GSM4626952: Input_C6eIF2α cKICaMKIIα 1hr post-FC_rep2 [D705-D501]; Mus musculus; RIP-Seq
Frozen hippocampi from one mouse were pooled and homogenized in 1 ml of buffer as described27. After centrifugation, 4 μl of anti-HA antibody (Extended Data Table 2) were added to 800 μl of the cleared lysate and incubated for 4 h at 4 °C. Remaining lysate was saved as input sample. After incubation, 200 μl of protein A/G magnetic beads (Thermo Fisher Scientific) were added and incubated overnight at 4 °C with rotation. Immunoprecipitates (IPs) were washed in high salt buffer and RNA from inputs and IPs extracted as described in the original protocol RNA-Seq libraries were prepared from 200 ng of total RNA using the NEBNext Ultra II Directional RNA Library Prep Kit for Illumina with the NEBNext Poly(A) mRNA Magnetic Isolation Module and NEBNext Multiplex Oligos (New England Biolabs). Libraries were pair-end sequenced (2 × 50 bp) on an Illumina Illumina Novaseq 6000 Instrument.
GEO Accession
GSM4626952