Enhanced genome assembly and a new official gene set for Tribolium castaneum

Abstract

Background: The red flour beetle Tribolium castaneum has emerged as an important model organism for the study of gene function in development and physiology, for ecological and evolutionary genomics, for pest control and a plethora of other topics. RNA interference (RNAi), transgenesis and genome editing are well established and the resources for genome-wide RNAi screening have become available in this model. All these techniques depend on a high quality genome assembly and precise gene models. However, the first version of the genome assembly was generated by Sanger sequencing, and with a small set of RNA sequence data limiting annotation quality. Results: Here, we present an improved genome assembly (Tcas5.2) and an enhanced genome annotation resulting in a new official gene set (OGS3) for Tribolium castaneum, which significantly increase the quality of the genomic resources. By adding large-distance jumping library DNA sequencing to join scaffolds and fill small gaps, the gaps in the genome assembly were reduced and the N50 increased to 4,753kbp. The precision of the gene models was enhanced by the use of a large body of RNA-Seq reads of different life history stages and tissue types, leading to the discovery of 1,452 novel gene sequences. We also added new features such as alternative splicing, well defined UTRs and microRNA target predictions. For quality control, 399 gene models were evaluated by manual inspection. The current gene set was submitted to Genbank and accepted as a RefSeq genome by NCBI. Conclusions: The new genome assembly (Tcas5.2) and the official gene set (OGS3) provide enhanced genomic resources for genetic work in Tribolium castaneum. The much improved information on transcription start sites supports transgenic and gene editing approaches. Further, novel types of information such as splice variants and microRNA target genes open additional possibilities for analysis.

CenH3 distribution reveals extended centromeres in the model beetle Tribolium castaneum

“…We performed TBLASTN search using cCENH3 protein sequence to query the latest T . castaneum genome assembly Tcas5.2 [ 24 ] in order to examine whether multiple copies of the gene are present in the genome. The TBLASTN search mapped the cCENH3 gene with 100% identity (E-value 1e-67) through entire length of the sequence only to one location at chromosome 9 (positions LG9:11651877–11652254) spanning a 378 bp long sequence.…”

“…The T . castaneum genome has been assembled into ten chromosome/linkage groups, but the current Tcas5.2 assembly lacks 20% of the genome sequence attributed to the heterochromatic (peri)centromeric regions [ 23 , 24 , 26 ], so it could not serve as a reliable repeat database reference. Therefore, we re-sequenced the T .…”

“…We identified the gene ID TC012577 from the current T . castaneum genome assembly Tcas5.2 [ 24 ] as the CenH3 homologue, and its coding sequence is identical to a candidate computationally predicted by Drinnenberg and coauthors [ 6 ]. By demonstrating cCENH3 chromosomal position, we validated cCENH3 experimentally as an authentic centromeric variant of histone H3.…”

“…In spite of that, 20% of the genome (44 Mb out of 204 Mb), primarily from the (peri)centromere regions, remained unmapped to genome assembly. The genome assembly with the contigs assembled into 10 chromosome groups was recently upgraded to the improved version Tcas5.2 [ 24 ], but the centromeric gaps are still unpatched. It was experimentally estimated that 17% of the T .…”

Metabolic and transcriptome responses of RNAi-mediated AMPKα knockdown in Tribolium castaneum

“…To ensure the accuracy of sequence alignment, raw reads were cleaned by removing adapter and primer sequences, reads with ambiguous nucleotides larger than 5% and low-quality reads. Clean reads were aligned to the reference genome of T. castaneum (Reference genome version: GCF_000002335.3_Tcas5.2_genomic.fna; Reference genome site: ftp://ftp.ncbi.nlm.nih.gov/genomes/all/GCF/000/002/335/GCF_000002335.3_Tcas5.2/GCF_000002335.3_Tcas5.2_genomic.fna.gz ) [ 21 ] using HISAT2 (CCB, Johns Hopkins University, USA). Subsequently, the transcripts were subjected to blast against GO, KOG and KEGG databases.…”