Changes in DNA methylation associated with type of delivery

A type of delivery represents an epigenetic factor with potential to affect further development by multiple mechanisms. One of them can be DNA methylation, which represents major epigenetic mechanism inolving direct chemical modification to the DNA. This study aims to examine wheather type of delivery induces changes of DNA methylation. The blood samples from umbilical cord blood and peripheral blood of 12 infants, born by vaginal delivery and cesarean section, were used to preparation of the Methylseq library according to NEBNext enzymatic Methyl-seq Methylation Library Preparation Kit with further generation of target-enriched DNA libraries by Twist Human Methylome Panel. DNA methylation status was measured using Illumina next-generation sequencing (NGS) systems NextSeq 500 and NextSeq 2000 (Illumina) using 2x75 or 2x100 paired-end sequencing kits and protocols. The entire analysis was done in the BaseSpace Sequence Hub (BSSH) cloud service. In the first step, the BCL files were converted into FASTQ files. Then FASTQ files were submitted as the input to the DRAGEN Methylation pipeline. A conversion of the bases from C-to-T, and G-to-A was done. The bisulfite converted reads were aligned to the reference genome (hg19 alt-aware UCSC reference genome). Targeted methylation calling was performed once the converted reads were aligned to the reference. The regions subdued to the analysis were provided in the BED file from Twist Bioscience. In the background, BSSH utilizes the Bismark Bisulfite Read Mapper to map the reads and to identify the local cytosine methylation status. As a result, we have obtained the BAM files as well as the Cytosine Report TXT file which serves as an input to the MethylKit application. The MethylKit application inside the BSSH represents the second phase of the computational analysis. Here the input data, produced as an output in the previous phase, had to be provided. At the same time, the conceptual division between the samples considered to be the experiment and the samples used as a reference set had to be stated.