title: GSM7162725: Prostate Cancer Patient 7; Homo sapiens; RNA-Seq
library_construction_protocol: Purified double-stranded cDNA was generated from 150 ng of RNA and amplified using 3' poly(A) selection and random priming throughout the transcriptome. Depending on the degree of sample degradation, double-stranded cDNA products generated with the NuGEN Ovation RNA-Seq System were sheared using the Covaris E220 Focused-ultrasonicator. The fragment sizes were confirmed on the Agilent Bioanalyzer High Sensitivity DNA chip to verify proper shearing. A double-stranded DNA library was created using 100 ng of sheared, double-stranded cDNA, preparing the fragments for hybridization onto a flow cell. Libraries were then pooled in equimolar amounts based on the qPCR quantification and a qPCR was run on the pooled libraries to determine the concentration of the pool for bridge amplification. Using the concentration from the Bio-Rad qPCR instrument, 15 pM of the library pool was loaded onto a flow cell and amplified by bridge amplification using the Illumina cBot instrument. A paired-end 100 bp run was used to sequence the flow cell on a HiSeq 4000 or NovaSeq6000 Sequencing platforms.