GSM5733005: PDX2_RNA_sgNS_rep2; Homo sapiens; RNA-Seq
Qiagen miRNeasy Mini Kit Each RNA sample was spiked in with an appropriate amount of either Mix1 or Mix2 according to Life Technologies' guidelines which would lead to about 1% of the total number of RNA-Seq reads mapping to the 92 ERCC control sequences. 300 ng total RNA of each sample was used for library preparation according to the manufacture's protocol of KAPA mRNA HyperPrep Kit (Illumina Platforms). 10 cycles of PCR amplification were performed. KAPA Pure Beads (Kapa Biosystems) were used for library purification. The quality of each library was verified using a Qubit fluorometer (Thermo Fisher Scientific) and further evaluated by the 2100 Bioanalyzer (Agilent Technologies). Sequencing was performed on an Illumina® Hiseq 2500 (Illumina, San Diego, CA) instrument using the TruSeq SR Cluster Kit V4-cBot-HS (Illumina®) to generate 51-nt single-end reads sequencing with v4 chemistry. Quality control of RNA-Seq reads was performed using FastQC.