title: GSM2826030: Injured Input  Rep2; Mus musculus; ChIP-Seq
library_construction_protocol: Seven days after surgery, mice were sacrificed and the three DRGs innervating the sciatic nerve on each side (six ganglia in total) were extracted and placed in HibernateE (BrainBits). Once the DRGs from three mice had been extracted and pooled together, the samples were gently spun down at 80 x g for 2 minutes. The pellet was then resuspended in 10mL of Complete Tissue Fixation Solution (Active Motif) containing 1% fresh formaldehyde and fixed at room temperature for ten minutes. The ActiveMotif ChIP-IT High Sensitivity Kit was then used to perform the remainder of the immunoprecipitation process with the following alterations: 1) TE + 1% SDS was used instead of ActiveMotif's Chromatin Prep Buffer in Section C step 9; 2) 400µL of ChIP Buffer was used for the sonication, which was carried out in 15mL Bioruptor Pico Tubes with 250mg of sonication beads (Diagenode; Catalog # C01020031); and 3) the sonication was performed on a Bioruptor Pico sonicator for 7 cycles of 30 seconds on and 30 seconds off per cycle with the water bath maintained at 4C continuously (Diagenode). Each chromatin preparation made in this manner yielded enough material for an input and two immunoprecipitation reactions. Immunoprecipitations were carried out according to the ChIP-IT High Sensitivity kit instructions using 4µg of polyclonal rabbit anti-Jun (SantaCruz Biotechnology; Catalog # sc-1694X) antibody. The eluates of each pair of IPs using the anti-Jun antibody from the same chromatin preparation were pooled together. Kapa Hyperprep was used for library preparation. Samples were sequenced on an Illumina NextSeq 500 generating 75bp single-end reads
attributes.tag: GEO Accession
attributes.value: GSM2826030