Quantification and Characterisation of the DNA phosphorothioate modification in the gut microbiome

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Nucleic acids, like DNA and RNA, are the basis of all life. In addition to the known four letter code of DNA, there are several more natural modifications of the canonical nucleosides. Only recently, the first modification of the DNA backbone was discovered, the phosphorothioate modification. This modification is only formed in a few bacteria. Over the last years, research on the connection of human health and the natural bacteria that inhabit the human body has been in an increased focus of attention. However, this research has not yet analyzed the possible influence of the phosphorothioate modification on the gut microbiome. It is my goal to study the influence of thus modified bacteria on the gut microbiome and its possible role in diseases like chronic inflammatory bowel disease. During my 2 years of research with the Dedon group it will be my goal to quantify phosphorothioate modifications in bacterial communities of the mouse gut microbiome. I will establish a sensitive LC-MS/MS method for analysis of the global phosphorothioate modification level. This method will be used to observe changes in the phosphorothioate modification level in antibiotic treated and control mice. An increase in the phosphorothioate levels would be a first hint for an advantage of bacteria, producing the modification, confronted with antibiotic-induced oxidative stress. To study the possible shifts in bacterial populations upon antibiotic treatment, deep sequencing will be applied. Therefore, enrichment of phosphorothioate modified DNA will be optimized by Hg-affinity purification. The final goal is the comparison of the global phosphorothioate modification level (by LC-MS/MS) and the changes of bacterial populations (deep sequencing) induced by antibiotic treatment. The comparison of the data received from both analytical methods will allow an insight into the natural function of the phosphorothioate modification and its connection to health and disease.