Originally sourced in Germany. Procured commercially: Innovative Environmental services (IES) Ltd, Switzerland.
Next Generation Sequencing
i) Illumina Hi-C sequencing 150 bp paired end data:
925,594,532 reads and 723x coverage.
ii) PacBio HiFi data, of mean read length 14,960, total reads 615,477, read length N50 16,105, and total bases 9,207,869,729. 450ng extracted using MagAttract method from a single individual at Rothamsted Research.
DNA was extracted using a modified MagAttract method from the published Mosquito protocol (https://www.biorxiv.org/content/10.1101/499954v1). The volume was 30ul, concentration 15ng/ul by Qubit analysis at Rothamsted (450ng gDNA).
Single female individual DNA used for PacBio HiFi and Hi-C Illumina sequencing (University College London, Arima Genomics respectively). Hifiasm was used to assemble the PacBio HiFi, with Juicer and 3d-dna and Hi-C data used for chromosome level assembly. Haplotigs were removed (purge_haplotigs). Unmapped reads were mapped back to the original assembly to check for missing sequence and incorporated into the final assembly. Manual curation was done to bring the genome together and check for miss-assemblies.
Public RNA-seq was assembled (BUSCO: C:94.7%[S:53.7%,D:41.0%],F:0.4%,M:4.9%) and used in the Maker2 annotation pipeline with trained Augustus and Genemark gene predictors. Data: PRJEB15223 (Larvae), PRJNA166085 (egg ropes, all four larval stages, pupae and male and female adults. Also included are larvae exposed to different concentrations of several model toxicants), PRJNA229141 (anterior and posterior early embryo), PRJNA675286 (larvae - TMO/LCO exposure). PASA was used to update the gene models to add UTR, correct existing models and add isoforms. Non-coding RNA was annotated using Infernal v1.1.4.
A Pfam genomic track was created by converting to six reading frames and utilizing hmmer to identify loci of interest. Using this information, loci of interest including UDP, P450, ABC and IRAC gene models were found and curated using mapped RNA-seq and a Maker gene annotation.
Two Endosymbionts were assembled which included an unknown Enterobacter (1,661,850 bp) and Wolbachia (559,667 bp).