Epigenomics

The epigenome comprises the chemical modifications that shape the physical structure of the genome. When epigenomic compounds attach to DNA and modify its function, they are said to have "marked" the genome. These marks do not change the sequence of the DNA. Rather, they change the way cells use the DNA's instructions. The marks are sometimes passed on from cell to cell as cells divide. They also can be passed down from one generation to the next. It regulates gene expression, enabling select genes to be active at different times during the cell life cycle. Epigenetics play a significant role in growth, development, and disease progression. Different factors make the epigenome change like response to diet, stress, and other environmental factors which lead to cause malfunction of protein that read and write epigenomics marks.

Eurofins provide comprehensive in detail wet and dry lab solution epigenome/methylome study which stimulate due to response of various chemical factors either targeted or whole genome sequence. We are offering

Whole genome Bisulfite sequencing (WGBS)

The gold-standard approach in methylation analysis is the whole genome bisulfite sequencing which gives comprehensive base-pair resolution and quantitative information at most genomic cytosine. WGBS is one of the robust global 5-methylcytosine (5mc) detection. This method rely on bisulfite conversion of DNA to detect unmethylated cytosines. This method uses cytosine bisulfite conversion converting unmethylated cytosines to uracil followed by random priming, tagging at the 5’ and 3’ end and introduction of Illumina adapters by PCR amplification. Converted bases are identified (after PCR) as thymine in the sequencing data, and read counts are used to determine the % methylated cytosines.

Advantage

• Discover methylation patterns of all CpG, CHH, and CHG regions across the entire genome.
• Capture full sample diversity with small amounts of DNA
• View methylation at practically every cytosine in the genome across most species
  
  

Methodology

Sample Requirement:

• Submit 2-4 µg of RNA-free genomic DNA (NGS grade) of molecular weight >40kb, Nanodrop A_260/280 ratio >1.8; A_260/230=2.0-2.2; and at a concentration of ~50 ng/µl
• Or 5-10 g of fresh tissue fast frozen in liquid nitrogen and shipped in dry ice to Eurofins facility.

Sample QC:

• RNA free NGS grade gDNA isolation will be carried out using commercially available kit with further purification making it suitable for NGS library preparation.
• Further quality checking will be carried out using agarose electrophoresis and DIN value estimation using Agilent Tape Station. Quantification will be carried out using Qubit 3.0 Fluorometer.

Library Preparation:

• Appropriate amount of gDNA will be bisulfite converted and NGS library will be prepared using TruSeq DNA methylation kit from illumina.
• Library validation will be carried out using Agilent Tape station kit.

Sequencing:

• Prepared library will be sequenced using 2 x 75 bp using NextSeq500 or 2 x 125 bp using HiSeq 2500 using latest sequencing chemistries.
• Delivery of data up to 10X-30X coverage of the genome (depending on size of genome) or customer specified data requirement.

Deliverables :

• Raw data will be available for download as a compressed archive of FASTQ files for each sample.
• Comprehensive compiled report and data set will be shared in link or Pen Drive or Hard Disk.
• For analysis please visit Bioinformatics Services