Skip to main page content
Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2013 Nov;76(4):718-27.
doi: 10.1111/tpj.12319. Epub 2013 Oct 10.

Anchoring and Ordering NGS Contig Assemblies by Population Sequencing (POPSEQ)

Free PMC article

Anchoring and Ordering NGS Contig Assemblies by Population Sequencing (POPSEQ)

Martin Mascher et al. Plant J. .
Free PMC article


Next-generation whole-genome shotgun assemblies of complex genomes are highly useful, but fail to link nearby sequence contigs with each other or provide a linear order of contigs along individual chromosomes. Here, we introduce a strategy based on sequencing progeny of a segregating population that allows de novo production of a genetically anchored linear assembly of the gene space of an organism. We demonstrate the power of the approach by reconstructing the chromosomal organization of the gene space of barley, a large, complex and highly repetitive 5.1 Gb genome. We evaluate the robustness of the new assembly by comparison to a recently released physical and genetic framework of the barley genome, and to various genetically ordered sequence-based genotypic datasets. The method is independent of the need for any prior sequence resources, and will enable rapid and cost-efficient establishment of powerful genomic information for many species.

Keywords: Hordeum vulgare; barley; genetic mapping; genome assembly; next-generation sequencing; population sequencing; technical advance.


Figure 1
Figure 1
Schematic representation of POPSEQ.(a) A segregating population (80–100 individuals) is constructed from a bi-parental cross.(b) A whole-genome shotgun is generated for one parent, and used to construct a gene space assembly (alternatively, the POPSEQ data itself may be used for this purpose). On this assembly, gene models (green arrows) are defined using RNA–seq. In parallel, POPSEQ, and, if necessary, genotyping-by-sequencing (GBS), is performed on the population, and a medium-density framework genetic map is calculated (thousands to tens of thousands of loci).(c) SNPs detected and typed by POPSEQ along with associated WGS contigs are integrated into the framework map through nearest-neighbor search.(d) The result of POPSEQ is a sequence assembly in linear order that contains comprehensive information on the gene space. It may be enhanced by performing POPSEQ on additional populations.
Figure 2
Figure 2
POPSEQ validation. WGS contigs anchored to three genetic maps. These plots show the colinearity of contigs anchored to the Morex × Barke iSelect framework map and (a) the physical and genetic framework of barley (International Barley Genome Sequencing Consortium, 2012), (b) a Morex × Barke genetic map constructed by genotyping-by-sequencing (GBS), (c) a GBS map (Poland et al., 2012) constructed in the OWB. WGS contigs are shown as dots, and are mostly within 5 cM of the diagonal: 90.8% in (a), 99.2% in (b) 93.2% in (c).

Similar articles

  • Combinatorial pooling enables selective sequencing of the barley gene space.
    Lonardi S, Duma D, Alpert M, Cordero F, Beccuti M, Bhat PR, Wu Y, Ciardo G, Alsaihati B, Ma Y, Wanamaker S, Resnik J, Bozdag S, Luo MC, Close TJ. Lonardi S, et al. PLoS Comput Biol. 2013 Apr;9(4):e1003010. doi: 10.1371/journal.pcbi.1003010. Epub 2013 Apr 4. PLoS Comput Biol. 2013. PMID: 23592960 Free PMC article.
  • A sequence-ready physical map of barley anchored genetically by two million single-nucleotide polymorphisms.
    Ariyadasa R, Mascher M, Nussbaumer T, Schulte D, Frenkel Z, Poursarebani N, Zhou R, Steuernagel B, Gundlach H, Taudien S, Felder M, Platzer M, Himmelbach A, Schmutzer T, Hedley PE, Muehlbauer GJ, Scholz U, Korol A, Mayer KF, Waugh R, Langridge P, Graner A, Stein N. Ariyadasa R, et al. Plant Physiol. 2014 Jan;164(1):412-23. doi: 10.1104/pp.113.228213. Epub 2013 Nov 15. Plant Physiol. 2014. PMID: 24243933 Free PMC article.
  • Barley whole exome capture: a tool for genomic research in the genus Hordeum and beyond.
    Mascher M, Richmond TA, Gerhardt DJ, Himmelbach A, Clissold L, Sampath D, Ayling S, Steuernagel B, Pfeifer M, D'Ascenzo M, Akhunov ED, Hedley PE, Gonzales AM, Morrell PL, Kilian B, Blattner FR, Scholz U, Mayer KF, Flavell AJ, Muehlbauer GJ, Waugh R, Jeddeloh JA, Stein N. Mascher M, et al. Plant J. 2013 Nov;76(3):494-505. doi: 10.1111/tpj.12294. Epub 2013 Aug 24. Plant J. 2013. PMID: 23889683 Free PMC article.
  • Genetic anchoring of whole-genome shotgun assemblies.
    Mascher M, Stein N. Mascher M, et al. Front Genet. 2014 Jul 7;5:208. doi: 10.3389/fgene.2014.00208. eCollection 2014. Front Genet. 2014. PMID: 25071835 Free PMC article. Review.
  • Prospects of pan-genomics in barley.
    Monat C, Schreiber M, Stein N, Mascher M. Monat C, et al. Theor Appl Genet. 2019 Mar;132(3):785-796. doi: 10.1007/s00122-018-3234-z. Epub 2018 Nov 16. Theor Appl Genet. 2019. PMID: 30446793 Review.
See all similar articles

Cited by 111 articles

See all "Cited by" articles


    1. Andolfatto P, Davison D, Erezyilmaz D, Hu TT, Mast J, Sunayama-Morita T, Stern DL. Multiplexed shotgun genotyping for rapid and efficient genetic mapping. Genome Res. 2011;21:610–617. - PMC - PubMed
    1. Brenchley R, Spannagl M, Pfeifer M, et al. Analysis of the bread wheat genome using whole-genome shotgun sequencing. Nature. 2012;491:705–710. - PMC - PubMed
    1. Chandler VL, Brendel V. The Maize Genome Sequencing Project. Plant Physiol. 2002;130:1594–1597. - PMC - PubMed
    1. Chen M, Presting G, Barbazuk WB, et al. An integrated physical and genetic map of the rice genome. Plant Cell. 2002;14:537–545. - PMC - PubMed
    1. Comadran J, Kilian B, Russell J, et al. Natural variation in a homolog of Antirrhinum CENTRORADIALIS contributed to spring growth habit and environmental adaptation in cultivated barley. Nat. Genet. 2012;44:1388–1392. - PubMed

Publication types

LinkOut - more resources