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Targeted Bisulfite Sequencing

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Targeted bisulfite sequencing

CD genomics is providing novel targeted methylation sequencing by combining proprietary target enrichment techniques and NGS technology to offer a viable solution for methylation state analysis of targeted genomic regions.

DNA methylation is a major type of epigenetic modifications that plays a crucial role in a variety of physiological and pathological processes such as gene regulation, biological development, disease initiation/progression, etc. Target-BS is an accurate, efficient and economic research method of DNA methylation, which is a hybridization-based approach on platforms containing pre-designed oligos that capture the CpG island, gene promoter and other significant methylated regions, followed by methylation sequencing, or PCR-based approach to amplify multiple bisulfite-converted DNA samples in a single reaction. There are commercially designed capture libraries available with a range of epigenetic features that cover ~12% to ~24% of all genome CpGs. In addition, we are available to design the primer sets to capture the region of interest and evaluate site-specific DNA methylation changes.

Compared to whole genome methylation profiling studies, methylation detection of only selected candidate regions is a more viable approach that makes methylation analysis with large sample size more feasible and accessible in different settings. Target-BS dramatically increases data throughput and brings down the cost, it is also able to detect the methylation state of regions that cannot be detected by RRBS or immunoprecipitation approaches. Target-BS has been widely applied for the methylation sequencing and validation of large sample cohort in target region.

Targeted bisulfite sequencing

Workflow

In the process of targeted bisulfite sequencing, the gDNA is bisulfite converted and multiplex amplified using specific designed primers, the amplicon is pooled through barcoding and adapterization. The amplicon library is then sequenced on Illumina sequencing instrument.

Sequencing Strategy and Recommended Sequencing Depth

  • Illumina HiSeq platform, paired-end 150 bp
  • Sequencing depth > 100X

Data Analysis

  1. Raw data statistics
  2. QC results
  3. Alignment against reference genome
  4. Methylation site prediction & CG, CHG, CHH categorization
  5. Estimation of DNA methylation level and distribution trend
  6. 5mC level in different gene structural elements
  7. Identification of differential methylated regions
  8. Function annotation of the proximate genes near differential methylated regions
  9. Multi sample methylation differential analysis

Sample Requirements

  • Sample Type: Genomic DNA without RNA contamination and severe degradation
  • Amount: DNA ≥ 2 µg; concentration ≥ 30 ng/µl
  • Purity: OD260/280 = 1.8~2.0

Key Features and Advantages

  • High precision and resolution. Methylation state detection at single-nucleotide resolution in the target region
  • High efficiency. Effective screening of methylation state through selected genomic regions on NGS platform
  • High-quality probe and primer design. Years of experience in the area of targeted genomic region analysis ensures the enrichment quality.
  • Cost effective. Spend much less on same task compared to Sanger method and whole genome bisulfite sequencing, and ensure more effective utilization of data

CD Genomics provides end-to-end assay and comprehensive service for targeted bisulfite sequencing including bisulfite conversion, primer/probe design synthesis, and validation, hybridization/PCR amplification, library preparation, DNA sequencing and data analysis. CD Genomics use both commercially designed capture arrays and custom-designed capture libraries to fully meet your research objectives. Our experts are ready to offer consultation for your project.

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CONTACT CD GENOMICS

45-1 Ramsey Road, Shirley, NY 11967, USA
Tel: 1-631-275-3058
Fax: 1-631-614-7828
Email: info@cd-genomics.com