Journal: PLoS Genetics
Article Title: Transcriptome-Wide Mapping of 5-methylcytidine RNA Modifications in Bacteria, Archaea, and Yeast Reveals m5C within Archaeal mRNAs
Figure Lengend Snippet: RNA immunoprecipitation with modification-specific antibodies. Shown is the coverage of Illumina-sequenced cDNA following RNA fragmentation, antibody pulldown, reverse transcription and sequencing. Black line, pulldown performed with an anti-5-methylcitosine (hm 5 C) antibody; green line, pulldown performed with an anti-5-hydroxy-methylcitosine antibody; red line, input RNA (no antibody applied). X-axis, position along the genome; Y-axis (right), read coverage of the sequenced anti-m 5 C library; Y-axis (left), fold enrichment of peaks related to median coverage along the gene. The coverage of the anti-hm 5 C and input libraries was normalized using the median of the anti-m 5 C library as a reference point. (A) The 23S gene of S. solfataricus . Peaks corresponding to positions 2121 and 2643 in the gene (875,473 and 875,995 relative to the S. solfataricus genome, respectively) are marked. Another peak, which did not come up in our bisulfite-based analysis, is observed around position 2760. (B) The 16S gene of S. solfataricus . A single peak corresponding to position 1369 in the gene (position 873,040 relative to the genome) is marked. (C–E) Antibody pulldown of m 5 C modifications in protein-coding genes from Table 3 .
Article Snippet: The mixed bisulfite-treated RNA sample was used as a template for cDNA library preparation according to the mRNA-seq Illumina protocol, omitting the polyA-based mRNA purification step.
Techniques: Immunoprecipitation, Modification, Sequencing