Homo sapiens
Protocols: MCF7 cells transfected with either scramble siRNA or DDX54 targeting siRNAs (#9 or 12), or DDX3X targeting siRNA (#6 or #8) harvested 72 h after transfection. Total RNA was purified from cells using miRNeasy Mini Kit (Qiagen) following manufacturer instructions. Excess of contaminating DNA was removed by On-Column DNase Digestion with the RNase-Free DNase Set (Qiagen), following manufacturer's instructions. Ribosomal RNA was depleted with Ribo-Zero rRNA Removal Kit (Human/Mouse/Rat) from Illumina, following manufacturer's instructions. All conditions were produced in three independent biological replicates. RNA seq libraries were produced with NEXTflex RNA-Seq Kit for Illumina Sequencing from Bioo Scientific, following manufacturer's instructions. The following Illumina barcodes were used. Sample Sequence TruSeq code Scrambled REP 1 CGATGT A002 DDX3 siRNA #6 REP1 TGACCA A004 DDX3 siRNA #8 REP1 ACAGTG A005 DDX54 siRNA #9 REP1 GCCAAT A006 DDX54 siRNA #12 REP1 CAGATC A007 Scrambled REP 2 CTTGTA A012 DDX3 siRNA #6 REP 2 ATCACG A001 DDX3 siRNA #8 REP 2 TTAGGC A003 DDX54 siRNA #9 REP 2 ACTTGA A008 DDX54 siRNA #12 REP 2 GATCAG A009 Scrambled REP 3 TAGCTT A010 DDX3 siRNA #6 REP 3 GGCTAC A011 DDX3 siRNA #8 REP 3 AGTCAA A013 DDX54 siRNA #9 REP 3 AGTTCC A014 DDX54 siRNA #12 REP 3 ATGTCA A015 RNA seq libraries were produced with NEXTflex RNA-Seq Kit for Illumina Sequencing from Bioo Scientific, following manufacturer's instructions.