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hexylena committed Dec 20, 2023
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In this section we will look at practical aspects of manipulation of next-generation sequencing data. We will start with the FASTQ format produced by most sequencing machines and will finish with the SAM/BAM format representing mapped reads. The cover image above shows a screen dump of a SAM dataset.

# Introduction to sequencing data

## FASTQ manipulation and quality control

[FASTQ](https://en.wikipedia.org/wiki/FASTQ_format) is not a very well defined format. In the beginning various manufacturers of sequencing instruments were free to interpret FASTQ as they saw fit, resulting in a multitude of FASTQ flavors. This variation stemmed primarily from different ways of encoding quality values as described [on the Wikipedia article for FASTQ](https://en.wikipedia.org/wiki/FASTQ_format) (below you will find an explanation of quality scores and their meaning). Today, the [FASTQ Sanger](https://www.ncbi.nlm.nih.gov/pubmed/20015970) version of the format is considered to be the standard form of FASTQ. Galaxy is using FASTQ Sanger as the only legitimate input for downstream processing tools and provides [a number of utilities for converting FASTQ files](https://www.ncbi.nlm.nih.gov/pubmed/20562416) into this form (see **FASTQ Quality Control** section of Galaxy tools).
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Thus in both cases (paired-end and mate-pair) a single physical piece of DNA (or RNA in the case of RNA-seq) is sequenced from two ends and so generates two reads. These can be represented as separate files (two FASTQ files with first and second reads) or a single file were reads for each end are interleaved. Here are examples:

#### Two single files
### Two single files

File 1

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> Note that read IDs are **identical** in two files and they are listed in **the same** order. In some cases read IDs in the first and second file may be appended with `/1` and `/2` tags, respectively.
{: .comment}

#### Interleaved file
### Interleaved file

```
@1/1
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