Bisulfite genomic sequencing of DNA from dried blood spot microvolume samples

Abstract

DNA methylation is an important event in epigenetic changes in cells, and a fundamental regulator of gene transcription. Bisulfite genomic sequencing is a powerful technique used in studies of DNA methylation. However, the established procedures often require relatively large amounts of DNA. In everyday practice, samples submitted for analysis might contain very small amounts of poor quality material, as is often the case with forensic stain samples. In this study, we assess a modified, more efficient method of bisulfite genomic sequencing. Genomic DNA extracted from 3-mm dried blood spots using QIAamp micro kit was treated with sodium bisulfite (using EpiTect kit). Subsequent methylation-specific PCR (MSP) followed by DNA sequencing displayed the differentially methylated region of imprinted gene SNRPN. Our results show that this new combination of efficient DNA extraction and bisulfite treatment provides high quality conversion of unmethylated cytosine to uracil for bisulfite genomic sequencing analysis. This reliable method substantially improves the DNA methylation analysis of forensic stain samples.