title: GSM2310148: BS-seq_E14_mPGCLC_d4_4; Mus musculus; OTHER
library_construction_protocol: PBAT: For small sample amounts (PGCLC samples) post-bisulfite adaptor tagging (PBAT) libraries were prepared directly from cell lysates following recently described protocols (Miura et al., 2012; Smallwood et al., 2014). PBAT libraries were prepared by bisulfite converting Proteinase K digested cell lysates using the EZ DNA Methylation-Direct Kit (Zymo). After cleanup, 1st strand synthesis was performed using 6N-forward oligos at 37°C for 90 minutes. Subsequently, samples were treated with Exonuclease I for 1 hour at 37°C, and DNA was purified using AMPure XP beads (Agencourt). Samples were eluted in 2nd strand synthesis mix with 6N-reverse oligos and incubated at 37°C for 90 minutes. DNA was purified and amplified with KAPA HiFi HotStart DNA Polymerase (KAPA Biosystems). All amplified libraries were purified and assessed for quality and quantity using High-Sensitivity DNA chips on the Agilent Bioanalyzer. High-throughput sequencing of all libraries was carried out with a 125 bp paired-end protocol on a HiSeq 2000 instrument (Illumina). Genomic DNA and total RNA were isolated from the samples using the Beckman Coulter's Agencourt RNAdvance Cell v2 kit following the manufacture’s instructions.
attributes.tag: GEO Accession
attributes.value: GSM2310148