m6A RNA Enrichment & Quantification Kit

Product Description

N6-methyladensoine (m⁶A), the methylation of adenosine in the N6 position, is a prevalent and reversible post-transcriptional RNA modification that decorates mRNA, tRNA and rRNA. Recent studies have revealed the crucial roles of m⁶A RNA modification in multiple cellular processes, including RNA structural stability, folding, interactions with proteins, cell viability, impaired self-renewal ability, cell proliferation, and cell death. Methylated RNA immunoprecipitation (RIP) is a gold standard method to monitor the status and map the location of RNA modifications in specific genes and in transcriptome wide.

• Fast, streamlined procedure, finished in 5 hours
• Hands-on time is less than 1 hour
• Enriched RNA for transcriptome-wide profiling

The RayBio® m⁶A (N6-methyladenosine) RNA Enrichment and quantification kit (QMERIP-M6A) identifies the abundance and enrichment location of m⁶A by MeRIP and quantitative real-time PCR. The enriched m⁶A RNA could also be used for transcriptome-wide profiling of m⁶A. In this assay, RNA sample is digested into fragments consisting of 100~500 nucleotides followed by incubation with an antibody against m⁶A conjugated to a magnetic bead. Then, the enriched RNA is released, purified, and eluted for qRT-PCR analysis. Included in the kit is a non-immune IgG control conjugated to a magnetic bead and two pairs of primer controls for Hela cell RNA. QMERIP-M6A uses a fast and streamlined procedure so the assay can be finished in 5 hours with a hands-on time of less than 1 hour. This kit has been validated for human, mouse, and rat m⁶A enrichment.

Workflow

Supporting Data

Figure 2. QMERIP-M6A enrichment test by PCR and RT-qPCR. (A) Representative images of agarose gel running after PCR amplification for the enrichment of m⁶A targeted SMAD7 and ACTB mRNA. (B) Bar graph showing the percentage enrichment of m⁶A targeted SMAD7 and ACTB mRNA using RT-qPCR assays. 2% Input RNA sample before IP was used as the positive control and IgG-IP was used as the negative control. Almost no PCR products were amplified from the m⁶A-IP ACTB RNA samples. Error bars, SD (n = 3 independent experiments). *** P-value <0.001 compared to IgG-IP was calculated using the student’s t-test.