title: GSM4178145: SKOV3.ip1 cells, Lipo_nov_n1; Homo sapiens; RNA-Seq
library_construction_protocol: RNA was extracted from mock-transfected and siRNA transfected cells 72h post-transfection. cDNA library produced with TGIRT (thermostable group II reverse transcriptase)  Purified total RNA was ribodepleted by using a RiboZero Gold (Human/Mouse/Rat) kit (Illumina). The resulting ribodepleted RNAs were fragmented with an NEBNext Magnesium RNA Fragmentation Module (New England Biolabs) by incubation at 94°C for 7 minutes and were then treated with T4 polynucleotide kinase (T4PNK; Epicentre) to remove 3' phosphates or 2', 3' cyclic monophosphates.  cDNAs were synthesized via TGIRT template-switching with either 0.5 or 1µM TGIRT-III reverse transcriptase (Ingex, LLC) for 15 min at 60o C, during which a DNA oligonucleotide containing the complement of an Illumina Read 2 sequencing primer-binding site becomes seamlessly linked to the 5' cDNA end. After reaction cleanup, a 5' adenylated DNA oligonucleotide containing the complement of an Illumina Read 1 sequencing primer-binding site is then ligated to the 3' cDNA end with Thermostable 5' AppDNA / RNA Ligase (New England Biolabs). Properly ligated cDNAs were amplified by PCR (12 cycles) to synthesize the second strand and add Illumina flowcell capture and index sequences. Libraries were size-selected with Ampure XP beads (Beckman-Coulter) and evaluated on an Agilent 2100 Bioanalyzer
identifiers.namespace: GEO
identifiers.id: GSM4178145