title: GSM3082824: WT BOBSC; Gallus gallus; Homo sapiens; OTHER
library_construction_protocol: Upon mixing the 50-60 million BOBSC iPS cells with the DT40 cells (10:1 ratio) , the nuclear fraction was extracted  (then treated with rnaseH for the control),  genomic DNA containing containing DNA:RNA hybrid purified, soluble contaminating RNA was digested with RNase I, sonicated to average size of 300 bp with Bioruptor Diagenodtreatments and immunoprecipitated DNA:RNA hybrids with S9.6 antibody. Following low, high, LiCl and TE buffer washes of Protein-G captured immunocomplexes, DNA:RNA hybrids were eluted, phenol:chloroform purified, denatured for 5 minutes at 95 C, DNA moity digested with DNaseI, and RNA moiety of the DNA:RNA hybrid purified with Trizol. Directional RNA libraries were prepared using NEBNext Ultra II Directional RNA Library Prep Kit (New England Biolabs, E7645) per instructions provided. Steps from random priming to End Prep of cDNA was followed as instructed in for rRNA depleted FFPR RNA samples, and further steps where followed as instructed for purified mRNA or rRNA depleted RNA.
attributes.tag: GEO Accession
attributes.value: GSM3082824