Long-read sequence assembly: A technical evaluation in barley

Martin Mascher*, Thomas Wicker, Jerry Jenkins, Christopher Plott, Thomas Lux, Chu Shin Koh, Jennifer Ens, Heidrun Gundlach, Lori B. Boston, Zuzana Tulpová, Samuel Holden, Inmaculada Hernández-Pinzón, Uwe Scholz, Klaus F.X. Mayer, Manuel Spannagl, Curtis J. Pozniak, Andrew G. Sharpe, Hana Simková, Matthew J. Moscou, Jane GrimwoodJeremy Schmutz, Nils Stein

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review


Sequence assembly of large and repeat-rich plant genomes has been challenging, requiring substantial computational resources and often several complementary sequence assembly and genome mapping approaches. The recent development of fast and accurate long-read sequencing by circular consensus sequencing (CCS) on the PacBio platform may greatly increase the scope of plant pan-genome projects. Here, we compare current long-read sequencing platforms regarding their ability to rapidly generate contiguous sequence assemblies in pan-genome studies of barley (Hordeum vulgare). Most long-read assemblies are clearly superior to the current barley reference sequence based on short-reads. Assemblies derived from accurate long reads excel in most metrics, but the CCS approach was the most cost-effective strategy for assembling tens of barley genomes. A downsampling analysis indicated that 20-fold CCS coverage can yield very good sequence assemblies, while even five-fold CCS data may capture the complete sequence of most genes. We present an updated reference genome assembly for barley with near-complete representation of the repeat-rich intergenic space. Long-read assembly can underpin the construction of accurate and complete sequences of multiple genomes of a species to build pan-genome infrastructures in Triticeae crops and their wild relatives.

Original languageEnglish
Pages (from-to)1888-1906
Number of pages19
JournalPlant Cell
Issue number6
StatePublished - Jun 2021
Externally publishedYes


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