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DNA Hydroxymethylation
DNA hydroxymethylation (5hmC) is an essential epigenetic modification that regulates gene expression. To gain insight into the role of 5hmC, researchers must be clear about the distribution of 5hmC in the genome. However, traditional bisulfite-based methods cannot distinguish between 5hmC and 5mc. The 5hmC monoclonal antibody capture method is a powerful tool for studying DNA hydroxylation modification. Lifeasible, combined with microarray technology and bioinformatic analysis, can obtain a genome-wide hydroxymethylation distribution map, which can help researchers to analyze the molecular mechanisms of species' morphological traits and stressful environmental adaptations from a new perspective.
- Sonication breaks the genome
We sonically interrupt genomic DNA into 400 bp-500 bp DNA fragments.
- Immunoprecipitation of hydroxymethylated DNA
- Linear amplification of hMeDIP and input DNA fragments
The above two DNA fragments (hMeDIP and Input) are amplified. This step significantly increases the sensitivity of the assay, allowing accurate results to be obtained with minute quantities of the test sample.
- Fluorescent labelling
The hMeDIP (Cy5) and Input (Cy3) samples are labeled separately.
- Microarray hybridization
The labeled hMeDIP and input samples are mixed, denatured, and hybridized to the methylation microarray detection chip.
- Image acquisition and data analysis
We use a high-resolution microarray scanner to detect the hybridization signals and professional analysis software to extract, normalize, peak analyze, and report the hybridization results.
- Provide experimental reports, including detailed practical methods and microarray experimental data and graphs
| Scanning image | Cy3, Cy5 fluorescence scanning image. |
| Raw data | Raw data, including fluorescence signal intensity of each probe. |
| Probe report | The log2(hMeDIP/input) value and p-value value of each probe were obtained after correction. |
| The log2(hMeDIP/input) value represents the relative enrichment intensity of each probe in hMeDIP DNA and input DNA. The p-value indicates the probability that the difference between the red and green signals of the probes is caused by abiotic factors. The lower the p-value is, the more likely the probe represents a methylation event, and the p-value is calculated by the modified KS test algorithm. The P-Value is calculated by the modified KS test algorithm. | |
| Peaks report | Peaks represent possible hydroxymethylated regions of DNA, calculated by specialized software; the report includes information on the chromosomal localization of possible peaks and information about the genes and CpG islands surrounding the peaks. |
| Summary report | Provides a comparison of peak regions between multiple samples and a summary for reference. |
- Differential enrichment peaks (DEP)
We use the average of the log2ratio of multiple samples in the replicate groups to analyze the differentially methylated regions between groups, which makes it possible to compare the methylation results and identify differentially methylated regions with the replicate sample data, which is very important for subsequent experiments and analysis.
Lifeasible will ultimately deliver the microarray data to you, along with complete protocols, lab reports (including software analysis results), and lab materials. Please feel free to contact us to submit your request.
The services provided by Lifeasible cover all aspects of plant research, please contact us to find out how we can help you achieve the next research breakthrough.
Contact*If your organization requires the signing of a confidentiality agreement, please contact us by email.
For research or industrial use.
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