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Long-read RNA-Seq analysis of the full-length F8 transcript in iPSCs

ABSTRACT: Long-read RNA-Seq analysis of the full-length F8 transcript in iPSCs

PROVIDER: PRJNA955178 | ENA |

REPOSITORIES: ENA

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Long-read RNA-Seq analysis of the full-length F8 transcript in iPSCs

Project description:We repaired the variant in iPSCs derived from a patient with moderate hemophilia A. To confirm the restoration of the full-length F8 transcript in the gene-corrected iPSCs, we performed long-read RNA-Seq analysis in iPSCs with the EF1α promoter.

2023-04-24 | GSE229621 | GEO

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F8 intein precise CodopN6 reconstitution

Project description:Liver gene therapy with adeno-associated viral (AAV) vectors is under clinical investigation for hemophilia A (HemA), the most common inherited X-linked bleeding disorder. Major limitations are the large size of the F8 transgene, which makes packaging in a single AAV vector a challenge, as well as the development of circulating anti-F8 antibodies which neutralize F8 activity. Taking advantage of split inteins-mediated protein trans-splicing, we divided the coding sequence of the large and highly secreted F8-N6 variant in two separate AAV split-inteins vectors whose co-administration to HemA mice results in F8 protein reconstitution and expression of therapeutic levels of F8 over time without anti-F8 antibodies development.

2022-03-31 | PXD031884 | Pride

Single-molecule, full-length transcript isoform sequencing reveals disease-associated RNA isoforms

Project description:Alternative splicing generates differing RNA isoforms that govern phenotypic complexity of eukaryotes. Its malfunction underlies many diseases, including cancer and cardiovascular diseases. Comparative analysis of RNA isoforms at the genome-wide scale has been difficult. Here, we established an experimental and computational pipeline that accurately quantifies transcript isoforms in their entire length from cDNA sequences with a full-length isoform detection accuracy of 97.6%. We generated a searchable, quantitative human transcriptome annotation with 31,025 known and 5,740 novel transcript isoforms (http://steinmetzlab.embl.de/iBrowser/). By analyzing the isoforms in the presence of RNA Binding Motif Protein 20 (RBM20) mutations associated with aggressive dilated cardiomyopathy (DCM), we identified 121 differentially expressed transcript isoforms in 107 cardiac genes. By establishing an isoform-differential expression test, our approach revealed that 11 of these genes displayed no detectable change in overall RNA expression. However, significant differences in the expression of specific isoforms in these genes was observed. These isoform specific effects demonstrate the need of analyzing RNA isoform expression levels rather than total gene expression levels.

2021-06-16 | E-MTAB-7334 | biostudies-arrayexpress

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Project description:Accurate annotations of genes and their transcripts is a foundation of genomics, but no annotation technique presently combines throughput and accuracy. As a result, the GENCODE reference collection of long noncoding RNAs remains far from complete: many are fragmentary, while thousands more remain uncatalogued. To accelerate lncRNA annotation, we have developed RNA Capture Long Seq (CLS), combining targeted RNA capture with third generation long-read sequencing. We present an experimental re-annotation of the entire GENCODE intergenic lncRNA populations in matched human and mouse tissues. CLS approximately doubles the complexity of targeted loci, both in terms of validated splice junctions and transcript models. Through its identification of full-length transcript models, CLS allows the first definitive measurement of promoter features, gene structure and protein-coding potential of lncRNAs. Thus CLS removes a longstanding bottleneck of transcriptome annotation, generating manual-quality full-length transcript models at high-throughput scales.

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Project description:Selective roles of vertebrate PCF11 in premature and full-length transcript termination (mNET-seq)

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Selective roles of vertebrate PCF11 in premature and full-length transcript termination (ChIP-seq)

Project description:Selective roles of vertebrate PCF11 in premature and full-length transcript termination (ChIP-seq)

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