Our recent experience has shown us that there can be considerable variation in quality and cross-purposes in project design when researchers first start conducting next generation sequencing (NGS) projects. It is a complex world of terminology and constantly changing technology.
We hope that with the transfer of some background knowledge and a few tips, researchers might more accurately communicate their needs to their sequencing facility. The purpose of this advisory document is to highlight options for mouse researchers in terms of exome capture and sequencing capabilities, and then discusses data output standards and expectations to help guide analysis.
Some of the questions we considered are:
- What technologies are used for exome capture and sequencing?
- Should exomes be indexed pre- or post-capture?
- How many exomes can be indexed per lane?
- Is it preferable to request Gb per exome over raw coverage?
- How to interpret quality assurance measures?
- What methods are used for sequence alignment and single nucleotide variation (SNV) detection?
- To what extent are SNV lists filtered or quality checked?
- What information might be usefully contained in a sequencing report?
- How can SNV's be validated?