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Plant Biotechnol J. 2018 Apr;16(4):877-889. doi: 10.1111/pbi.12835. Epub 2017 Oct 13.

Genotyping-by-sequencing through transcriptomics: implementation in a range of crop species with varying reproductive habits and ploidy levels.

Author information

1
Agriculture Victoria, AgriBio, Centre for AgriBioscience, 5 Ring Road, Bundoora, Victoria 3083, Australia.
2
School of Applied Systems Biology, La Trobe University, Bundoora, Victoria 3086, Australia.

Abstract

The application of genomics in crops has the ability to significantly improve genetic gain for agriculture. Many marker-dense tools have been developed, but few have seen broad adoption in plant genomics due to issues of significant variations of genome size, levels of ploidy, single nucleotide polymorphism (SNP) frequency and reproductive habit. When combined with limited breeding activities, small research communities and scant sequence resources, the suitability of popular systems is often suboptimal and routinely fails to effectively balance cost-effectiveness and sample throughput. Genotyping-by-sequencing (GBS) encompasses a range of protocols including resequencing of the transcriptome. This study describes a skim GBS-transcriptomics (GBS-t) approach developed to be broadly applicable, cost-effective and high-throughput while still assaying a significant number of SNP loci. A range of crop species with differing levels of ploidy and degree of inbreeding/outbreeding were chosen, including perennial ryegrass, a diploid outbreeding forage grass; phalaris, a putative segmental allotetraploid outbreeding forage grass; lentil, a diploid inbreeding grain legume; and canola, an allotetraploid partially outbreeding oilseed. GBS-t was validated as a simple and largely automated, cost-effective method which generates sufficient SNPs (from 89 738 to 231 977) with acceptable levels of missing data and even genome coverage from c. 3 million sequence reads per sample. GBS-t is therefore a broadly applicable system suitable for many crops, offering advantages over other systems. The correct choice of subsequent sequence analysis software is important, and the bioinformatics process should be iterative and tailored to the specific challenges posed by ploidy variation and extent of heterozygosity.

KEYWORDS:

Brassica napus; Lens culinaris; Lolium perenne; Phalaris aquatic; RNA

PMID:
28913899
PMCID:
PMC5866951
DOI:
10.1111/pbi.12835
[Indexed for MEDLINE]
Free PMC Article

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