GSM6476915: Bb A3 MOI 1 8 hrs, Rep_F [MS8_72]; Homo sapiens; RNA-Seq
Total RNA was extracted from 107 mo-DCs using RLT Lysis Buffer (Qiagen) by following manufacturer's instructions NEBNext Poly(A) Magnetic Isolation Module (NEB #E7490) and NEBNext Ultra II Directional RNA Library Prep Kit for Illumina (Cat# E7765) were used to generate libraries.  Specifically, Poly A RNAs were isolated from total RNAs and fragmented for cDNA synthesis. End repair is performed where 3' to 5' exonuclease activity of enzymes removes 3' overhangs and the polymerase activity fills in the 5' overhangs. An 'A' base is then added to the 3' end of the blunt phosphorylated DNA fragments which prepares the DNA fragments for ligation to the sequencing adapters, which have a single 'T' base overhang at their 3' end. Ligated fragments are subsequently size selected through purification using the Sample Purification Beads included in the kit and undergo PCR amplification techniques to prepare the libraries. The BioAnalyzer is used for quality control of the libraries to ensure adequate concentration and appropriate fragment size.  The resulting library insert size is 200bp-500bp with a median size around 300bp. Libraries were uniquely barcoded and pooled for NovaSeq6000 sequencing.