Question

When performing quality control on FASTQ data, what specific statistical pattern in the base-call distribution indicates that the sequencing library contains adapter contamination?

Accepted Answer

When performing quality control on FASTQ data, adapter contamination is identified by a specific rise in the frequency of non-genomic sequences at the 3&#x27; end of the reads. FASTQ data consists of sequences of nucleotides, which are the building blocks of DNA represented by the letters A, T, C, and G, accompanied by quality scores. During sequencing, adapters are short, synthetic DNA molecules attached to the ends of target DNA fragments to allow them to bind to the sequencing flow cell. If a DNA fragment is shorter than the number of cycles performed by the sequencer, the machine will read through the entire target fragment and begin sequencing the adapter sequence attached to the end. In the base-call distribution plot, which shows the frequency of each nucleotide at every position along the read, this appears as an abrupt, unnatural increase in the specific nucleotide content of the adapter sequence occurring simultaneously across a significant portion of the reads. For instance, if the adapter sequence begins with a string of guanines, you will see a sharp spike in the proportion of guanine base calls starting at the position where the target DNA fragment ends and the adapter begins. This pattern is distinct because it is not random; it follows the exact, known order of the synthetic adapter nucleotides rather than the expected biological variation of the DNA sample being studied.

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When performing quality control on FASTQ data, what specific statistical pattern in the base-call distribution indicates that the sequencing library contains adapter contamination?