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Whole Genome Bisulfite Sequencing

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whole genome bisulfite sequencing
By Christoph Bock (Max Planck Institute for Informatics) (Own work) [CC BY-SA 3.0
(http://creativecommons.org/licenses/by-sa/3.0)], via Wikimedia Commons
DNA methylation at the C5 position of cytosine plays a crucial role in gene expression and chromatin remodelling, and perturbations in methylation patterns are implicated in the development of cancer, neurodegenerative diseases, and neurological disorders. Therefore, the mapping of methylated bases (the methylome) is critical to understanding gene expression and other processes subject to epigenetic regulation.

Novogene provides whole genome sequencing of bisulfite-converted DNA, as an effective method to identify individually methylated cytosines on a genome-wide scale. Methylome analysis is an increasingly valuable research tool with a range of applications, including studies on gene regulation, stem cell differentiation, embryogenesis, aging, cancer and other diseases, and phenotypic diversity and evolution in plants and animals.

The Novogene Advantage

  • Extensive experience: over 500 projects successfully completed and a bioinformatics analysis team composed entirely of Ph.D. scientists.
  • Unsurpassed data quality: We guarantee that ≥ 80% of bases have a sequencing quality score ≥ Q30, exceeding Illumina’s official guarantee of ≥ 75%.
  • Comprehensive data analysis: We use widely-accepted mainstream software such as Bismark and a mature in-house pipeline for mapping, DMR analysis, functional analysis and data visualization.

Project Workflow

whole genome bisulfite sequencing project workflow

Sequencing Strategy

  • 250~300-bp insert bisulfite treated DNA library
  • HiSeq platform, paired-end 150 bp

Data Quality Guarantee

  • Over 80% of bases have a sequencing quality score ≥ Q30. See examples of our high quality data.

Sample Requirements

  • DNA amount:
    • For genome size ≤ 1.5 G: ≥ 3.0 μg
    • For 1.5G < genome size ≤ 3.5 G: ≥ 5.0 μg
  • DNA concentration: ≥ 50 ng/μl
  • DNA volume: ≥ 20 μl
  • Purity: No degradation, no DNA contamination

Turnaround Time

  • 15 working days from verification of sample quality without data analysis
  • The data analysis turnaround is project-dependent.

Recommended Sequencing Depth

  • Sequencing depth > 30X

Analysis Pipeline

whole genome biosulfite sequencing analysis pipeline


Table. Representative samples showing data quality of Novogene’s whole genome bisulfite sequencing service.
Sample NameRaw ReadsRaw BasesClean ReadsClean BasesError Rate (%)Q20 (%)Q30 (%)GC (%)bisulfite Conversion Rate (%)
Sample 1750864918112.63 G686418034102.96 G0.0394.6286.5223.0199.29
Sample 2762188782114.14 G61849526092.61 G0.0494.3586.4121.0899.66
Sample 3789026912118.35 G721080112108.16 G0.0296.5490.6321.1299.66
Sample 4784535120117.68 G760489944114.07 G0.0392.8185.4920.7099.63

Project Example

The following study utilized Novogene's WGBS and RNA-Seq services.

Lineage-specific functions of TET1 in the postimplantation mouse embryo
Nature Genetics, 49 (7): 1061-1072 (2017)

The TET enzymes are essential epigenetic regulators and also play a significant role in embryo implantation. Novogene’s whole genome bisulfite sequencing (WGBS) service, together with RNA sequencing, was adopted to explore the non-redundant functions of one TET enzyme, TET1, in the post-implantation mouse embryo. TET1 enzyme was shown to regulate lineage-specific genes in the pre-streak mouse epiblast and suppress metabolic genes in the extraembryonic ectoderm. TET1 was also found to be involved in the DNA methylome regulation of epiblast-like cells. While loss of TET1 affected the EpiLC methylome, the gene expression of more than 90% of associated genes was not affected. Further studies demonstrated the catalytic activity of telomere maintenance, via the ZSCAN4 locus, and the catalytic-independent repression of target genes. The interplay between catalytic and non-catalytic activities in TET1 was regulated through the transcriptional repressor, JMJD8. This study highlights how the complex interplay and functions of TET1 in normal embryo development were identified using WGBS as a tool.

wgbs-data
Figure. Heat map cluster of the differentially expressed genes that are regulated by TET1 in the epiblast

Publications Using Novogene’s Expertise

JournalTitle
Nature Communications, 3:850 (2012)An atlas of DNA methylomes in porcine adipose and muscle tissues.
Genome Biology, 15: R49 (2014)Whole-genome analysis of 5-hydroxymethylcytosine and 5-methylcytosine at base resolution in the human brain.
Bioinformatics, 33(11):1591-1595 (2017)Genome-wide analysis of DNA methylation profiles in a senescence-accelerated mouse prone 8 brain using whole-genome bisulfite sequencing.
Nature Genetics, 49(7):1061-1072 (2017)Lineage-specific functions of TET1 in the postimplantation mouse embryo.
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