Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Universal Enrichment and Sequencing of DNA Modifications Using a Click-Coupled IgG-Protein G System

ABSTRACT: Efficient nucleic acid enrichment is pivotal for deciphering epigenetic modifications and disease biomarkers, yet current methods are constrained by insufficient specificity, poor versatility, and high costs. We developed a universal strategy named ‘Click-IP-Seq’ by leveraging the high-affinity binding between Protein G and Fc region of DBCO-modified IgG. This enabled the directional conjugation of DBCO-IgG with azide-modified nucleic acids via copper-free strain-promoted azide-alkyne cycloaddition (SPAAC) click chemistry, achieving specific capture and enrichment of modified nucleic acids. Firstly, this method efficiently enriched two major DNA modifications, 8-oxo-7,8-dihydroguanine (8-oxo-dG) and 5-hydroxymethylcytosine (5hmC) in model DNA systems. Then, genome-wide distribution of 8-oxodG from human cells and tissues align with previously reports. Finally, employing ‘Click-IP-Seq’, we performed the first comprehensive analysis of 8-oxo-dG spatial distribution and associated biological functions in human colorectal carcinoma tissues. This technology provides a high-specificity and versatile enrichment platform for nucleic acid modifications, which is expected to promote the application of cancer molecular diagnosis.

ORGANISM(S): Homo sapiens mixed organisms

PROVIDER: GSE305517 | GEO | 2025/09/05

REPOSITORIES: GEO

ACCESS DATA

Json Xml

Dataset's files

Source:

			Action	DRS
		Other

Items per page:

1 - 1 of 1

Similar Datasets

Epigenomic profiling of active regulatory elements by enrichment of unmodified CpG dinucleotides

Project description:In this study we present an enrichment-based and bisulfite-free approach for epigenomic profiling named ‘Active-Seq’ (Azide Click Tagging for In Vitro Epigenomic sequencing) that achieves genome-wide profiling of DNA, by enriching for non-modified CpG sites using a mutated methyltransferase enzyme. Active-Seq is a fast epigenetic profiling platform with a simple and streamlined workflow, performed in tandem with sequencing library preparation. The enzymatic chemistry is non-damaging toward the DNA which is critical for working with low concentration DNA input and will facilitate the future development of multiomics assays.

2025-05-04 | GSE266366 | GEO

DNA-seq of mitochondrial DNA from liver of OGG1 and MUTYH deficient mice that have been bred for five consecutive generations

Project description:Studying whether removal of base-excision repair from mitochondria will result into increase in mitochondrial DNA (mtDNA) mutation load. The endogenous genes of OGG1 and MUTYH DNA glycosylases were modified to lack the genomic region encoding for the predicted mitochondrial targeting sequence. The mouse lines used: A mouse line that lacks the region encoding for the mitochondrial targeting sequence (L2 to W23) of OGG1 (Ogg1 dMTS mice). A mouse line that lacks the region encoding the mitochondrial targeting sequence (K2 to P33) of MUTYH (Mutyh dMTS mice). To accumulate mutations to the mitochondrial DNA these mice were bred double homozygous Mutyh dMTS x Ogg1 dMTS mice as a maternal lineage for five consecutive generations and mitochondrial DNA from liver was extracted from the offspring and sequenced with Illumina. OGG1 and MUTYH are involved in repair of 8-oxo-dG from DNA. 8-oxo-dG can be a mutagenic lesion because some DNA repair polymerases are known to erroneously incorporate adenosine opposite to 8-oxo-dG during replication leading to GC>TA transversion mutations.

2018-07-13 | E-MTAB-6532 | biostudies-arrayexpress

RNA-seq of mouse heart that is both heart SOD2 knockout and deficient for mitochondrial DNA repair

Project description:To study whether increase in mitochondrial oxidative stress (SOD2 removal) and decrease in mitochondrial DNA repair (Ogg1 dMTS) results into increase in mutations in mitochondrial RNA (mtRNA). Oxidative stress has been suggested to induce mutations in mtDNA and as DNA is used as a template in transcription, mutations or 8-oxo-dG on DNA can induce GC>TA transversion accumulation in mtRNA. To verify this, we extracted and sequenced (Illumina) total RNA from heart Sod2 knockout mice alone and mice that were also deficient for mitochondrial base-excision repair. The repair deficiency was induced by removing the genomic region encoding for the predicted mitochondrial targeting sequence from endogenous OGG1 (L2 to W23) called Ogg1 dMTS mice, thus excluding the protein from mitochondria. OGG1 is a DNA glycosylase that recognizes and repairs 8-oxo-dG damage from DNA. Oxidative stress can induce 8-oxo-dG lesions, thus we removed the mitochondrial matrix localized superoxide dismutase (SOD2) from these mice to increase the level of oxidative stress. 8-oxo-dG lesion can be mutagenic because some DNA repair polymerases are known to erroneously incorporate adenosine opposite to 8-oxo-dG during replication leading to GC>TA transversion mutations.

2018-07-13 | E-MTAB-6534 | biostudies-arrayexpress

RNA-Seq of A375 cells treated with tetracycline-based crosslinking probes, with and without competition by parent tetracyclines, to show specific binding of the tetracycline analogs Col-3 and doxycycline on rRNA substructures of the human ribosome

Project description:Upon finding that the tetracyclines COL-3 and doxycycline target unique rRNA substructures of the 80S ribosome (E-MTAB-7147), we confirmed these putative ribosomal binding sites by showing that tetracycline-based crosslinking probes are specifically competed from these rRNA structures by their parent drugs. In short, we incorporated dual bioorthogonal handles into tetracycline-based probes, containing both a photoactive diazirine to enable direct probe crosslinking to the human ribosome and an azide handle to allow selective enrichment of crosslinked biomolecules via copper-free ‘click’ chemistry. The COL-3 and doxycycline probes were each incubated with A375 cells, followed by irradiation at 365 nm to induce photolysis of the diazirine moiety and subsequent crosslinking to adjacent ribosomal components. Pulldown and RNA-Seq of the crosslinked RNAs from our experiments were used to identify enrichment of reverse transcription (RT) stops at ribosomal RNA sites caused by local crosslinking of our probes. In these experiments, UV crosslinking was preformed with the COL-3 and doxycycline probe compounds in the presence and absence of free COL-3 and doxycycline (i.e., parent tetracycline molecules lacking the bifunctional diazirine-azide moiety), which were used as competitors to specifically diminish crosslinking to the ribosomal RNA. Crosslinking to ribosomal RNA was determined using RNA-Seq based RT stop enrichment, which was was also compared to inactive tetracycline probes containing a modified bifunctional linker, a non-specific aniline probe, and untreated controls.

2018-08-31 | E-MTAB-7161 | biostudies-arrayexpress

DNA-seq of mitochondrial DNA from mouse heart that is both heart SOD2 knockout and deficient for mitochondrial DNA repair

Project description:To study whether increase in mitochondrial oxidative stress (SOD2 removal) and decrease in mitochondrial DNA repair (Ogg1 dMTS) results into increase in mitochondrial DNA mutation load. Oxidative stress has been suggested to induce mutations in mtDNA. To verify this, we extracted and sequenced (Illumina) mitochondrial DNA from heart Sod2 knockout animals that were also deficient for mitochondrial base-excision repair. The repair deficiency was induced by removing the genomic region encoding for the predicted mitochondrial targeting sequence from endogenous OGG1 (L2 to W23) called Ogg1 dMTS mice, thus excluding the protein from mitochondria. OGG1 is a DNA glycosylase that recognizes and repairs 8-oxo-dG damage from DNA. Oxidative stress can induce 8-oxo-dG lesions, thus we removed the mitochondrial matrix localized superoxide dismutase (SOD2) from these mice to increase the level of oxidative stress. 8-oxo-dG lesion can be mutagenic because some DNA repair polymerases are known to erroneously incorporate adenosine opposite to 8-oxo-dG during replication leading to GC>TA transversion mutations.

2018-07-13 | E-MTAB-6533 | biostudies-arrayexpress

Universal Enrichment and Sequencing of DNA Modifications Using a Click-Coupled IgG-Protein G System

Project description:Universal Enrichment and Sequencing of DNA Modifications Using a Click-Coupled IgG-Protein G System

| PRJNA1306169 | ENA

RNA-Seq of A375 cells treated with tetracycline-based crosslinking probes to identify binding sites for the tetracycline analogs Col-3 and doxycycline on the human ribosome

Project description:Following the identification of the 80S ribosome as a putative target of the tetracycline analogs Col3 and doxycycline, we next sought to identify the precise binding sites for these tetracyclines within the ribonucleoprotein complex. We incorporated dual bioorthogonal handles into tetracycline-based probes, containing both a photoactive diazirine to enable direct probe crosslinking to the human ribosome and an azide handle to allow selective enrichment of crosslinked biomolecules via copper-free click chemistry. The Col-3 and doxycycline probes were each incubated with A375 cells, followed by irradiation at 365 nm to induce photolysis of the diazirine moiety and subsequent crosslinking to adjacent ribosomal components. Pulldown and RNA-Seq of the crosslinked RNAs from our experiments were used to identify enrichment of reverse transcription (RT) stops at ribosomal RNA sites caused by local crosslinking of our probes. This RNA-Seq based RT stop enrichment analysis was compared to results using an non-specific aniline control probe and untreated controls.

2018-08-30 | E-MTAB-7147 | biostudies-arrayexpress

DYn-2 based identification of Arabidopsis sulfenomes

Project description:Identifying the sulfenylation state (-SOH) of stressed cells. Here, we optimized an in vivo trapping method of sulfenic acids in hydrogen peroxide (H2O2) stressed plant cells. With click chemistry, the dimedone based alkyne functionalized DYn-2 probe was biotinylated for subsequent streptavidin enrichment of sulfenylated proteins. the DYn-2 modification and DYn-2-Biotin-azide modification were unknown modifications, we therefore chose N-D-glucuronylglycine [MOD00067] and N-palmitoyl-S-(sn-1-2,3-dipalmitoyl-glycerol)cysteine [MOD00444]

2016-06-27 | PXD001562 | Pride

Impact of azide metabolic glycoengineering and bioorthogonal click chemistry cell-cell tethering in adipose tissue-derived mesenchymal stromal cells

Project description:We report RNA sequencing data from adipose tissue-derived mesenchymal stromal cells (hASCs) functionalized via metabolic glycoengineering with pendant azide sialosides. Beyond the impact of this cell surface engineering on transcriptome, we explore the expression profiles of azide-hASCs covalently tethered to an interfacial glycosaminoglycan via bioorthogonal click chemistry and yielding a 3D multicellular construct. We find global gene expression changes in response to metabolic glycoengineering in 2D cell culture conditions as well as those arising from cell-to-cell tethering into 3D environments.

2024-04-15 | GSE231507 | GEO

Selective chemical tracking of Dnmt1 catalytic activity in live cells

Project description:Enzymatic methylation of cytosine to 5-methylcytosine in DNA is a fundamental epigenetic mechanism involved in mammalian development and disease. DNA methylation is brought about by collective action of three AdoMet-dependent DNA methyltransferases (DNMTs), whose catalytic interactions and temporal interplay are poorly understood. We used structure-guided engineering of the Dnmt1 methyltransferase to enable catalytic transfer of azide tags onto DNA from a synthetic cofactor analog, Ado-6-azide, in vitro . We then CRISPR-edited the Dnmt1 locus in mouse embryonic stem cells to install the engineered codon, which, following pulse- internalization of the Ado-6-azide cofactor by electroporation, permitted selective azide-tagging of Dnmt1-specific genomic targets in cellula . The deposited covalent tags were exploited as ‘click’ handles for reading adjoining sequences and precise genomic mapping of the methylation sites. The proposed approach, Dnmt-TOP-seq, enables high-resolution temporal tracking of the Dnmt1 catalysis in mammalian cells paving the way to selective studies of other methylation pathways in eukaryotic systems.

2022-02-04 | GSE182445 | GEO

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