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:Primary objectives: The primary objective is to investigate circulating tumor DNA (ctDNA) via deep sequencing for mutation detection and by whole genome sequencing for copy number analyses before start (baseline) with regorafenib and at defined time points during administration of regorafenib for treatment efficacy in colorectal cancer patients in terms of overall survival (OS).
Primary endpoints: circulating tumor DNA (ctDNA) via deep sequencing for mutation detection and by whole genome sequencing for copy number analyses before start (baseline) with regorafenib and at defined time points during administration of regorafenib for treatment efficacy in colorectal cancer patients in terms of overall survival (OS).
Project description:Human utilization of the mulberry-silkworm interaction started at least 5,000 years ago and greatly influenced world history through the Silk Road. Complementing the silkworm genome sequence, here we describe the genome of a mulberry species (Morus notabilis C. K. Schneider). In the 330 Mb genome assembly of M. notabilis, we identified 128 Mb of repetitive sequences and 29,338 genes, 60.8% of which were supported by transcriptome sequencing. Mulberry gene sequences appear to evolve ~3 times faster than other Rosales, perhaps facilitating its spread to Europe, Africa, and America. It is among few eudicots but several Rosales not preserving genome duplications in more than 100 million years – however neopolyploid series in mulberry and several others suggest that new duplications may confer benefits. Strikingly, five predicted mulberry miRNAs were found in the hemolymph and silkglands of silkworm, suggesting profound molecular level interactions that promise to expand knowledge of plant-herbivore relationship which constitute key elements of most terrestrial habitats. In addition, we investigated the characters of hemolymph small RNA. small mRNA profiles of silkworm hemolymph in the fifth instar day-5 silkworm were generated by deep sequencing, in twice, using Illumina Hiseq 2000.
Project description:We applied Single Molecule Real-Time long-read whole-genome sequencing in Dux knockout mouse and confirmed the success of our Dux knockout mouse model.
Project description:Mulberry (Morus alba L.) is considered a millenary medicinal plant and a food source for silkworms. Different M. alba extracts offer a variety of biological and pharmacological properties that are largely attributed to stilbenoids, a small group of phenylpropanoids, including resveratrol and oxyresveratrol. These are naturally present in non-renewable parts of mulberry trees, impeding their efficient extraction. As a way to bypass this spatiotemporal restriction, we generated cell suspensions from mulberry leaf petioles and demonstrated that the combined use of methyl jasmonate and two different types of cyclodextrins were able to elicit a high production of resveratrol and oxyresveratrol. As oxyresveratrol-producing enzymes are still unknown, we improved the structural and functional annotation of the mulberry genome by integrating short and long-read sequencing data and combined it with time series transcriptome, metabolite and proteome data in response to cell elicitation to identify a complete set of phenylpropanoid and stilbenoid related genes. These included 23 stilbene synthase (STS) genes and a group of six p-coumaroyl-CoA 2'-hydroxylases (C2’Hs), all being highly co-expressed with resveratrol and oxyresveratrol production. We transiently transformed grapevine (Vitis vinifera L.) calli and Nicotiana benthamiana plants to functionally validate the role of C2’Hs in the first committed step of oxyresveratrol synthesis, which provides an alternative substrate for STSs by hydroxylating p-coumaroyl-coA into 2’4’-dihydroxycinnamoyl-CoA. Finally, we offer tools for genomic and transcriptomic exploration also in the context of jasmonate elicitation aiding in the characterization of novel stilbenoid-modifying and regulatory genes in the Morus genus.
Project description:Adenovirus is a common human pathogen that relies on host cell processes for transcription and processing of viral RNA and protein production. Although adenoviral promoters, splice junctions, and cleavage and polyadenylation sites have been characterized using low-throughput biochemical techniques or short read cDNA-based sequencing, these technologies do not fully capture the complexity of the adenoviral transcriptome. By combining Illumina short-read and nanopore long-read direct RNA sequencing approaches, we mapped transcription start sites and cleavage and polyadenylation sites across the adenovirus genome. In addition to confirming the known canonical viral early and late RNA cassettes, our analysis of splice junctions within long RNA reads revealed an additional 35 novel viral transcripts. These RNAs include fourteen new splice junctions which lead to expression of canonical open reading frames (ORF), six novel ORF-containing transcripts, and fifteen transcripts encoding for messages that potentially alter protein functions through truncations or fusion of canonical ORFs. In addition, we also detect RNAs that bypass canonical cleavage sites and generate potential chimeric proteins by linking separate gene transcription units. Of these, an evolutionary conserved protein was detected containing the N-terminus of E4orf6 fused to the downstream DBP/E2A ORF. Loss of this novel protein, E4orf6/DBP, was associated with aberrant viral replication center morphology and poor viral spread. Our work highlights how long-read sequencing technologies can reveal further complexity within viral transcriptomes.
Project description:Human utilization of the mulberry-silkworm interaction started at least 5,000 years ago and greatly influenced world history through the Silk Road. Complementing the silkworm genome sequence, here we describe the genome of a mulberry species (Morus notabilis C. K. Schneider). In the 330 Mb genome assembly of M. notabilis, we identified 128 Mb of repetitive sequences and 29,338 genes, 60.8% of which were supported by transcriptome sequencing. Mulberry gene sequences appear to evolve ~3 times faster than other Rosales, perhaps facilitating its spread to Europe, Africa, and America. It is among few eudicots but several Rosales not preserving genome duplications in more than 100 million years – however neopolyploid series in mulberry and several others suggest that new duplications may confer benefits. Strikingly, five predicted mulberry miRNAs were found in the hemolymph and silkglands of silkworm, suggesting profound molecular level interactions that promise to expand knowledge of plant-herbivore relationship which constitute key elements of most terrestrial habitats. In addition, we investigated the characters of hemolymph small RNA.
Project description:Objectives: To perform long-read transcriptome and proteome profiling of pathogen-stimulated peripheral blood mononuclear cells (PBMCs) from healthy donors. We aim to discover new transcripts and protein isoforms expressed during immune responses to diverse pathogens. Methods: PBMCs were exposed to four microbial stimuli for 24 hours: the TLR4 ligand lipopolysaccharide (LPS), the TLR3 ligand Poly(I:C), heat-inactivated Staphylococcus aureus, Candida albicans, and RPMI medium as negative controls. Long-read sequencing (PacBio) of one donor and secretome proteomics and short-read sequencing of five donors were performed. IsoQuant was used for transcriptome construction, Metamorpheus/FlashLFQ for proteome analysis, and Illumina short-read 3’-end mRNA sequencing for transcript quantification. Results: Long-read transcriptome profiling reveals the expression of novel sequences and isoform switching induced upon pathogen stimulation, including transcripts that are difficult to detect using traditional short-read sequencing. We observe widespread loss of intron retention as a common result of all pathogen stimulations. We highlight novel transcripts of NFKB1 and CASP1 that may indicate novel immunological mechanisms. In general, RNA expression differences did not result in differences in the amounts of secreted proteins. Interindividual differences in the proteome were larger than the differences between stimulated and unstimulated PBMCs. Clustering analysis of secreted proteins revealed a correlation between chemokine (receptor) expression on the RNA and protein levels in C. albicans- and Poly(I:C)-stimulated PBMCs. Conclusion: Isoform aware long-read sequencing of pathogen-stimulated immune cells highlights the potential of these methods to identify novel transcripts, revealing a more complex transcriptome landscape than previously appreciated.