Solexa file

Illumina Inc. High-throughput genotyping by whole-genome resequencing. Zerbino D , Birney E. Velvet: algorithms for de novo short read assembly using de Bruijn graphs. Ultrafast and memory-efficient alignment of short DNA sequences to the human genome. Genome Biol. Oxford University Press is a department of the University of Oxford. It furthers the University's objective of excellence in research, scholarship, and education by publishing worldwide.

Sign In or Create an Account. Sign In. Advanced Search. Search Menu. Article Navigation. Close mobile search navigation Article Navigation. Volume Cock , Peter J. Oxford Academic. Christopher J. Naohisa Goto. Michael L. Peter M. Cite Cite Peter J. Select Format Select format. Permissions Icon Permissions. ABSTRACT FASTQ has emerged as a common file format for sharing sequencing read data combining both the sequence and an associated per base quality score, despite lacking any formal definition to date, and existing in at least three incompatible variants.

This introduced the PHRED quality score of a base call, defined in terms of the estimated probability of error:. Table 1. Open in new tab. Although the FASTQ format only records a single quality score per letter, Solexa also produced other files with quality scores for all four bases, and in order to represent low-quality information more fully an alternative logarithmic mapping was used Solexa quality scores are defined as:.

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Please check for further notifications by email. Furthermore, as part of this switch for GAPipeline 1. While the Illumina 1. Even reading the literature can be confusing, for example Huang et al. Although some of these tools can convert from the Solexa and in some cases also the Illumina 1. Therefore, most users will do this conversion very early in their workflows perhaps using OBF software.

We hope that this trend will lead to Illumina themselves switching to the original FASTQ convention at a later date, which would eventually relegate this confusion of incompatible variants to a historical concern. In addition to simple conversion between FASTQ variants, other common steps in a sequencing pipeline include quality and adaptor trimming, and contaminant or quality-based filtering.

C , Funding for open access charge: P. National Center for Biotechnology Information , U. Journal List Nucleic Acids Res v. Nucleic Acids Res.

Published online Dec Peter J. Fields , 2 Naohisa Goto , 3 Michael L. Heuer , 4 and Peter M. Rice 5. Christopher J. Michael L. Peter M.

Author information Article notes Copyright and License information Disclaimer. Published by Oxford University Press. This article has been cited by other articles in PMC. Abstract FASTQ has emerged as a common file format for sharing sequencing read data combining both the sequence and an associated per base quality score, despite lacking any formal definition to date, and existing in at least three incompatible variants.

Table 1. Open in a separate window. Figure 1. Conflict of interest statement. None declared. Supplementary Material [Supplementary Data] Click here to view. Improved tools for biological sequence comparison. Natl Acad. Bennett S. Solexa Ltd. Genome sequencing in microfabricated high-density picolitre reactors.

In: Janitz M, editor. Wiley; Cock P. A gaussian is fitted to the resulting histogram. This image is then thresholded. The factor currently used is 4. After thresholding object detection takes place. The algorithm is reasonably straightforward. Every pixel in the image is itterated over when a pixel above the threshold in encountered a boundary is created. The boundary is created by walking all adjacent above threshold pixels clockwise. The area covered by this newly identified cluster is also calculated, and the pixels in this position on the original image unset to avoid detecting this cluster again.

During this process the maximum pixel intensity in this cluster is also found. It is the maximum pixel intensity that is passed to the base caller rather than a function of all the pixels in this cluster. Split objects no longer have boundaries, and so only the maximum pixel is shown. Once final objects have been identified local background can be compensated for.

All pixels that are not part of a cluster are extracted and sorted. The procedure is iterated and each time 3-sigma outliers are removed to avoid large contaminations by residual low-level signal. This iteration terminates if less than 2 pixels are removed or less than 20 pixels remain. Bustard is the Illumina basecaller, the base call is simply the maximum intensity after Crosstalk and Phasing corrections have been applied. Quality score calculation is more involved NOTE: add more. Phasing is a term used to describe the signal contribution of bases from the previous and next cycles.

Phasing and prephasing are averaged over the run and used to derive a correction matrix. NOTE: add detail and check. Chastity is a similar metric, and is defined as the ratio of the largest intensity to the sum of the largest and second largest intensities. As pointed out by Irina, the logic of this definition is somewhat faulty. If the second highest intensity is negative as it can be due to background compensation it will contribute to the sum incorrectly.

To further complicate the situation Chastity is also sometimes used to mean a limit for purity for example in PhasingEstimate. Chastity of 0.