De Novo Genome Assembly at Chromosome-Scale of Hermetia illucens (Diptera Stratiomyidae) via PacBio and Omni-C Proximity Ligation Technology

Abstract

Hermetia illucens is a species of great interest for numerous industrial applications. A high-quality reference genome is already available for H. illucens. However, the worldwide maintenance of numerous captive populations of H. illucens, each with its own genotypic and phenotypic characteristics, made it of interest to perform a de novo genome assembly on one population of H. illucens to define a chromosome-scale genome assembly. By combining the PacBio and the Omni-C proximity ligation technologies, a new H. illucens chromosome-scale genome of 888.59 Mb, with a scaffold N50 value of 162.19 Mb, was assembled. The final chromosome-scale assembly obtained a BUSCO completeness of 89.1%. By exploiting the Omni-C proximity ligation technology, topologically associated domains and other topological features that play a key role in the regulation of gene expression were identified. Further, 65.62% of genomic sequences were masked as repeated sequences, and 32,516 genes were annotated using the MAKER pipeline. The H. illucens Lsp-2 genes that were annotated were further characterized, and the three-dimensional organization of the encoded proteins was predicted. A new chromosome-scale genome assembly of good quality for H. illucens was assembled, and the genomic annotation phase was initiated. The availability of this new chromosome-scale genome assembly enables the further characterization, both genotypically and phenotypically, of a species of interest for several biotechnological applications.

Keywords: Hermetia illucens; Larval serum protein-2; de novo genome assembly; gene annotation; topology associated domains.

Figure 1

Hermetia illucens adult (left); larvae and pupae (right).

Figure 2

BUSCO scores of our Hermetia illucens assemblies generated from different databases. The initial draft assembly presented a reduced number of missing genes (M) compared to the subsequent draft and final assemblies. Databases used to assess completeness against eukaryotes, insects, and dipterans: “Eukaryote_odb10”, “insecta_odb10”, and “diptera_odb10” databases, respectively. BUSCO = Benchmarking Universal Single-Copy Orthologs.

Figure 3

Taxon-annotated GC-coverage plot of the initial draft assembly. Each scaffold or contig is represented in the scatter plot by a single filled circle with a diameter proportional to the sequence length and with a color specific to the taxonomic affiliation. Each circle is placed on the X-axis based on its GC proportion and on the Y-axis based on the base coverage of the sequence in the coverage library. The legend in the top right-hand corner shows the number of scaffolds or contig, total span, and N50 of the sequences belonging to each taxonomic group. nt = nucleotides, Kb = kilobase, plot generated using blobtools (v1.1.1).

Figure 4

HiGlass contact map of chromosomal interactions. In the contact map, each point represents how often two regions of the genome were found to be close enough together to be ligated, and regions of the map associated with more intense color represent genomic regions with the greater physical association. In this representation, chromosomes are visible as large squares and are highlighted here with black frames. The length of each chromosome is shown in the contact map above each of them. Mb = megabase.

Figure 5

BUSCO score of Hermetia illucens reference genome and our soft-masked final assembly. Scores were generated from the “insecta_odb10” and “diptera_odb10” datasets. BUSCO = Benchmarking Universal Single-Copy Orthologs.

Figure 6

BUSCO score of the gene annotation made on the soft-masked final assembly. The gene annotation missed the prediction of several conserved genes (12.5 and 18.3% of Insecta and Diptera conserved genes, respectively). Databases used to assess gene annotation completeness against insects and dipterans: “insecta_odb10” and “diptera_odb10” databases, respectively. BUSCO = Benchmarking Universal Single-Copy Orthologs.

Figure 7

Visualization of gene structure for ANN16409. The light blue rectangle represents the CDS of ANN16409 organized into a single exon. The white arrowhead on the left of the light blue rectangle (CDS) represents the gene orientation.

Figure 8

The 3D protein structure prediction of ANN16409 (A) and ANN16413 (B). The color representation of each amino acid depends on the accuracy of its prediction within the protein structure according to the legend on the right. pLDDT = predicted local distance difference test.

Figure 9

Structural alignment of ANN16409 and ANN16413 (A), ANN16409 and dmeLsp-2 (B), and ANN16413 and dmeLsp-2. The structural alignments were created on iCn3D Structure Viewer, using TM-align as the algorithm for protein structure comparisons. RMSD = root mean square deviation, TM-score = template modelling score. In these representations, all matching chains are superposed, while the aligned residues are colored in red for identical residues or blue for non-identical residues. In panel B, non-aligned residue of dmeLsp-2 and ANN16409 are colored light grey and brown, respectively; in panel (C), non-aligned residue of dmeLsp-2 and ANN16413 are colored light grey and brown, respectively.