m5C lncRNA and CircRNA Sequencing

In m5C modifications, a methyl group is attached to the fifth carbon of the cytosine ring in DNA and RNA molecules. This modification was first identified on DNA and later on RNA in the 1970s. It is an important post-transcriptional modification that has significant roles in many biological processes. Using a high-throughput sequencing method, it was verified that m5C was ubiquitous in non-coding RNA and some mRNA. Currently, the primary detection methods for m5C modifications include RNA bisulfite sequencing (RNA-BisSeq) and m5C meRIP sequencing.

LncRNA is a non-coding RNA with a length of more than 200 nucleotides. Studies have shown that lncRNA plays an important role in many life activities, such as dosage compensation effect, epigenetic regulation, cell cycle regulation and cell differentiation regulation, and has become a hotspot in genetic research.

CircRNA RNA is a special class of non-coding RNA molecules, and it is also the latest research hotspot in RNA field. Recent studies have shown that circRNA molecules are rich in microRNA (miRNA) binding sites and play the role of miRNA sponge in cells, thereby relieving the inhibition of miRNA on its target genes and increasing the expression level of target genes; This mechanism is called competitive endogenous RNA (ceRNA) mechanism. CircRNA plays an important role in disease regulation by interacting with disease-associated miRNAs.

Transcriptome-wide mapping RNA modification is crucial to understand the distribution and function of RNA modifications. CD Genomics adopts the method of rRNA depletion to comprehensively detect the m5C modification on mRNA, lncRNA, and circRNA.

m5C lncRNA and CircRNA SequencingFig1. Dynamically reversible process of 5mC RNA (Song, 2022)

Library Construction of m5C lncRNA&CircRNA Sequencing

Library Construction of m5C lncRNA&CircRNA Sequencing – CD Genomics

Features

Transcriptome-Wide Selectively High Resolution Heterogeneity Analysis
This method can comprehensively detect the m5C modification on mRNA, lncRNA, and circRNA. There are two sequencing strategies, which can be selected according to actual needs. RNA-BisSeq can identify the m5C modified site with single base resolution. can specifically enrich m5C-modified RNA fragments.

Project Workflow

Sample Preparation

1. Sample Preparation

Library Preparation

2. Library Preparation

Sequencing

3. Sequencing

Data Analysis

4. Data Analysis & Delivery

Data analysis

Sample Requirements

RNA sample quantity ≥ 30 µg.

1.8 ≤ OD260/280 ≤ 2.2, 28S:18S ≥ 1.0, RIN ≥ 7.

Please make sure that the RNA is not significantly degraded.

Deliverable: FastQ, BAM, QC report, Distribution of m5C, landscape of DMC, enrichment analysis, PCA analysis, custom bioinformatics analysis.

References:

  1. Guo, Gangqiang, et al. Advances in mRNA 5-methylcytosine modifications: Detection, effectors, biological functions, and clinical relevance. Molecular Therapy-Nucleic Acids 26 (2021): 575-593.
  2. Song, Hang, et al. "Biological roles of RNA m5C modification and its implications in Cancer immunotherapy." Biomarker Research 10.1 (2022): 1-15.
* For Research Use Only. Not for use in diagnostic procedures.


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