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Article|12 Jan 2018|OPEN
Reference quality assembly of the 3.5-Gb genome of Capsicum annuum from a single linked-read library
Amanda M. Hulse-Kemp1,2,3, Shamoni Maheshwari1, Kevin Stoffel1, Theresa A. Hill1, David Jaffe4, Stephen R. Williams4, Neil Weisenfeld4, Srividya Ramakrishnan5, Vijay Kumar4, Preyas Shah4, Michael C. Schatz5, Deanna M. Church4 & Allen Van Deynze1,
1USDAARS Genomics and Bioinformatics Research Unit, Raleigh, NC, USA
2Department of Crop and Soil Sciences, North Carolina State University, Raleigh, NC, USA
3Department of Plant Sciences, University of California, Davis, CA, USA
410x Genomics, Inc, 7068 Koll Center Parkway, Suite 401, Pleasanton, CA, USA
5Department of Computer Science, Johns Hopkins University, Baltimore, MD, USA

Horticulture Research 5,
Article number: 4 (2018)
doi: 10.1038/hortres.2018.4
Views: 619

Received: 05 Jul 2017
Revised: 13 Nov 2017
Accepted: 16 Nov 2017
Published online: 12 Jan 2018

Abstract

Linked-Read sequencing technology has recently been employed successfully for de novo assembly of human genomes, however, the utility of this technology for complex plant genomes is unproven. We evaluated the technology for this purpose by sequencing the 3.5-gigabase (Gb) diploid pepper (Capsicum annuum) genome with a single Linked-Read library. Plant genomes, including pepper, are characterized by long, highly similar repetitive sequences. Accordingly, significant effort is used to ensure that the sequenced plant is highly homozygous and the resulting assembly is a haploid consensus. With a phased assembly approach, we targeted a heterozygous F1 derived from a wide cross to assess the ability to derive both haplotypes and characterize a pungency gene with a large insertion/deletion. The Supernova software generated a highly ordered, more contiguous sequence assembly than all currently available C. annuum reference genomes. Over 83% of the final assembly was anchored and oriented using four publicly available de novo linkage maps. A comparison of the annotation of conserved eukaryotic genes indicated the completeness of assembly. The validity of the phased assembly is further demonstrated with the complete recovery of both 2.5-Kb insertion/deletion haplotypes of the PUN1 locus in the F1 sample that represents pungent and nonpungent peppers, as well as nearly full recovery of the BUSCO2 gene set within each of the two haplotypes. The most contiguous pepper genome assembly to date has been generated which demonstrates that Linked-Read library technology provides a tool to de novo assemble complex highly repetitive heterozygous plant genomes. This technology can provide an opportunity to cost-effectively develop high-quality genome assemblies for other complex plants and compare structural and gene differences through accurate haplotype reconstruction.