GSM1668879: Replicate_2; Mus musculus; ChIP-Seq
Nascent strands (NS) were purified as previously described in details (Cayrou et al., 2012b). Briefly, after extraction with DNAzol, NS were first separated from genomic DNA by sucrose gradient. Several fractions of interest containing NS DNA (0.5 to 2 Kb range) were phosphorylated by T4 polynucleotide kinase, and digested with -Exo (100-150 units that correspond to 300-500 -Exo units/g of DNA) overnight at 37 °C to eliminate contaminating DNA. A second round of phosphorylation/digestion, using the same conditions, is performed before purification of the samples for library preparation. Three samples were purified from three independent cell cultures. The RNase A controls were obtained by treating NS-containing fractions with 100 ng/ml RNAse A before -Exo digestion to remove RNA primers at the 5' end of the NS. Another control we also used was genomic DNA sonicated to the same size of the NS DNA and treated by -Exo in the same conditions used for NS samples. To allow reliable comparison with the NS samples, around 1g of sonicated DNA was thereby treated. Libraries were prepared for sequencing using standard Illumina protocols
GEO Accession
GSM1668879