Phased Diploid Genome Assembly with Single Molecule Real-Time Sequencing

By Chen-Shan Chin, P. Peluso, Fritz J. Sedlazeck, Maria Nattestad, G. Concepcion, Alicia Clum, Christopher Dunn, Ronan C. O’Malley, Rosa Figueroa-Balderas, Abraham Morales-Cruz and 7 more. Nature Methods, 2016

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While genome assembly projects have been successful in many haploid and inbred species, the assembly of noninbred or rearranged heterozygous genomes remains a major challenge. To address this challenge, we introduce the open-source FALCON and FALCON-Unzip algorithms (https://github.com/PacificBiosciences/FALCON/) to assemble long-read sequencing data into highly accurate, contiguous, and correctly phased diploid genomes. We generate new reference sequences for heterozygous samples including an F1 hybrid of Arabidopsis thaliana, the widely cultivated Vitis vinifera cv. Cabernet Sauvignon, and the coral fungus Clavicorona pyxidata, samples that have challenged short-read assembly approaches. The FALCON-based assemblies are substantially more contiguous and complete than alternate short- or long-read approaches. The phased diploid assembly enabled the study of haplotype structure and heterozygosities between homologous chromosomes, including the identification of widespread heterozygous structural variation within coding sequences.

Subfields: Molecular Biology, Plant Science

View Paper: https://www.semanticscholar.org/paper/9af8db5ae66e5009b4deb8bcdc4beb93752e0e74

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Citations in the corpus

Nanopore sequencing and the Shasta toolkit enable efficient de novo assembly of eleven human genomes (2020, background): “We believe that one of the most promising directions for the assembly of diploid samples is the integration of phasing into the assembly algorithm itself, as pioneered by others 17,51,52 .”
Improved maize reference genome with single-molecule technologies (2017, positive): “De novo assembly of the long reads from SMRT Sequencing was performed using two assemblers: the Celera Assembler PBcR –MHAP pipeline 33 and Falcon 34 with different parameter settings.”
Purge Haplotigs: allelic contig reassignment for third-gen diploid genome assemblies (2018, methodology): “Furthermore, recent advances in ‘diploid aware’ genome assemblers have considerably improved the quality of highly heterozygous diploid genome assemblies [4, 5].”
WhatsHap: fast and accurate read-based phasing (2016, background): “It is sometimes also called haplotype assembly, but should not be confused with haplotype-resolved (de novo) genome assembly, which has recently also been explored [12, 13].”
Structural variant calling: the long and the short of it (2019, methodology): “Nevertheless, there are de novo assembly methods to account for this such as trio-sga [78], Falcon-Unzip [79], or Trio-Canu [80] that often require additional coverage and/or parental information.”
Accurate detection of complex structural variations using single molecule sequencing (2017, positive): “With the accuracy established, we next used genuine sequencing of an Arabidopsis thaliana trio (Col-0, CVI and the Col-0 x CVI F1 progeny) 38 and Ashkenazi human trio data set from Genome-in-a-Bottle (GiaB) 39 to assess the recall and Mendelian consistency (Table 1 and Supplementary Notes 4 & 5).”
Merqury: reference-free quality, completeness, and phasing assessment for genome assemblies (2020, methodology): “For a comparison of multiple assemblies, we demonstrate Merqury on haplotype-resolved (TrioCanu [10]), pseudo-haplotype (FALCON-Unzip [9]), and mixed-haplotype (Canu [33]) assemblies of this hybrid genome.”
Assembly of long error-prone reads using de Bruijn graphs (2016, methodology): “A common solution is to mask highly repetitive k -mers as done in the Celera Assembler (31) and Falcon (32).”