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GFF and GTF I/O

Learning Objective

In this tutorial, you will learn how to read and write GFF and GTF files.

Difficulty

Average

Duration

45 min

Prerequisites

tutorial-sequences, tutorial-input-output-overview, GFF Format Specification

This tutorial shows how to read and write GFF and GTF files using the GffFileIn and GffFileOut classes. It starts out with a quick reminder on the structure of GFF and GTF files and will then continue with how to read and write GFF and GTF files.

The GFF and GTF formats are used for annotating genomic intervals (an interval with begin/end position on a contig/chromosome). GFF exist in versions 2 and 3 and GTF is sometimes called "GFF 2.5". There are specifications for GFF 2, GFF 3, and GTF available elsewhere. GFF and GTF are TSV-based formats and in general have the same structure. The main difference is the underlying system/ontology for the annotation but also smaller differences in the format.

In this tutorial, we will focus on the format GFF 3 since it is the most current one with most complete tool support. The information of this tutorial can easily be translated to the other two formats.

The SeqAn module gff_io allows the reading and writing of the GFF and GTF formats.

Tip

Format Version Support in SeqAn

GffFileIn allows to read GFF files in version 2 and 3 and GTF files. For writing, GffFileOut supports only GFF 3 and GTF.

GFF Format

The following is an example of a GFF 3 file:

demos/tutorial/gff_and_gtf_io/example.gff

The meaning of the columns are as follows:

seq id (1)

Name of the reference sequence.

source (2)

Free text field describing the source of the annotation, such as a software (e.g. "Genescan") or a a database (e.g. "Genebank"), "." for none.

type (3)

The type of the annotation.

start (4)

The 1-based begin position of the annotation.

end (5)

The 1-based end position of the annotation.

score (6)

The score of the annotation, "." for none.

strand (7)

The strand of the annotation, "+" and "-" for forward and reverse strand, "." for features that are not stranded.

phase (8)

Shift of the feature regarding to the reading frame, one of "0", "1", "2", and "." for missing/dont-care.

attributes (9)

A list of key/value attributes. For GFF 3, this is a list of key=value pairs, separated by semicolons (e.g. ID=cds00003;Parent=mRNA00003). For GTF and GFF 2, this is a list of tuples, separated by semicolon. The first entry gives the key, the following entries are values. Strings are generally enclosed in quotes (e.g. Target "HBA_HUMAN" 11 55 ; E_value 0.0003)

Tip

1-based and 0-based positions.

There are two common ways of specifying intervals.

  1. Start counting positions at 1 and give intervals by the first and last position that are part of the interval (closed intervals). For example, the interval [1,000; 2,000] starts at character 1,000 and ends at character 2,000 and includes it. This way is natural to non-programmers and used when giving coordinates in GFF files or genome browsers such as UCSC Genome Browser and IGV.
  2. Start counting positions at 0 and give intervals by the first position that is part of the interval and giving the position behind the last position that is part of the interval. The interval from above would be [999; 2,000) in this case.

In text representations, such as GFF and GTF, 1-based closed intervals are used whereas in the internal binary data structures, SeqAn uses 0-based half-open intervals.

A First Working Example

The following example shows an example of a program that reads the file with the path example.gff and prints its contents back to the user on standard output.

demos/tutorial/gff_and_gtf_io/example1.cpp

The program first opens a GffFileIn for reading and a GffFileOut for writing. The GFF records are read into GffRecord objects which we will focus on below.

Assignment 1

Type

Reproduction

Objective

Create a file with the sample GFF content from above and adjust the path "example.gff" to the path to your GFF file (e.g. "/path/to/my_example.gff").

Solution

demos/tutorial/gff_and_gtf_io/solution1.cpp

Accessing the Records

The class GffRecord stores one record in a Gff file.

demos/tutorial/gff_and_gtf_io/base.cpp

The static members INVALID_POS, INVALID_REFID store sentinel values for marking positions and reference sequence ids as invalid. The static funtion INVALID_SCORE() returns the IEEE float "NaN" value.

Assignment 2

Counting Records

Type

Review

Objective

Change the result of Assignment 1 by counting the number of variants for each chromosome/contig instead of writing out the records.

Solution

demos/tutorial/gff_and_gtf_io/solution2.cpp

The output is

demos/tutorial/gff_and_gtf_io/solution2.cpp.stdout

Creating a New File

Assignment 3

Generating GFF From Scratch

Type

Application

Objective

Write a program that prints the following GFF file. Create GffRecord objects and write them to a GffFileOut using writeRecord().

demos/tutorial/gff_and_gtf_io/solution3.cpp.stdcout

Solution

demos/tutorial/gff_and_gtf_io/solution3.cpp

Next Steps

  • Continue with the tutorial.