Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

ABSTRACT: AAV-genome population sequencing detects the repair of mutated ITR structures and the impact of guide RNA cassette designs on vector genome integrity

PROVIDER: PRJNA608034 | ENA |

REPOSITORIES: ENA

ACCESS DATA

Json Xml

Dataset's files

Source:

			Action	DRS
	SRR11171703_subreads.fastq.gz	Fastqsanger.gz
	SRR11171704_subreads.fastq.gz	Fastqsanger.gz
	SRR11171705_subreads.fastq.gz	Fastqsanger.gz
	SRR11171706_subreads.fastq.gz	Fastqsanger.gz
	SRR11171707_subreads.fastq.gz	Fastqsanger.gz

Items per page:

1 - 5 of 9

Similar Datasets

Adeno Associated Virus (AAV)

Project description:AAV-genome population sequencing detects the repair of mutated ITR structures and the impact of guide RNA cassette designs on vector genome integrity

| PRJNA607794 | ENA

A Self-Complementary AAV Proviral Plasmid System to Reduce Aberrant Cross-Packaging and ITR Promoter Activity in AAV Vector Preparations (Long-read)

Project description:Adeno-associated viral vectors (AAV) are a leading delivery system for gene therapy in animal models and humans. With several FDA-approved AAV gene therapies on the market, issues related to vector manufacturing have become increasingly important. In this study, we focused on potentially toxic DNA contaminants that can arise from AAV proviral plasmids, the raw materials required for manufacturing recombinant AAV in eukaryotic cells. Typical AAV proviral plasmids are circular DNAs containing a therapeutic gene cassette flanked by natural AAV inverted terminal repeat (ITR) sequences, and a plasmid backbone carrying prokaryotic sequences required for plasmid replication and selection in bacteria. While the majority of AAV particles package the intended therapeutic payload, some capsids instead package the bacterial sequences located on the proviral plasmid backbone. Since ITR sequences also have promoter activity, potentially toxic bacterial open reading frames can be produced in vivo, thereby representing a safety risk. In this study, we describe a new AAV proviral plasmid for vector manufacturing that (1) significantly decreases cross-packaged bacterial sequences; (2) increases correctly packaged AAV payloads; and (3) blunts ITR-driven transcription of cross-packaged material to avoid expressing potentially toxic bacterial sequences. This system may help improve the safety of AAV vector products.

2024-07-26 | GSE272650 | GEO

A Self-Complementary AAV Proviral Plasmid System to Reduce Aberrant Cross-Packaging and ITR Promoter Activity in AAV Vector Preparations (Short-read)

2024-07-26 | GSE272649 | GEO

On the design of CRISPR-based single cell molecular screens

Project description:We demonstrate that vector designs for such screens that rely on cis linkage of guides and distally located barcodes suffer from swapping of intended guide-barcode associations at rates approaching 50% due to template switching during lentivirus production, greatly reducing sensitivity.

2018-01-29 | GSE108699 | GEO

Identification of liver-specific enhancer-promoter activity in the 3’ UTR of the wild-type AAV2 genome

Project description:We describe and functionally characterize a previously unknown liver-specific enhancer-promoter element in the wild-type AAV2 (wtAAV2) genome lying between the cap stop codon and right-hand inverted terminal repeat (ITR). Remarkably, this element falls within the 163-nucleotide common insertion region of the AAV genome implicated in HCC oncogenesis, thereby providing a mechanistic explanation for the participation of AAV integration events in the development of HCC.

2017-06-12 | GSE93341 | GEO

A Self-Complementary AAV Proviral Plasmid System to Reduce Aberrant Cross-Packaging and ITR Promoter Activity in AAV Vector Preparations (Long-read)

Project description:A Self-Complementary AAV Proviral Plasmid System to Reduce Aberrant Cross-Packaging and ITR Promoter Activity in AAV Vector Preparations (Long-read)

| PRJNA1137884 | ENA

A Self-Complementary AAV Proviral Plasmid System to Reduce Aberrant Cross-Packaging and ITR Promoter Activity in AAV Vector Preparations (Short-read)

Project description:A Self-Complementary AAV Proviral Plasmid System to Reduce Aberrant Cross-Packaging and ITR Promoter Activity in AAV Vector Preparations (Short-read)

| PRJNA1137885 | ENA

Human α-synuclein overexpression upregulates SKOR1 in a rat model of simulated nigrostriatal aging

Project description:Parkinson’s disease is characterised by progressive loss of dopaminergic (DA) neurons from the substantia nigra (SN) and intraneuronal accumulation of α-synuclein (αSyn) protein. Age is the biggest risk factor for Parkinson’s and has been suggested to create a vulnerable pre-parkinsonian state, but the drivers of this association are unclear. It is known that aging leads to increased αSyn expression in DA neurons and it is possible that this may alter molecular processes that are central to maintaining nigrostriatal integrity. To investigate this, adult female Sprague-Dawley rats received unilateral intranigral injection of adeno-associated viral (AAV) vector carrying wild-type human αSyn (AAV-αSyn) or control vector (AAV-Null). AAV-αSyn induced no detrimental effects on motor behaviour, but there was expression of human wild-type αSyn throughout the midbrain and ipsilateral striatum at 20 weeks post-surgery. Microarray analysis revealed that the gene most-upregulated in the ipsilateral SN of the AAV-αSyn group was the SKI Family Transcriptional Corepressor 1 (SKOR1). Bioenergetic state analysis of mitochondrial function found that SKOR1 overexpression reduced the maximum rate of cellular respiration. Furthermore, SKOR1 impaired neurite growth to the same extent as α-synuclein, and inhibited BMP-SMAD-dependent transcription, a pathway that promotes DA neuron survival and growth. Given the normal influence of ageing on DA neuron loss in human SN, the extent to which αSyn upregulates SKOR1 may influence whether an individual undergoes normal nigrostriatal ageing or reaches a threshold for prodromal PD. This provides new insight into mechanisms through which age-related changes may influence molecular mechanisms controlling neuronal integrity across the lifespan.

2024-08-08 | GSE252918 | GEO

AAV vector-derived elements integrate into Cas9-generated double strand breaks and disrupt gene transcription

Project description:We previously found that an adeno-associated virus (AAV) vector containing S. aureus Cas9 and a multi-target guide (g)RNA could integrate into the genome and prematurely terminate transcription of Ube3a-ATS, a long non-coding RNA. Here, we assessed the performance of three additional AAV vectors containing S. aureus Cas9 and twenty-five vectors containing N. meningococcus Cas9, all targeting single sites within Ube3a-ATS. We found that none of these single-target gRNA vectors were as effective as multi-target gRNA vectors at reducing Ube3a-ATS expression in neurons. We also developed a new anchored multiplex PCR sequencing (AMP-seq) method and analysis pipeline to quantify the relative frequency of all possible editing events at target sites, including unresolved double-stranded breaks (DSBs) and capture of foreign DNA. We found that integration of AAV was the most frequent editing event (67-89% of all edits) at three different single target sites, far surpassing insertions and deletions (indels). None of the most frequently observed indels was capable of blocking transcription when incorporated into a Ube3a-ATS minigene reporter, whereas two vector derived elements—the polyA and reverse promoter—reduced downstream transcription by up to 50%. Since not all editing events disrupt gene transcription, our findings suggest that the probability that a gene trapping AAV integration event occurs is influenced by which vector-derived element(s) are integrated and by the number of target sites.

2024-10-16 | GSE271068 | GEO

Generation of LAMC2-EGFP knock-in human pancreatic cancer cells

Project description:Since LAMC2 is a secreted molecule present in the extracellular matrix of the cells, was designed a strategy based on CRISPR/Cas9-mediated homologous recombination to mark LAMC2 cells in human PDACs. A Cas9 single-guide RNAs complementary to sequences overlapping the stop codon of the LAMC2 locus was designed and a donor vector that contained LAMC2 homology arms flanking an EGFP reporter cassette positioned immediately upstream of the stop codon was generated. LF2A self-cleavage peptide in frame with EGFP so that LAMC2-EGFP locus was expressed as a single mRNA was added, whereas the resulting polypeptide was cleaved in the two encoded proteins, LAMC2 and EGFP. L3.6pl and PANC-1 cells were nucleofected with the donor vector together with a guide-RNA-Cas9 (guide). The engineered cells were subcutaneously injected in CD1 male mice and RNA-seq was performed on LAMC2-EGFP+ and EGFP- -derived tumors.

2022-04-11 | E-MTAB-11597 | biostudies-arrayexpress

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data