Similar Datasets
Project description:The goal of these experiments were to test the on-target and target-adjacent editing efficiencies of different single-nucleobase editing systems. Previous studies have shown that tethering DNA mutating enzymes to Cas9-nickase-UGI complexes results in editing of chromosomal DNA. However, these editing events encompass undesirable target-adjacent nucleobase edits. Here, we characterize a novel approach that reduces the frequency of target-adjacent editing while maintaining a high level of on-target editing.
2020-02-23 | E-MTAB-8742 | biostudies-arrayexpress
Project description:A Scalable Epitope Tagging Approach for High Throughput ChIP-seq Analysis ChIP-seq comparison between CRISPR editing cells using epitope antibody and non-editing cells using endogeneous TF antibody
2016-09-01 | E-GEOD-78064 | biostudies-arrayexpress
Project description:Enrichment of histone marks was assessed by CUT&RUN-sequencing after epigenetic editing with dCas9::KRAB (or dCas9::GFP as a control) and subsequently upon release of the trigger. Samples were collected after 7 days of DOX induction (epigenetic editing) and after 4 and 7 days of DOX washout (release of the trigger).
2021-11-23 | E-MTAB-10525 | biostudies-arrayexpress
Project description:HEK293T/17 cells were genetically edited by SMART editing to assess the effectiveness of SMART editing. There are 4 target genes: CXCR4, Lmnb1, Nrl and NRXN3. Cultured cells were first synchronized at the G2/M phase. Then, cells were transfected with Cas9 RNP along with SMART or traditional templates. After culture for two to three days, cells were harvested, and genomic DNA was extracted. The targeted loci were amplified by PCR and sequenced on an Illumina MiSeq.
2025-04-21 | E-MTAB-14909 | biostudies-arrayexpress
Project description:Background: Adenosine deaminases that act on RNA (ADARs) bind to double-stranded and structured RNAs and deaminate adenosines to inosines. This A to I editing is widespread and required for normal life and development. Besides mRNAs and repetitive elements, ADARs can target miRNA precursors. Editing of miRNA precursors can affect processing efficiency and alter target specificity. Interestingly, ADARs can also influence miRNA abundance independent of RNA-editing. In mouse embryos where editing levels are low, ADAR2 was found to be the major ADAR protein that affects miRNA abundance. Here we extend our analysis to adult mouse brains where high editing levels are observed. Results: Using Illumina deep sequencing we compare the abundances of mature miRNAs and editing events within them, between wild-type and ADAR2 knockout mice in the adult mouse brain. Reproducible changes in abundance of specific miRNAs are observed in ADAR2 deficient mice. Most of these quantitative changes seem unrelated to A to I editing events. However, many A to G transitions in cDNAs prepared from mature miRNA sequences, reflecting A to I editing events in the RNA, are observed with frequencies reaching up to 80%. About half of these editing events are primarily caused by ADAR2 while a few miRNAs show increased editing in the absence of ADAR2, suggesting preferential editing by ADAR1. Moreover, novel, previously unknown editing events were identified in several miRNAs. In general 64% of all editing events are located within the seed region of mature miRNAs. In one of these cases retargeting of the edited miRNA could be verified in reporter assays. Also, altered processing efficiency upon editing near a processing site could be experimentally verified. Conclusions: ADAR2 can significantly influence the abundance of certain miRNAs in the brain. Only in a few cases changes in miRNA abundance can be explained by miRNA editing. Thus, ADAR2 binding to miRNA precursors, without editing them, may influence their processing and thereby abundance. ADAR1 and ADAR2 have both overlapping and distinct specificities for editing of miRNA editing sites. Over 60% of editing occurs in the seed region possibly changing target specificities for many edited miRNAs. Examination of the effect of ADAR2 on mature miRNA abundance and sequence in adult mouse brain.
2014-10-02 | E-GEOD-61066 | biostudies-arrayexpress
Project description:The genome-wide identification, tissue-specificity and functional implications of Apobec-1 mediated C-to-U RNA editing remains incomplete. Deep sequencing, data filtering and validation from wild-type and Apobec-1 deficient mice revealed 56 novel editing sites in 54 intestinal mRNAs and 22 novel sites in 17 liver mRNAs (74-81% true-positive), all within 3' untranslated regions. Eleven of 17 liver RNAs shared editing sites with intestinal RNAs, while 6 sites were unique to liver. Changes in RNA editing led to corresponding changes in intestinal mRNA and protein levels in 11 genes. We found distinctive polysome profiles for several editing targets and demonstrated nuclear but not cytoplasmic editing of novel exonic sites in intestinal (but not hepatic) apoB RNA. RNA editing was validated using cell-free extracts from wild-type but not Apobec-1 deficient mice. These studies define selective, tissue-specific targets of Apobec-1 dependent RNA editing and show the functional consequences of editing are both transcript- and tissue-specific. Examination of C-to-U RNA editing in mouse liver and intestine
2014-05-23 | E-GEOD-57910 | biostudies-arrayexpress
Project description:Background: Adenosine deaminases that act on RNA (ADARs) bind to double-stranded and structured RNAs and deaminate adenosines to inosines. This A to I editing is widespread and required for normal life and development. Besides mRNAs and repetitive elements, ADARs can target miRNA precursors. Editing of miRNA precursors can affect processing efficiency and alter target specificity. Interestingly, ADARs can also influence miRNA abundance independent of RNA-editing. In mouse embryos where editing levels are low, ADAR2 was found to be the major ADAR protein that affects miRNA abundance. Here we extend our analysis to adult mouse brains where high editing levels are observed. Results: Using Illumina deep sequencing we compare the abundances of mature miRNAs and editing events within them, between wild-type and ADAR2 knockout mice in the adult mouse brain. Reproducible changes in abundance of specific miRNAs are observed in ADAR2 deficient mice. Most of these quantitative changes seem unrelated to A to I editing events. However, many A to G transitions in cDNAs prepared from mature miRNA sequences, reflecting A to I editing events in the RNA, are observed with frequencies reaching up to 80%. About half of these editing events are primarily caused by ADAR2 while a few miRNAs show increased editing in the absence of ADAR2, suggesting preferential editing by ADAR1. Moreover, novel, previously unknown editing events were identified in several miRNAs. In general 64% of all editing events are located within the seed region of mature miRNAs. In one of these cases retargeting of the edited miRNA could be verified in reporter assays. Also, altered processing efficiency upon editing near a processing site could be experimentally verified. Conclusions: ADAR2 can significantly influence the abundance of certain miRNAs in the brain. Only in a few cases changes in miRNA abundance can be explained by miRNA editing. Thus, ADAR2 binding to miRNA precursors, without editing them, may influence their processing and thereby abundance. ADAR1 and ADAR2 have both overlapping and distinct specificities for editing of miRNA editing sites. Over 60% of editing occurs in the seed region possibly changing target specificities for many edited miRNAs. Examination of the effect of ADAR2 on gene expression in mature mouse wildtype and ADAR2 knockout brain using AffymetrixM-BM-. GeneChipM-BM-. Whole Transcript (WT) Expression Arrays (Analysis by KFB Regensburg, Germany)
2014-10-02 | E-GEOD-61067 | biostudies-arrayexpress
Project description:RNA-editing is a tightly regulated, and essential cellular process for a properly functioning brain. Dysfunction of A-to-I RNA editing can have catastrophic effects, particularly in the central nervous system. Thus, understanding how the process of RNA-editing is regulated has important implications for human health. However, at present, very little is known about the regulation of editing across tissues, and individuals. Here we present an analysis of RNA-editing patterns from 9 different tissues harvested from a single mouse. For comparison, we also analyzed data for 5 of these tissues harvested from 15 additional animals. We find that tissue specificity of editing largely reflects differential expression of substrate transcripts across tissues. We identified a surprising enrichment of editing in intronic regions of brain transcripts, that could account for previously reported higher levels of editing in brain. There exists a small but remarkable amount of editing which is tissue-specific, despite comparable expression levels of the edit site across multiple tissues. Expression levels of editing enzymes and their isoforms can explain some, but not all of this variation. Together, these data suggest a complex regulation of the RNA-editing process beyond transcript expression levels. Tissues include: brain, heart, kidney, liver, lung, skin, spleen, testis, thymus mRNA profiles of 9 tissues from an 8 week old, male C57BL/6 mouse were generated by deep sequencing using Illumina HiSeq.
2016-01-19 | E-GEOD-74747 | biostudies-arrayexpress
Project description:A genome-wide CRISPR screen was combined with a tdTomato reporter-based epigenetic memory assay to identify factors that erase epigenetic memory in ESC. After introducing genome wide perturbation and dCas9::KRAB-mediated epigenetic editing of the Esg1-tdTomato reporter, the trigger was released and cells that maintained the silencing sorted at FACS. Samples were collected out of sorted tdTomato negative (TOMminus) and positive (TOMplus) cells after 6 days of DOX treatment (epigenetic editing) and 3 or 7 days of DOX washout (release of the trigger), using a gating strategy to separate the bottom 2.5% negative cells (2.5%gate) and cells ranging from mildly to fully repressed (widegate).
2021-11-23 | E-MTAB-10524 | biostudies-arrayexpress
Project description:We used transgenic mouse embryos that are deficient in the two enzymatically active RNA editing enzymes ADAR1 and ADAR2 to compare relative frequencies but also sequence composition of mature miRNAs in these genetically modified backgrounds to wild-type mice by Illumina next gen sequencing. Deficiency of ADAR2 leads to a reproducible change in abundance of specific miRNAs and their predicted targets. Changes in miRNA abundance seem unrelated to editing events. Additional deletion of ADAR1 has surprisingly little impact on the mature miRNA repertoire, indicating that miRNA expression is primarily dependent on ADAR2. A to G transitions reflecting A to I editing events can be detected at few sites and at low frequency during the early embryonic stage investigated. Again, most editing events are ADAR2 dependent with only few editing sites being specifically edited by ADAR1. Besides known editing events in miRNAs a few novel, previously unknown editing events were identified. Some editing events are located to the seed region of miRNAs opening the possibility that editing leads to their retargeting. GSM852140-8: sequencing of mature miRNAs of wt, ADAR2-/- and ADAR1-/-/ADAR2-/- female mouse embryos at E11.5 GSM863778-81: Gene expression was measured in wiltype, ADAR2-/- and ADAR1-/-/ADAR2-/- E11.5 whole female mouse embryos using Agilent Whole Mouse Genome Oligo Microarrays 8x60K.
2012-02-02 | E-GEOD-34626 | biostudies-arrayexpress