The methylation of cytosines is a major epigenomic mechanism that modulates the primary genomic code. This leads to the formation of 5-methylcytosines (5mCs) at select sites of the genome. Cytosine methylation regulates gene expression and chromatin remodeling, and as a result plays important roles in many biological functions including embryonic development, cell differentiation, and stem cell pluripotency. Abnormal DNA methylation can lead to diseases, such as cancer.
DNA methylome analysis has for many years been conducted with the use of microarrays, including the widely used Illumina Infinium MethlyationEPIC BeadChips that NUSeq processes. DNA methylation sequencing is a newer technology that is usually based on bisulfite conversion to differentiate methylated vs. unmethylated cytosines. Upon treatment with bisulfite, unmethylated cytosines are converted to uracils, while 5mCs are nonreactive and retained. In the sequencing step, unmethylated cytosines are read as thymines, while methylated cytosines still as cytosines.
Based on genomic coverage, bisulfite conversion based sequencing can be conducted in NUSeq as either Whole-Genome Bisulfite Sequencing (WGBS) or Reduced Representation Bisulfite Sequencing (RRBS). WGBS costs more and the associated data analysis is much more involved. RRBS instead provides a cost effective approach to survey DNA methylation by sampling CpG-rich regions of the genome. To perform RRBS, genomic DNA is digested with a methylation-insensitive restriction enzyme, such as MspI. The digested DNA fragments are then subjected to adapter ligation, bisulfite conversion, and PCR, to generate a library for sequencing.
WGS and RRBS DNA Methylation Sequencing Library Prep Pricing
Library prep costs are listed on the Core Pricing page.