Similar Datasets
Project description:Recombinant AAV vectors have the unique ability to promote targeted integration of transgenes via homologous integration at specified genomic sites reaching frequencies of 0.1-1%. We studied genomic parameters that influence targeting efficiencies on a large scale. To do this, we generated more than 1000 engineered, doxycycline-inducible target sites in the human HAP1 cell line and infected this polyclonal population with a library of AAV-DJ targeting vectors each carrying a unique barcode. The heterogeneity of barcode integration at each target site provided an assessment of the targeting efficiency at that locus. We compared targeting efficiency with and without target site transcription for identical chromosomal positions, finding that targeting efficiency was enhanced by target site transcription. Chromatin states associated with active transcription were also predictive of higher targeting efficiency. Furthermore, there was an effect on the amenability of a site to targeting due to other factors such as the level of transcription from intersecting genes. These results define important parameters that may not only assist in designing optimal targeting vectors for genome editing, but also provide new insights into the mechanism of AAV-mediated homologous recombination.
2020-06-04 | GSE151740 | GEO
Project description:We have previously developed a modified iteration of a viral chromosome conformation capture (V3C-seq) assay to show that the autonomous parvovirus Minute Virus of Mice (MVM) localizes spatially with cellular sites of DNA damage to establish viral replication centers. Similar V3C-seq assays to map AAV genome localization show that both replicating and non-replicating AAV2 genomes in the absence of helper virus colocalize with cellular sites of DNA damage. The AAV non-structural protein Rep 68/78, when ectopically expressed in the absence of viral infection or during AAV2 infection in the absence of helper proteins also localizes to cellular sites of DNA damage. Strikingly however, recombinant AAV gene therapy vector genomes derived from AAV do not colocalize with AAV and Rep at cellular DDR sites.
2021-10-20 | GSE178316 | GEO
Project description:To study monocyte and macrophage activation in ANCA-associtated vasculitis (AAV), we performed bulk RNA sequencing of bead-selected monocytes and in vitro cultured monocyte-derived macrophages from AAV patients and healthy controls. Overview patients included for sequencing monocytes: - AAV active disease, n=4, MPO-AAV=4 - AAV remission, n=10, PR3-AAV=5, MPO-AAV=5 - Healthy controls, n=6 Overview patients included for sequencing monocyte-derived macrophages: - AAV active, n=1, PR3-AAV=1 - AAV remission, n=3, PR3-AAV=3 - Healthy controls, n=3
2024-06-20 | E-MTAB-13676 | biostudies-arrayexpress
Project description:Deciphering patterns of connectivity between neurons in the mammalian brain is a critical step toward understanding brain function. Conventional imaging based neuroanatomical tracing methods identify area-to-area or sparse neuron-to-neuron connectivity patterns, but with extremely limited throughput. Recently developed barcode-based connectomics methods can efficiently map large numbers of single-neuron projections, but linking these data to single-cell transcriptomics remains a challenge. Here, we established a retro-AAV barcode-based multiplexed tracing method called MERGE-seq (Multiplexed projection neuRons retroGrade barcodE sequencing), which is capable of simultaneously characterizing the projectome and transcriptome at the single neuron level. We uncovered dedicated and collateral projection patterns of ventromedial prefrontal cortex (vmPFC) neurons to five downstream targets (AI, DMS, BLA, MD and LH). We found that projection-defined vmPFC neurons are molecularly heterogeneous, which are composed of different neuronal subtypes. We further identified transcriptional signatures of various dedicated and bifurcated vmPFC neurons, and verified Pou3f1 as the marker gene of neurons sending collateral axons to DMS and LH. Finally, we fitted our single-neuron connectome/transcriptome data into a machine learning-based model and revealed groups of genes that were predictive of certain projection pattern. In summary, we have developed a new multiplexed technique whose paired connectome and gene expression data can help reveal organizational principles that form neural circuits and process information.
2024-02-25 | GSE210173 | GEO
Project description:Deciphering patterns of connectivity between neurons in the mammalian brain is a critical step toward understanding brain function. Conventional imaging based neuroanatomical tracing methods identify area-to-area or sparse neuron-to-neuron connectivity patterns, but with extremely limited throughput. Recently developed barcode-based connectomics methods can efficiently map large numbers of single-neuron projections, but linking these data to single-cell transcriptomics remains a challenge. Here, we established a retro-AAV barcode-based multiplexed tracing method called MERGE-seq (Multiplexed projection neuRons retroGrade barcodE sequencing), which is capable of simultaneously characterizing the projectome and transcriptome at the single neuron level. We uncovered dedicated and collateral projection patterns of ventromedial prefrontal cortex (vmPFC) neurons to five downstream targets (AI, DMS, BLA, MD and LH). We found that projection-defined vmPFC neurons are molecularly heterogeneous, which are composed of different neuronal subtypes. We further identified transcriptional signatures of various dedicated and bifurcated vmPFC neurons, and verified Pou3f1 as the marker gene of neurons sending collateral axons to DMS and LH. Finally, we fitted our single-neuron connectome/transcriptome data into a machine learning-based model and revealed groups of genes that were predictive of certain projection pattern. In summary, we have developed a new multiplexed technique whose paired connectome and gene expression data can help reveal organizational principles that form neural circuits and process information.
2024-02-25 | GSE210172 | GEO
Project description:To explore how Mier1 affects liver regeneration, we specifically knocked out Mier1 in mouse liver through adeno-associated virus (AAV). The mice we used were knocked in a Cre-induced Cas9 expression cassette. Through tail vein injection, we delivered the AAV expressing Cre-recombinase under TBG promoter, and sgRNA targeting Mier1 (AAV-Mier1 sgRNA), into the adult Cas9 knockin mice to knock out the Mier1 gene in liver. AAV vectors with no sgRNA inserted (AAV-Cre) were used in control animals. To assess the role of MIER1 in liver regeneration, we performed 70% partial hepatectomy, 3 weeks after AAV injection. We then performed ATAC-seq in Control and Mier1 liver-sepcific knockout mice liver tissues at 0 h and 24 h after PHx to explore the influence of Mier1 on the chromation opening during liver regeneration.
2022-12-31 | GSE212626 | GEO
Project description:Genome wide DNA methylation profiling of 4 human tissues of various ages: 83 samples of normal adjacent resected kidney tissue .The Illumina Infinium 27k Human DNA methylation Beadchip v1.2 was used to obtain DNA methylation profiles across approximately 27,000 CpGs. Sentrix barcode provided for batch normalization
2013-08-16 | GSE49907 | GEO
Project description:CRISPR-Cas9 delivery by AAV holds promise for gene therapy but faces critical barriers due to its potential immunogenicity and limited payload capacity. Here, we demonstrate genome engineering in postnatal mice using AAV-split-Cas9, a multi-functional platform customizable for genome-editing, transcriptional regulation, and other previously impracticable AAV-CRISPR-Cas9 applications. We identify crucial parameters that impact efficacy and clinical translation of our platform, including viral biodistribution, editing efficiencies in various organs, antigenicity, immunological reactions, and physiological outcomes. These results reveal that AAV-CRISPR-Cas9 evokes host responses with distinct cellular and molecular signatures, but unlike alternative delivery methods, does not induce detectable cellular damage in vivo. Our study provides a foundation for developing effective genome therapeutics mRNA-Seq from muscles (9 samples; 3 mice x 3 conditions) and lymph nodes (9 samples; 3 mice x 3 conditions).
2016-07-29 | E-GEOD-84534 | biostudies-arrayexpress