Project description:Here we describe CapTrap-Seq, an experimental workflow designed to address the problem of reduced transcript end detection by long-read RNA sequencing methods, especially at the 5' ends. We apply CapTrap-Seq to profile transcriptomes of the human heart and brain and we compared the obtained results with other library preparation approaches. CapTrap-Seq is a platform-agnostic method and here tested the method by using 3 different long-read sequencing platforms: MinION (ONT), Sequel (PacBaio) and Sequel II (PacBio).
Project description:Two PacBio Hifi sequencing runs from the kidney of a single male NMR sample used to make the mHetGla4.1.primary genome assembly. Specifically, we assembled a second NMR genome from an unrelated male of a separate captive colony in Toronto, Canada, using PacBio HiFi (155.8 Gb, read N50 = 11.45 Kb) and ONT-LSK (299.6 Gb, read N50 = 10.1 Kb) reads (contig N50 = 75.7 Mb, Compleasm S = 98%). This accession stores the Pacbio Hifi data for this independent assembly.
Project description:One PacBio Hifi sequencing run from the kidney of a single male CDMR (Bathyergus suillus) sample used to make the mBatSui1.1.primary genome assembly as an evolutionary comparator to our telomere-to-telomere naked mole-rat genome assembly. Specifically, we assembled a CDMR from a wild-derived sample in South African cape and sequenced in Toronto, Canada, using PacBio HiFi (89 Gb, read N50 = 18 Kb) and ONT-ULK (55 Gb, read N50 = 43 Kb) reads (contig N50 = 33 Mb, Compleasm S = 99%, QV = 71.0). This accession stores the Pacbio Hifi data for this assembly.
Project description:Long-read RNA sequencing technologies offer unparalleled in- sights into transcriptomes by enabling full-length sequencing of RNA molecules, uncovering novel isoforms and alternative splicing events. While long-read sequencing platforms, such as Pacific Biosciences (PacBio) and Oxford Nanopore Technologies (ONT), have historically been associated with higher error rates, recent advancements in both platforms have significantly en- hanced read accuracy, broadening their applicability for tran- scriptomic studies. With the rapid evolution of sequencing protocols and bioin- formatics tools, the trade-offs between sequencing throughput, read length, accuracy, and cost present significant challenges in selecting the optimal approach. Systematic benchmarking studies that compare these options are crucial to inform fu- ture research directions. However, many existing benchmark- ing datasets with matched data across multiple platforms have limitations, including: 1) a lack of realistic biological replicates, which may restrict the generalisability of differential analysis results to real-world scenarios, and 2) the use of earlier sequenc- ing kits, which may not reflect the latest advancements in se- quencing technology, limiting their relevance for future studies that typically use newer sequencing protocols. Here we present LongBench, a comprehensive benchmarking dataset designed to fill these critical gaps. Derived from eight lung cancer cell lines with synthetic RNA spike-ins, LongBench includes bulk, single-cell, and single-nucleus RNA-seq data from three state-of-the-art long-read sequencing platforms — ONT PCR-cDNA, ONT direct RNA, PacBio Kinnex — alongside Il- lumina short-read data for robust cross-platform comparisons. The LongBench dataset is a valuable resource for benchmarking and improving sequencing protocols and bioinformatics tools. With the LongBench dataset we present a systematic evaluation of transcript capture, quantification, and differential expression analyses, examining the strengths and limitations of each se- quencing platform in various biological contexts, enabling re- searchers to make more informed decisions on platform and method selection.
Project description:Long-read sequencing technologies such as Iso-Seq (PacBio Inc.) generate highly accurate sequences of full-length mRNA transcript isoforms. Long-read transcriptomics may be especially useful in the context of lymphocyte functional plasticity as it relates to human health and disease. However, no long-read isoform-aware reference transcriptomes of human circulating lymphocytes seem to be publicly available despite being valuable as benchmarks in a variety of transcriptomic studies. To begin to fill this gap, we purified four lymphocyte subsets (CD4 T, CD8 T, NK, and Pan B cells) from the peripheral blood of a healthy male donor and obtained high-quality RNA (RIN>8) for PacBio Iso-Seq analysis and parallel RNA-Seq analysis.
Project description:Long-read sequencing technologies such as Iso-Seq (PacBio Inc.) generate highly accurate sequences of full-length mRNA transcript isoforms. Long-read transcriptomics may be especially useful in the context of lymphocyte functional plasticity as it relates to human health and disease. However, no long-read isoform-aware reference transcriptomes of human circulating lymphocytes seem to be publicly available despite being valuable as benchmarks in a variety of transcriptomic studies. To begin to fill this gap, we purified four lymphocyte subsets (CD4 T, CD8 T, NK, and Pan B cells) from the peripheral blood of a healthy male donor and obtained high-quality RNA (RIN>8) for PacBio Iso-Seq analysis and parallel RNA-Seq analysis.
2022-09-16 | GSE202327 | GEO
Project description:microbiota in the soil of contaminated site (long-read sequencing by PacBio platform)
Project description:The amoebozoan Acanthamoeba castellanii is an important cause of infectious blindness and a key evolutionary outgroup to humans and fungi. We conducted complementary proteomic and transcriptomic profiling of an Acanthamoeba encystation timecourse. For the transcriptomics, we used both short read RNA-seq (Illumina) and long-read MAS-ISO-seq (PacBio). The long-read MAS-ISO-seq data are reported here.
Project description:Two PacBio Hifi sequencing runs from the kidney of a single male NMR sample used to make the mHetGla4.1.primary genome assembly. Specifically, we assembled a second NMR genome from an unrelated male of a separate captive colony in Toronto, Canada, using PacBio HiFi (155.8 Gb, read N50 = 11.45 Kb) and ONT-LSK (299.6 Gb, read N50 = 10.1 Kb) reads (contig N50 = 75.7 Mb, Compleasm S = 98%). This accession stores the ONT-LSK data for this independent assembly.