An ultra-high density genetic linkage map of perennial ryegrass (Lolium perenne) using genotyping by sequencing (GBS) based on a reference shotgun genome assembly.
High density genetic linkage maps that are extensively anchored to assembled genome sequences of the organism in question are extremely useful in gene discovery. To facilitate this process in perennial ryegrass (Lolium perenne L.), a high density single nucleotide polymorphism (SNP)- and presence/absence variant (PAV)-based genetic linkage map has been developed in an F2 mapping population that has been used as a reference population in numerous studies. To provide a reference sequence to which to align genotyping by sequencing (GBS) reads, a shotgun assembly of one of the grandparents of the population, a tenth-generation inbred line, was created using Illumina-based sequencing.



The assembly was based on paired-end Illumina reads, scaffolded by mate pair and long jumping distance reads in the range of 3-40 kb, with >200-fold initial genome coverage. A total of 169 individuals from an F2 mapping population were used to construct PstI-based GBS libraries tagged with unique 4-9 nucleotide barcodes, resulting in 284 million reads, with approx. 1·6 million reads per individual. A bioinformatics pipeline was employed to identify both SNPs and PAVs. A core genetic map was generated using high confidence SNPs, to which lower confidence SNPs and PAVs were subsequently fitted in a straightforward binning approach.



The assembly comprises 424 750 scaffolds, covering 1·11 Gbp of the 2·5 Gbp perennial ryegrass genome, with a scaffold N50 of 25 212 bp and a contig N50 of 3790 bp. It is available for download, and access to a genome browser has been provided. Comparison of the assembly with available transcript and gene model data sets for perennial ryegrass indicates that approx. 570 Mbp of the gene-rich portion of the genome has been captured. An ultra-high density genetic linkage map with 3092 SNPs and 7260 PAVs was developed, anchoring just over 200 Mb of the reference assembly.



The combined genetic map and assembly, combined with another recently released genome assembly, represent a significant resource for the perennial ryegrass genetics community.
Teagasc, Crops, Environment and Land Use Programme, Oak Park Research Centre, Carlow, Ireland University College Dublin, School of Agriculture and Food Science, Dublin, Ireland.
Janaki
J
Velmurugan
Teagasc, Crops, Environment and Land Use Programme, Oak Park Research Centre, Carlow, Ireland Information and Computational Sciences Group, James Hutton Institute, Errol Road, Invergowrie, Dundee, UK Division of Plant Sciences, University of Dundee at the James Hutton Institute, Errol Road, Invergowrie, Dundee, UK.
Ewan
E
Mollison
Teagasc, Crops, Environment and Land Use Programme, Oak Park Research Centre, Carlow, Ireland.
Susanne
S
Barth
Information and Computational Sciences Group, James Hutton Institute, Errol Road, Invergowrie, Dundee, UK.
David
D
Marshall
Information and Computational Sciences Group, James Hutton Institute, Errol Road, Invergowrie, Dundee, UK.
Linda
L
Milne
Teagasc, Animal and Grassland Research and Innovation Centre, Grange, Ireland.
Christopher J
CJ
Creevey
University College Dublin, School of Agriculture and Food Science, Dublin, Ireland.
Bridget
B
Lynch
Teagasc, Crops, Environment and Land Use Programme, Oak Park Research Centre, Carlow, Ireland.
Helena
H
Meally
Teagasc, Animal and Grassland Research and Innovation Centre, Grange, Ireland.
Matthew
M
McCabe
Teagasc, Crops, Environment and Land Use Programme, Oak Park Research Centre, Carlow, Ireland dan.milbourne@teagasc.ie.
Dan
D
Milbourne
D002874:Chromosome Mapping; D008040:Genetic Linkage; D018745:Genome, Plant; D060005:Genotyping Techniques / Q000379:methods; D059014:High-Throughput Nucleotide Sequencing; D006720:Homozygote; D008129:Lolium / Q000235:genetics*; D020641:Polymorphism, Single Nucleotide
GBS; Lolium perenne; genome assembly; genotyping by sequencing; linkage mapping; perennial ryegrass; presence/absence variation; single nucleotide polymorphism