TASSEL-GBS: A High Capacity Genotyping by Sequencing Analysis Pipeline

Jeffrey C Glaubitz,James Harriman,Robert J Elshire,Fei Lu,Edward S Buckler,Terry M Casstevens,Qi Sun,Nicholas A Tinker

doi:10.1371/journal.pone.0090346

Jeffrey C Glaubitz, James Harriman + Show 6 more

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https://doi.org/10.1371/journal.pone.0090346

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Abstract

Genotyping by sequencing (GBS) is a next generation sequencing based method that takes advantage of reduced representation to enable high throughput genotyping of large numbers of individuals at a large number of SNP markers. The relatively straightforward, robust, and cost-effective GBS protocol is currently being applied in numerous species by a large number of researchers. Herein we describe a bioinformatics pipeline, tassel-gbs, designed for the efficient processing of raw GBS sequence data into SNP genotypes. The tassel-gbs pipeline successfully fulfills the following key design criteria: (1) Ability to run on the modest computing resources that are typically available to small breeding or ecological research programs, including desktop or laptop machines with only 8–16 GB of RAM, (2) Scalability from small to extremely large studies, where hundreds of thousands or even millions of SNPs can be scored in up to 100,000 individuals (e.g., for large breeding programs or genetic surveys), and (3) Applicability in an accelerated breeding context, requiring rapid turnover from tissue collection to genotypes. Although a reference genome is required, the pipeline can also be run with an unfinished “pseudo-reference” consisting of numerous contigs. We describe the tassel-gbs pipeline in detail and benchmark it based upon a large scale, species wide analysis in maize (Zea mays), where the average error rate was reduced to 0.0042 through application of population genetic-based SNP filters. Overall, the GBS assay and the tassel-gbs pipeline provide robust tools for studying genomic diversity.

Highlights

The advent of generation sequencing has elicited a revolution in biology buoyed by an advancing tidal wave of raw sequence data[1,2,3,4]
Because of its relative simplicity and robustness, the genotyping by sequencing (GBS) method of Elshire et al [12] or close derivatives thereof have already been applied in numerous species by many researchers (e.g., [7,17,20,21,22,23,24,25,26,27,28,29,30,31])
A production-ready TOPM will be applicable to a breeding population as long as the genetic diversity present in the founders of the breeding population is well represented in the individuals that comprised the corresponding Discovery Build

Summary

Introduction

The advent of generation sequencing has elicited a revolution in biology buoyed by an advancing tidal wave of raw sequence data[1,2,3,4]. The TASSEL-GBS pipeline first collapses all of the reads into a master tag list containing all of the sequence tags present at or above a user-specified minimum count, tallied across all of the samples in the Discovery Build (Figure 1A).

Results

Conclusion