title: GSM2255551: RNA-seq Pax5+/- Pro-B [12810]; Mus musculus; RNA-Seq
library_construction_protocol: Rneasy Plus Mini Kit, Dynabeads mRNA purification kit About 1-5 ng of cDNA or ChIP-precipitated DNA were used as starting material for the generation of sequencing libraries with the NEBNext Quick Ligation Module, NEBNext Endrepair Module and NEBNext dA-Tailing module or NEBNext Ultra Ligation Module and NEBNext End Repair/dA-Tailing module. DNA fragments of the following sizes were selected for the different experiments: 200–500 bp for ChIP-seq and 150–700 bp for RNA-seq with AMPure XP beads (Beckman Coulter). For strand-specific RNA-sequencing, the uridines present in one cDNA strand were digested with uracil-N-glycosylase (New England Biolabs) as described (Parkhomchuk et al. 2009) followed by PCR amplification with the KAPA Real Time Amplification kit (KAPA Biosystems). Completed libraries were quantified with the Agilent Bioanalyzer dsDNA 1000 assay kit and Agilent QPCR NGS Library Quantification kit. Cluster generation and sequencing was carried out by using the Illumina HiSeq 2000/2500 system with a read length of 50 nucleotides (single-read) or 100/125 nucleotides (paired-end) according to the manufacturer’s guidelines.