RRBS (Reduced Representation Bisulfite Sequencing)

single base pair resolution DNA methylation analysis

 

RRBS Service の概要

Reduced Representation Bisulfite Sequencing (RRBS)
 

Reduced Representation Bisulfite Sequencing(RRBS)は、最大500万個のCpG領域において1塩基対の分解能でDNAメチル化データを取得でき、サンプル間でのメチル化の差も容易に同定することが可能です。 Whole Genome Bisulfite Sequencingよりも費用効果が高く、ビーズアレイベースのDNAメチル化プラットフォームよりも広い範囲のデータを取得できるため、RRBSはサンプル間でDNAメチル化をプロファイリングする理想的なプラットフォームです。

RRBS is a method designed to isolate DNA containing CpGs by using DNA restriction endonuclease digestion and amplification of fragments smaller than 300 bp. In Brief, the methylation insensitive endonucleases, Msp1 and Taq1, are used to cut C^CGG and T^CGG respectively, thus generating DNA fragments with CpG at the end. Since CpG islands and promoters have a high density of CpGs they are cut at a high frequency and the fragments derived from these regions tend to be short. This subset of short fragments is preferentially amplified when the library is generated through PCR and additional selection occurs during sequencing cluster generation. The final fraction of DNA that is sequenced represents a small fraction of the total genome and is enriched in CpGs. These fragments are mapped to the genome to reveal the methylation status of millions of individual CpGs.

RRBS の特長:

  • サンプルあたり、最大500万CpGのメチル化データを取得
  • プロモーターおよびCpGアイランドを含む生物学的に関連性のある標的領域の情報
  • シークエンスの深さは、シングルエンド30,000,000 リード以上
  • DNAの必要量はわずか 100 ng
  • ランダムなバーコードの組み込みによりPCR重複を除去し、ライブラリの偏りを軽減

Deliverables:

  • Raw data (FASTQ) output files
  • Aligned data (BAM) files
  • Methylation table includes percent methylation value for each CpG
  • Differential methylation analysis
  • Figures & graphs
    – Includes coverage depth analysis, correlation plots, and pie charts

RRBSについての詳細はこちら

RRBS Overview flow chart

What our customers are saying about us

"We worked with Active Motif to process some FFPE and frozen liver samples sourced from 25+year old samples for an RRBS assay. The overall process of submitting samples, communicating with Active Motif, receiving the data, and obtaining the summary was quick and painless. The whole operation was smooth and professionally done."

Brian Chorley, PhD
Environmental Protection Agency
View complete list of testimonials >

 

RRBS Service受託解析サービスのデータ

RRBS data from human samples

Figure 1: RRBS data from human samples.

The displayed regions are representative regions from the genome-wide data set and shows differential DNA methylation at 1A) an exon of CD19 and 1B) an intron of PAX5. Each block is a separate data point with red representing a methylated cytosine and blue representing an unmethylated base.


RRBS data illustrating sequencing depth coverage for any given CpG

Figure 2: Graph illustrating sequencing depth coverage for any given CpG.

The minimum sequence depth for a CpG to be included in the analysis is 10. Over 1 million CpG sites have a coverage of at least 10 reads.


RRBS data illustrating the correlation between each sample analyzed

Figure 3: Graph illustrating the correlation between each sample analyzed.

The upper right corner shows the Pearson correlation coefficient for the two samples. The bar plots show the distribution of the percent methylation in each sample, with the majority of analyzed CpG sites having 0 or 100% methylation. The scatter plot in the lower left corner shows the level of methylation at shared CpG sites for each sample.


RRBS data showing the assignment of differentially methylated cytosines to genic features

Figure 4: Pie chart showing the assignment of differentially methylated cytosines to genic features.

A differential methylation analysis was performed on a per CpG basis, and identified sites were then aligned to annotated gene features such as promoters and exons.

 

RRBS Service 使用文献

Search our database of customer publications that have used our RRBS services.


 

 

 

RRBS Service 受託解析サービスの資料

 

 

 

RRBS Service Sample Submissionポータル

オンラインSample Submissionポータルは、解析したいサンプルの情報をアクティブ・モティフの担当者と共有し、依頼者には解析の進捗をご覧いただけるサイトです。 解析を依頼するために必要な連絡先、サンプルの情報、また解析の内容を記載していただきます。記載の方法や、サンプル提出方法について、ご不明な点がありましたら「[email protected]」までご連絡ください。

 

You might also be interested in:

 
Name Cat No. Price  
RRBS 25069 Get Quote