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ABSTRACT: Long read sequencing of lentiviral vectors

PROVIDER: PRJEB72210 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	ERR12650417.fastq.gz	Fastqsanger.gz
	ERR12650418.fastq.gz	Fastqsanger.gz
	ERR12650419.fastq.gz	Fastqsanger.gz
	ERR12650420.fastq.gz	Fastqsanger.gz
	ERR12650421.fastq.gz	Fastqsanger.gz

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Project description:Preclinical toxicity analyses of lentiviral vectors expressing the HIV-1 LTR-specific designer-recombinase Brec1

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PHLDA1-PRDM1 mediates the effect of lentiviral vectors on fate-determination of human retinal progenitor cells

Project description:Lentiviral vectors have markedly enhanced gene therapy efficiency in treating congenital diseases, but its long-term safety remains controversial. Most gene therapies for congenital eye diseases need to be carried out at early ages, yet the assessment of related risks to ocular development posed by lentiviral vectors is challenging. Utilizing single-cell transcriptomic profiling on human retinal organoids, this study explored the impact of lentiviral vectors on the retinal development and found that lentiviral vectors can cause retinal precursor cells to shift toward photoreceptor fate through the up-regulation of key fate-determining genes such as PRDM1. Further investigation demonstrated that the intron and intergenic region of PRDM1 was bound by PHLDA1, which was also upregulated by lentiviral vector exposure. Importantly, knockdown of PHLDA1 successfully suppressed the lentivirus-induced differentiation bias of photoreceptor cells. The findings also suggest that while lentiviral vectors may disrupt the fate determination of retinal precursor cells, posing risks in early-stage retinal gene therapy, these risks could potentially be reduced by inhibiting the PHLDA1-PRDM1 signaling axis.

2024-08-28 | GSE251752 | GEO

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Project description:Long-read assembly of Romanomermis culicivorax and Panagrolaimus PS1159

| PRJEB66727 | ENA

Transcriptomic analysis of MYC overexpression in LT-HSC.

Project description:Long-term hematopoietic stem cells (LT-HSC) were sorted from human cord blood and cultured overnight before transduction with lentiviral vectors (empty E CTRL or MYC). Six days after transduction, BFP+ transduced cells were sorted for RNA extraction and sequencing

| EGAS00001004970 | EGA

Transcriptomic analysis of TFEB knockdown in LT-HSC.

Project description:Long-term hematopoietic stem cells (LT-HSC) were sorted from human cord blood and cultured overnight before transduction with lentiviral vectors overexpressing an shRNA against Renilla (shCTRL) or TFEB (shTFEB). Six days later, mCherry+ transduced cells were sorted for RNA extraction and sequencing.

| EGAS00001004967 | EGA

Proteome of the snow alga Chloromonas typhlos

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

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Multi-omic profiling of pathogen-stimulated primary immune cells

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.

2023-09-16 | PXD045237 | Pride

Aberrant splicing isoforms detected by full-length transcriptome sequencing as transcripts of potential neoantigens in non-small cell lung cancer

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

Analysis of lentiviral vector splicing events with nanopore sequencing

Project description:All lentiviral vectors derived from HIV-1 have the major splice donor (SD1) in the 5’ leader region and splice acceptor 7 (SA7) within the env region. Splicing events decrease the amount of full-length RNA available for packaging into virions and could lead to packaging of genomes containing internal deletions. Because there are splice sites or splicing enhancer/silencer elements in both gag and env, we compared how deleting each region affected intracellular genomic RNA splicing in RNA isolated from transfected HEK293Tsa cells. Oxford Nanopore direct cDNA sequencing was used to characterize the splice variants because the long-read length allows full-length transcripts to be analyzed. We show that deleting 507 nt of env, including the SA7, decreased the number of splicing events per transcript and increased the proportion of unspliced genomic RNAs ~3-fold in the cell.

2021-01-30 | GSE165829 | GEO

Expression profiling of Burkitt's lymphoma cells 24h after FOXM1 shRNA or MYB shRNA lentivirus-mediated transduction

Project description:Human Burkitt's lymphoma ST486 cells were transduced with non-target control shRNA lentiviral vectors, FOXM1 shRNA, and MYB shRNA lentiviral vectors. Total RNA was isolated 24h later. cRNA was produced with the standard one-step IVT protocol (Affymetix) and hybridized in U95Av2 gene chips (Affymetrix).

2009-07-27 | GSE17172 | GEO

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