GSM7092432: 53371_cre (JAK1 KO); Mus musculus; RNA-Seq
RNA was harvested using Rneasy mini plus kit (Qiagen). Total RNA was extracted from flash-frozen cultured cancer cells using the RNeasy Mini Kit (QIAGEN). QIAGEN's protocol was followed to extract the total RNA. The concentration of the RNA was determined on a NanoDrop spectrophotometer, and the integrity of the RNA was validated using gel electrophoresis. Quality and quantity of the total RNA was re-confirmed by Novogene. Only high quality RNA was used for library prepration.   1.3 ug of total RNA was used for the construction of sequencing libraries. Library construction was performed by Novogene  with the following procedure: After the QC, mRNA was enriched using oligo(dT) beads. Next, the mRNA was fragmented randomly by adding fragmentation buffer, followed by cDNA was synthesis using mRNA template and random hexamer primers. Next a custom second-strand synthesis buffer (Illumina) , dNTPs, RNase H and DNA polymerase I were added to initiate the second-strand synthesis. After a series of terminal repair, A ligation and sequencing adaptor ligation, the double-stranded cDNA library was completed through size selection and PCR enrichment. The quality of the library was confirmed via: (1) Qubit 2.0: tests the library concentration preliminarily. (2) Agilent 2100: tests the insert size. (3) Q-PCR: quantifies the library effective concentration precisely.