Project description:To identify aberrant splicing isoforms and potential neoantigens, we performed full-length cDNA sequencing of lung adenocarcinoma cell lines using a long-read sequencer MinION. We constructed a comprehensive catalog of aberrant splicing isoforms and detected isoform-specific peptides using proteome analysis.
2020-12-09 | PXD019915 | JPOST Repository
Project description:full-length isoform of endosperm from Nongda3672
Project description:To investigate the impact of full-length (1-23ex) and short isoform (7-20ex) PIWIL2 expression, we established by knock-in (KI) the HEK293 cell lines, expressed full-length (1-23ex) and short isoform (7-20ex) PIWIL2. KI_pCI is knock-in control.
Project description:iRNA approaches were used to knock down the expression of dystrophin (DMD), including its full-length isoform DP427, and utrophin (UTRN) genes. Specifically, iRNA-mediated knockdown of both DMD (full-length dystrophin DP427) and UTRN was performed in HSkM myotubes. It is known that the shorter dystrophin isoform DP71 compensates for full-length dystrophin DP427 in the early stages of myofiber differentiation in cell models of Duchenne muscular dystrophy. Accordingly, DP427-deficient and DP71/DP427-deficient HSkM myotubes exhibit distinct changes in the expression of genes involved in membrane integrity and cytoskeletal functions. RNA-sequencing analyses revealed overlapping transcriptomic alterations in these HSkM cell lines, which corresponded to observed phenotypic abnormalities such as increased membrane permeability and intracellular calcium levels, mitochondrial aggregation, elevated reactive oxygen species (ROS), enhanced cyto- and genotoxicity, and induction of apoptosis.
Project description:Aging is a multifactorial process accompanied by profound alterations in endocrine function, which plays a central role in maintaining whole-body physiology and metabolism. Although age-related changes in pituitary hormone levels, including growth hormone (GH), have been documented, the molecular mechanisms underlying pituitary aging and its effects on full-length transcript isoforms remain poorly understood. Here, we performed full-length long-read transcriptomic sequencing (Iso-Seq) of anterior pituitary glands from young and old male mice to comprehensively characterize aging-associated changes in gene expression, isoform usage, and alternative splicing. Integration of differential expression, isoform-switch, and novel isoform analyses identified Pou1f1 as the most significantly downregulated gene in aged pituitaries (log₂FC = −2.02, Padj = 1.02 × 10⁻³⁰), accompanied by reduced expression of genes involved in neurosecretory function, including Syn1, Syt7, Dvl3, and Cacna2d2. Notably, aging was associated with a directional accumulation of novel isoforms, with more isoforms enriched in old than young pituitaries (100 vs. 55; binomial P = 0.0004), together with a global shift toward shorter 3′ UTRs (9.6% reduction in length). Together, these findings provide a high-resolution full-length transcriptomic landscape of pituitary aging and reveal previously unrecognized changes in transcript isoform composition and 3′ UTR architecture that may contribute to age-related endocrine dysfunction.
Project description:We developed a hybrid-sequencing workflow, combining next-generation and third-generation sequencing, to reconstruct full-length transcriptomes. Integrating with polysome profiling and ribosome footprinting data, we predicted isoform–specific translational status and reconstructed ORFeome. Moreover, we identified isoforms with specific subcellular localization pattern in neurons.
Project description:We consturcted a full-length transcript reference of CD4SP and CD8SP T cells using TGS (PacBio) data. We then used SGS (Illumina) CD4SP and CD8SP data to quantify isoform expressions and study alternative splicing patterns against the full-length transcript reference between CD4SP and CD8SP cells.
Project description:Single-cell isoform regulation is increasingly being studied. To get a full view of alternative isoform usage, from transcription start site and alternative splicing to transcription termination site, full-length sequences have to be studied. Here we use PacBio long read sequencing technology combined with unique molecular identities to get a full and accurate picture of alternative isoform usage of different stages in maturing oligodendrocyte single cells. We see that the majority of molecules in single-cells are separate isoforms, even after applying a conservative definition of what constitutes an isoform. Few isoforms are common between cells of the same cell-type but the common isoforms are higher expressed. We also see that exon junctions in coding regions are better regulated than exon junctions in non-coding regions and that genes often express more than one coding isoform.