Project description:In order to polish a long-read genome assembly, short-read illumina data was obtained from Heterodera schachtii cysts (Woensdrecht population from IRS, the Netherlands). Cysts where obtained from infected plant material. Nematodes were cleaned using a sucrose gradient centrifugation step. Thereafter DNA was extracted and used for library preparation and sequencing by Illumina NextSeq500.
Project description:New tools for improved long-read transcript assembly and coalescence with its short-read counterpart are required. Using our short- and long-read measurements from different cell lines with spiked-in standards, we systematically compared key parameters and biases in the read alignment and assembly of transcripts. We report a cDNA synthesis artifact in long-read datasets that impacts the identity and quantitation of assembled transcripts. We developed a computational pipeline to strand long-read cDNA libraries that markedly improves assembly of transcripts from long-reads. Incorporating stranded long-reads in a new hybrid assembly approach, we demonstrate its efficacy for improved characterization of challenging lncRNA transcripts. Our workflow can be applied to a wide range of transcriptomics datasets for superior demarcation of transcript ends and refined isoform structure, which can enable better differential gene expression analyses and molecular manipulations of transcripts.
Project description:New tools for improved long-read transcript assembly and coalescence with its short-read counterpart are required. Using our short- and long-read measurements from different cell lines with spiked-in standards, we systematically compared key parameters and biases in the read alignment and assembly of transcripts. We report a cDNA synthesis artifact in long-read datasets that impacts the identity and quantitation of assembled transcripts. We developed a computational pipeline to strand long-read cDNA libraries that markedly improves assembly of transcripts from long-reads. Incorporating stranded long-reads in a new hybrid assembly approach, we demonstrate its efficacy for improved characterization of challenging lncRNA transcripts. Our workflow can be applied to a wide range of transcriptomics datasets for superior demarcation of transcript ends and refined isoform structure, which can enable better differential gene expression analyses and molecular manipulations of transcripts.
2023-10-14 | GSE215355 | GEO
Project description:Long-read whole genome sequencing of Aegilops tauschii TA2450
Project description:a chromosome-level nuclear genome and organelle genomes of the alpine snow alga Chloromonas typhlos were sequenced and assembled by integrating short- and long-read sequencing and proteogenomic strategy
Project description:The naked mole-rat (NMR; Heterocephalus glaber) has recently gained considerable attention in the scientific community for its unique potential to unveil novel insights in the fields of medicine, biochemistry, and evolution. NMRs exhibit unique adaptations that include protracted fertility, cancer resistance, eusociality, and anoxia. This suite of adaptations is not found in other rodent species, suggesting that interrogating conserved and accelerated regions in the NMR genome will find regions of the NMR genome fundamental to their unique adaptations. However, the current NMR genome assembly has limits that make studying structural variations, heterozygosity, and non-coding adaptations challenging. We present a complete diploid naked-mole rat genome assembly by integrating long-read and 10X-linked read genome sequencing of a male NMR and its parents, and Hi-C sequencing in the NMR hypothalamus (N=2). Reads were identified as maternal, paternal or ambiguous (TrioCanu). We then polished genomes with Flye, Racon and Medaka. Assemblies were then scaffolded using the following tools in order: Scaff10X, Salsa2, 3d-DNA, Minimap2-alignment between assemblies, and the Juicebox Assembly Tools. We then subjected the assemblies to another round of polishing, including short-read polishing with Freebayes. We assembled the NMR mitochondrial genome with mitoVGP. Y chromosome contigs were identified by aligning male and female 10X linked reads to the paternal genome and finding male-biased contigs not present in the maternal genome. Contigs were assembled with publicly available male NMR Fibroblast Hi-C-seq data (SRR820318). Both assemblies have their sex chromosome haplotypes merged so that both assemblies have a high-quality X and Y chromosome. Finally, assemblies were evaluated with Quast, BUSCO, and Merqury, which all reported the base-pair quality and contiguity of both assemblies as high-quality. The assembly will next be annotated by Ensembl using public RNA-seq data from multiple tissues (SRP061363). Together, this assembly will provide a high-quality resource to the NMR and comparative genomics communities.