RNA-seq of Homo sapiens: UG11-70-RT
Total RNA was extracted using Qiagen RNeasy mini kit, according to the manufacturers protocol, including a DNase digestion. Concentration and quality of the total extracted RNA was checked by using the 'Quant-it ribogreen RNA assay' (Life Technologies, Grand Island, NY, USA) and the RNA 6000 nano chip (Agilent Technologies, Santa Clara, CA, USA), respectively. Subsequently, 500 ng of RNA was used to perform an Illumina sequencing library preparation using the QuantSeq 3' mRNA-Seq Library Prep Kits (Lexogen, Vienna, Austria) according to manufacturer's protocol. Libraries were quantified by qPCR, according to Illumina's protocol 'Sequencing Library qPCR Quantification protocol guide', version February 2011. A High sensitivity DNA chip (Agilent Technologies, Santa Clara, CA, US) was used to control the library's size distribution and quality. Sequencing was performed on a high throughput Illumina NextSeq 500 flow cell generating 75 bp single reads.