Abstract
A powerful method to determine the methylation status of specific cytosine residues within RNA is bisulfite sequencing. In combination with high-throughput sequencing methods cytosine methylation can be determined at nucleotide resolution on a transcriptome-wide level. Nevertheless, several critical aspects need to be considered before starting such a project. Below we describe a detailed step-by-step protocol for planning and performing a transcriptome-wide bisulfite sequencing experiment and subsequent data analysis to determine methyl-cytosine in poly(A)RNA from cells and tissues.
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