Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

DART-seq: an antibody-free method for global m6A detection

ABSTRACT: m6A is a widespread RNA modification which plays important roles in the regulation of gene expression. Methods for the global detection of m6A rely on immunoprecipitation of methylated transcripts using m6A antibodies. However, these methods are costly and require large amounts of input RNA, making them prohibitive for many experiments. Here, we describe DART-seq, an antibody-free method for m6A detection which enables transcriptome-wide mapping of m6A residues using low amounts of input material. DART-seq can be used to obtain global m6A maps using as little as 10 nanograms of total RNA and at single-molecule resolution. This method offers several improvements over current techniques and will facilitate detection of m6A in limiting cell and tissue types.

ORGANISM(S): Homo sapiens

PROVIDER: GSE125780 | GEO | 2019/09/23

REPOSITORIES: GEO

ACCESS DATA

Json Xml

Similar Datasets

Improved methods for deamination-based m6A detection

Project description:We present an improved version of DART-seq which utilizes a variant of the YTH domain engineered to achieve enhanced m6A recognition in APO1-YTH (D422N). In addition, we develop in vitro DART-seq and show that it performs similarly to cellular DART-seq and can map m6A in any sample of interest using nanogram amounts of total RNA.

2022-05-11 | GSE196416 | GEO

DART-seq: an antibody-free method for global m6A detection

Project description:DART-seq: an antibody-free method for global m6A detection

| PRJNA517490 | ENA

RNA m6A methylome from zebrafish zygotes, mouse liver polyA-selected RNAs, mouse embryonic stem cells, spike-in RNA controls, and single mouse oocytes and embryos

Project description:N6-methyladenosine (m6A) is a prevalent RNA modification that has key roles in distinct biological processes, including meiosis and early embryo development. Despite advances in methodology for m6A mapping, the applicability of current methods is limited by the need for large amounts of input material. We develop a sensitive picogram scale m6A RNA immunoprecipitation and sequencing (picoMeRIP-seq) method, also suitable for single-cell (scMeRIP-seq). We validate the down-scaling of the method by profiling m6A from picogram amounts of mouse liver polyA-selected RNA, mouse embryonic stem cells, and zebrafish zygotes. We provide proof-of-principle of m6A profiling in single mouse oocytes and preimplantation embryos. Our work opens a new avenue for studying m6A in single cells and scarce cell types in a transcriptome-wide manner.

2023-05-05 | GSE184893 | GEO

Transcriptome-wide profiling and quantification of N6-methyladenosine by enzyme-assisted adenosine deamination [eTAM-seq]

Project description:We report evolved TadA-assisted N6-methyladenosine sequencing (eTAM-seq), an enzyme-assisted sequencing technology for quantitative, base-resolution profiling of m6A. eTAM-seq functions by global adenosine deamination, enabling detection of m6A as persistent A. We demonstrate adenosine-to-inosine (I) conversion rates up to 99% using a hyperactive TadA variant. With eTAM-seq, we profile and quantify m6A in the whole transcriptomes of HeLa cells and mouse embryonic stem cells (mESCs), with simultaneous deconvolution of the transcriptome and epitranscriptome. Further, we showcase deep sequencing-free, site-specific m6A quantification with as few as 10 cells, an input demand that is at least 4 orders of magnitude lower than existing methods. Collectively, eTAM-seq enables sensitive detection and faithful quantification of m6A with limited RNA input, representing a novel solution to deciphering the epitranscriptome.

2022-10-24 | GSE211303 | GEO

Transcriptome-wide profiling and quantification of N6-methyladenosine by enzyme-assisted adenosine deamination [site_specific_quantification]

2022-10-24 | GSE201063 | GEO

Transcriptome-wide profiling and quantification of N6-methyladenosine by enzyme-assisted adenosine deamination [probes_assay]

2022-10-24 | GSE201062 | GEO

An antibody independent verification method quantifies m6A in the chicken β-actin zipcode

Project description:N6-Methyladenosine (m6A) in mRNA regulates almost every stage in the mRNA life cycle, and the development of the high throughput detection of methylated sites in mRNA using MeRIPSeq or miCLIP revolutionized the m6A research field. Both methods are based on immunoprecipitation of fragmented mRNA. However, it is well documented that antibodies often have nonspecific activities, thus verification of identified m6A sites using an antibody-independent method would be highly desirable. Currently such approaches are limited. Here we present RedBaron, an improved biochemical method for the site-specific detection and quantification of m6A in RNA. We demonstrate that the RedBaron method is able to accurately quantify m6A levels at specific transcripts in vivo. We used this assay for the site-specific detection and quantification of m6A within the chicken β-actin (ACTB) zipcode sequence in chicken embryos and in fibroblast cells. We demonstrate that methylation of this site in the β-actin zipcode enhances ZBP1 binding in vitro, whilst methylation of a nearby adenosine abolishes

2023-05-24 | GSE185078 | GEO

Evaluation of commercially available small RNA methods for plant samples

Project description:Plant small RNAs are a diverse and complex set of molecules, ranging in length from 21 to 24 nt, involved in a wide range of essential biological processes. Nowadays, high-throughput sequencing is the most commonly used method for the discovery and quantification of small RNAs. However, it is known that several biases can occur during the preparation of small RNA libraries, especially using low input RNA. We used two types of plant biological samples to evaluate the performance of seven commercially available methods for small RNA library construction, using different RNA input amounts. We show that when working with plant material, library construction methods have differing capabilities to capture small RNAs, and that different library construction methods provide better results when applied to the detection of microRNAs, phased small RNAs, or tRNA-derived fragments.

2021-08-01 | GSE156973 | GEO

Transcriptome-wide N6-methyladenine profiling in low input multiplex samples by a kit-free multi-barcode method

Project description:N6-methyladnine, which is the most abundant post-transcriptional RNA modification in eukaryotic mRNA, has been proved to be essential in various biological processes and related to numerous diseases. Transcriptome-wide m6A profiling by next generation sequencing is widely used to explore the distributions as well as quantity of m6A modifications. As traditional m6A-seq demands large amount of starting RNA which limited its application to clinical samples, we present a strategy of low input multi-barcode m6A-seq (SLIM-m6A-seq) to realize simplified m6A profiling of mixed clinical samples. This method maintains the advantages of preferable detection limit, low cost and simplified experiment procedures. Sequencing data of clinical blood samples from patients with diabetic myocardial infarction suggest that this method is practical in clinical application and m6A may play a critical role in the progression of diabetic cardiovascular complications.

2021-10-14 | GSE185749 | GEO

Improved methods for deamination-based m6A detection

Project description:Improved methods for deamination-based m6A detection

| PRJNA804865 | ENA