This post is part 7 of a series on file formats, written for the 2017 UK-KBRIN Essentials of Next Generation Sequencing Workshop at the University of Kentucky.

qual format

A .qual file provides Phred-based quality scores for a set of sequences (often a corresponding FASTA file). It looks something like this:

>my_sequence1
4 39 8 4 50 1 100 5 0
>my_sequence2 
3 3 40 42 35

Similar to FASTA format, each sequence is defined with a header starting with the > character, and all subsequente characters after the line break describe the sequence. Unlike FASTA, the individual nucleotides are not included in this file, only the corresponding quality Phred-format scores. At this point, all we need to understand is what are Phred quality scores. Every nucleotide in a sequence will have a quality score associated with it. The score Q describes the probability P that the base is called in correctly:

Q = –10 log(P)

Let's look at some example Phred scores below:

Phred scores typically range from 2-40 with current sequencing technology. As you can see, setting a cutoff somewhere around 30 or higher results in fairly strict confidence in invidual base calls.