Next Generation Sequencing (NGS)/Genotyping by sequencing

Genotyping By Sequencing (GBS) is a method of cataloguing genetic variation between individuals on a common reference genome by using Next Generation Sequencing.

Overview

Genetic variation between individuals is a key component of explaining phenotype, traits or characteristics. For this reason, cataloguing variations between individuals is an important step towards understanding, predicting, and ultimately engineering biology.

There are many methods of cataloguing variation between individuals. Genotyping By Sequencing (GBS) being a common method.

GBS relies on generating sequencing data for an individual which is aligned to the reference genome. Variations are then called on the reference using a variety of methods.

Useful background

There are two key 'biological' concepts related to GBS, sequencing, alignment and genotyping.

From an informatics perspective, imputation.

Objective

For a given individual, GBS produces a catalogue of variation for that individual. It is a relatively cheap and 'un-biased' method of discovering variations, not being limited to variations within genes, or transcripts, and without any prior expectation of polymorphic sites.

Biological questions

Variation from GBS can be used to explain quantitative differences between individuals, better understand genetic relationships between groups of individuals, allow high density genetic maps to be constructed. GWAS.

Inputs and outputs

Inputs

• A reference genome
• Whole Genome Sequencing over one or more individuals.

Outputs

• For each individual, a list of variant loci.
• Variations may include SNPs, InDels, or Structural Variations, depending on the sequencing technology used.

Experimental design

There are two broad approaches to GBS, 1) relatively high depth of coverage over N individuals, or 2) relatively low level coverage of N groups of individuals (with relativly high coverage per group).

If only short read sequencing technology is used, then only SNPs and small InDels will be discovered. If longer reads and paired-end sequencing technology is used, larger InDels and structural variation may be discovered.

Typical steps in the method

Sample collection

Summary

Brief bit of information.

Sequencing

Summary

Brief bit of information.

QC and filtering

Summary

Brief bit of information.

Alignment

Summary

Brief bit of information.

Genotype calling

Summary

Brief bit of information.

Data submission

Summary

Brief bit of information.

Next steps

Depending on the type of experiment, GWAS, GSEA, Genetic Mapping.

Workflows

Example galaxy workflow

Link to an example galaxy workflow for for the method (including example datasets) on a given galaxy instance or to the XML document describing the workflow.

Discussion

POV discussion about the method.

Links to related discussion on BioStar: Genotyping By Sequencing on Biostars