparse coverage due to higher concentrations of C-G bases. This can make alignment results less reliable. Plasmids may interfere with alignment or need to be isolated out before or during library prep. And you want to detect and discard any members of the population with any evidence of cancer, genetic disease, etc.

And you need good population coverage. What's a normal variant? Newer methods propose a graph alignment instead of just trying to build a single sequence reference genome.