Un-zipping Diploid Genomes - Revealing All Kinds of Heterozygous Variants from Comprehensive Haplotig Assembly

Outside of the simplest cases, genomic information is not carried in a single reference per individual, but rather has higher ploidy (n ≥ 2). The existence of two or more highly related sequences within an individual makes it difficult to build high-quality, contiguous genome assemblies from short DNA fragments. Based on earlier work on the polyploidy aware assembler FALCON, we developed new algorithms and software (FALCON-unzip) for de novo haplotype reconstructions from SMRT Sequencing data.

We apply the algorithms and the prototype software for a highly repetitive diploid fungal genome (Clavicorona pyxidata) and an F1 hybrid from two inbred Arabidopsis strains.

For the fungal genome, we achieve an N50 of 1.53 Mb of the ~42 Mb genome and an N50 of the haplotigs of 872 kb. We find that ~45% of the genome is highly heterozygous and ~55% of the genome is highly homozygous. We develop methods to assess the base-level accuracy and local haplotype phasing accuracy of the assembly with short-read data from an Illumina platform. For the Arabidopsis F1 hybrid genome, we find that 80% of the genome can be separated into haplotigs. The long-range accuracy of phasing haplotigs is evaluated by comparing them to the assemblies from the two inbred parental lines.  

The more complete view of both haplotypes can help to generate better annotations, provide useful biological insights, including information about all kinds of heterozygous variants, and be used for studying differential allele expression. Finally, we apply FALCON/FALCON-Unzip on three other organisms (grape, zebra finch, human) to demonstrate this is a general approach to resolve diploid assembly with PacBio long-read data.