Developmental and Injury-induced Changes in DNA Methylation in Regenerative versus Non-regenerative Regions of the Vertebrate (Xenopus laevis) Central Nervous System assayed by 5hmC MeDIP and by ChIP-seq for Histone Markers

Xenopus is uniquely suited for identifying core features of successful CNS axon regeneration, because parts of its CNS (e.g., eye), regenerate axons throughout life, whereas others (e.g., hindbrain) do so only as tadpoles. To aid in the interpretation of bisulfite whole genome methylation sequencing (WGBS) on juvenile frog eye after optic nerve injury, and on hindbrain samples from tadpole and juvenile frog after spinal cord injury during the peak phase of axon regeneration, we performed ChIP-seq for histone modifications associated with active gene expression (H3K4me3 & H3K27ac) and repressed gene expression (H3K27me3 & H3K9me3) on these same tissues, as well as DNA-immunoprecipitation sequencing (DIP seq) for 5-hydroxymethyl cytosine (5hmC) on eye samples during optic nerve regeneration. Overall design: Histone ChIP seq: Two biological replicates for each of the tissues and conditions used previously for WGBS (6 pooled eyes after optic nerve crush, from the operated side and contralateral unoperated side, plus 6 pooled eyes from unoperated animals; 5 pooled hindbrains from tadpoles and 5 from frogs subject to spinal cord injury (SCI) plus 5 pooled hindbrains from age-matched, uninjured tadpoles and frogs each. 5hmC DIP seq: performed on eye samples only, otherwise collected as for WGBS and histone ChIP-seq, with replicates (2) pooled for sequencing.