FASTQ format

FASTQ format is a text-based format for storing both a biological sequence (usually nucleotide sequence) and its corresponding quality scores. Both the sequence letter and quality score are each encoded with a single ASCII character for brevity.

It was originally developed at the Wellcome Trust Sanger Institute to bundle a FASTA sequence and its quality data, but has recently become the de facto standard for storing the output of high-throughput sequencing instruments such as the Illumina Genome Analyzer.

A FASTQ file normally uses four lines per sequence.

A FASTQ file containing a single sequence might look like this:

The character '!' represents the lowest quality while '~' is the highest. Here are the quality value characters in left-to-right increasing order of quality (ASCII):

The original Sanger FASTQ files also allowed the sequence and quality strings to be wrapped (split over multiple lines), but this is generally discouraged as it can make parsing complicated due to the unfortunate choice of "@" and "+" as markers (these characters can also occur in the quality string).

Sequences from the Illumina software use a systematic identifier:

Versions of the Illumina pipeline since 1.4 appear to use #NNNNNN instead of #0 for the multiplex ID, where NNNNNN is the sequence of the multiplex tag.

With Casava 1.8 the format of the '@' line has changed:

Note that more recent versions of Illumina software output a sample number (as taken from the sample sheet) in place of an index sequence. For example, the following header might appear in the first sample of a batch:

FASTQ files from the NCBI/EBI Sequence Read Archive often include a description, e.g.

In this example there is an NCBI-assigned identifier, and the description holds the original identifier from Solexa/Illumina (as described above) plus the read length. Sequencing was performed in paired-end mode (~500bp insert size), see SRR001666. Notably in the above output the paired-end information was lost when the data was extracted from the NCBI SRA using fastq-dump with default settings.

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