H1D1T3_fq
Total RNA, free from genomic DNA contamination, was extracted from confluent 35 mm dishes of H1 ESCs on Days 0, 1, 3, 5, 8 and 14 using the Qiagen RNeasy Plus Mini Kit (Valencia, CA).   All samples had RNA Integrity Numbers =9.40 as determined by Bioanalyzer 2100 (Agilent Technologies, Santa Clara, CA).  Using the TruSeq kit version 2.5 (Illumina, San Diego, CA), RNA sequencing libraries were prepared using 500 ng total RNA, spiked with Ambion® ERCC external RNA controls according to the manufacturers specifications (Life Technologies, Carlsbad, CA).  Briefly, magnetic bead technology was used to isolate and elute mRNA, which was primed and fragmented to sizes of approximately 300 bp.   First- and second-strand cDNA synthesis was performed using Superscript II reverse transcriptase (Invitrogen, Carlsbad, CA).  After end-repair and A-tailing reactions, unique indices were introduced according to the protocol in order to enable sample multiplexing during the sequencing run.  qPCR-quantitation of the library and sequencing services were performed on an  Illumina HiSeq 2000 platform by the Sequencing Core of the Human and Molecular Genetics Center (Medical College of Wisconsin).  Running two samples per lane generated ~90 million paired reads per sample.