Single-Molecule Sequencing of the Drosophila serrata Genome

Abstract

Long-read sequencing technology promises to greatly enhance de novo assembly of genomes for nonmodel species. Although the error rates of long reads have been a stumbling block, sequencing at high coverage permits the self-correction of many errors. Here, we sequence and de novo assemble the genome of Drosophila serrata, a species from the montium subgroup that has been well-studied for latitudinal clines, sexual selection, and gene expression, but which lacks a reference genome. Using 11 PacBio single-molecule real-time (SMRT cells), we generated 12 Gbp of raw sequence data comprising ∼65 × whole-genome coverage. Read lengths averaged 8940 bp (NRead50 12,200) with the longest read at 53 kbp. We self-corrected reads using the PBDagCon algorithm and assembled the genome using the MHAP algorithm within the PBcR assembler. Total genome length was 198 Mbp with an N50 just under 1 Mbp. Contigs displayed a high degree of chromosome arm-level conservation with the D. melanogaster genome and many could be sensibly placed on the D. serrata physical map. We also provide an initial annotation for this genome using in silico gene predictions that were supported by RNA-seq data.

Whole genome sequences of 23 species from the Drosophila montium species group (Diptera: Drosophilidae): A resource for testing evolutionary hypotheses

“…Estimated genome sizes within the montium group range from 155.1 Mb to 223.4 Mb (mean=196.4 Mb; median=198.1 Mb). These sizes are consistent with the previously assembled D. kikkawai [3] and D. serrata [19] genomes, with total sequence lengths of 164.3 Mb and 198.0 Mb, respectively. Our own D. serrata assembly (strain 14028-0681.02) has an estimated genome size of 184.7 Mb.…”

“…Our own D. serrata assembly (strain 14028-0681.02) has an estimated genome size of 184.7 Mb. The relatively small difference between our genome size estimate and the total contig length of the previously published PacBio D. serrata assembly (strain Fors4) [19] is likely a product of the imprecision of k -mer frequency spectrum-based genome size estimates, along with strain-level differences in genome size. Across all montium species, estimated genome size is strongly positively correlated with repeat content ( r =0.88, p <1e-06).…”

“…This produced a 164 Mb assembly with a scaffold N50 of 904 kb. The D. serrata genome [19] was sequenced to a depth of 63x coverage using PacBio long-reads. It yielded a 198 Mb assembly with a contig N50 of 943 kb.…”

Sex‐biased transcriptome divergence along a latitudinal gradient

“…Each EST was linked to an annotated feature from the draft D. serrata genome assembly (Allen et al . ; doi: ) via tblastx (NCBI stand‐alone blast version 2.3.0+). All ESTs were successfully linked to a D. serrata feature with a median e‐value of 3.76e −137 .…”

“…To determine whether genes underlying specific biological functions were more likely to have diverged than others, we performed gene ontology (GO) term enrichment analysis after assigning functions to D. serrata ESTs based on homolog identification as follows (Table S1, Supporting information). Each EST was linked to an annotated feature from the draft D. serrata genome assembly (Allen et al 2017;doi: 10.1101/090969) via tblastx (NCBI standalone blast version 2.3.0+). All ESTs were successfully linked to a D. serrata feature with a median e-value of 3.76e À137 .…”