FASTQ
Definition
The standard sequencing output file format containing four-line records per read: identifier, nucleotide sequence, separator, and Phred quality score string. All MPS pipelines begin with FASTQ input.
- Field
- Next-generation sequencing (NGS/MPS)
- Structure
- Four lines per read: ID, sequence, separator, Phred quality
- Role
- Standard raw output format from sequencing instruments
- Pipeline position
- First input stage before alignment and allele calling
Common questions
What does the Phred quality line in a FASTQ record actually represent?+
It is a per-base confidence score encoded as ASCII characters, indicating the sequencer's estimated probability that each called nucleotide is correct; downstream pipelines use these scores to filter or trim low-confidence bases before further analysis.
Why does FASTQ matter for the reliability of a forensic DNA sequencing result?+
Every quality-filtering, alignment, and allele-calling decision downstream starts from the FASTQ file, so if the raw read quality is poor or the file is mishandled at this stage, errors propagate through the entire analysis regardless of how good the later software steps are.
Related terms
- BAM File
- Binary Alignment/Map format, the compressed binary version of a SAM file. Stores aligned reads with their mapping positions, quality scores, and alignment...
- Isoallele
- Two STR alleles that share the same repeat count (and therefore appear as the same allele on CE) but differ by a...
- Phred Quality Score (Q)
- A logarithmic probability score for base-call accuracy. Q30 = 99.9% accuracy (1 in 1000 error probability). Q20 = 99% accuracy. Most forensic...
- Stutter (MPS)
- In MPS, stutter includes both length-based stutter (as in CE, an n-1 repeat artefact from PCR slippage) and sequence-based stutter (single-nucleotide errors...
- VCF
- Variant Call Format, the standard output from variant callers (GATK, FreeBayes). Records each variant position with reference allele, alternative allele(s), genotype likelihoods,...