Illumina HiSeq 2500 paired end sequencing; RNA sequencing of endocrine resistant breast cancer from human patients
RNA sequencing of endocrine resistant breast cancer from human patients
Surgical specimens were collected and either (i) snap-frozen and stored at -70oC or (ii) fixed in formalin and embedded in paraffin. For fresh frozen tissues, RNA and DNA were isolated using QIAGEN RNAeasy and DNAeasy kits, respectively, according to the manufacturer's protocols. For FFPE tissues, RNA and DNA were isolated with QIAGEN AllPrep FFPE Kit, according to the manufacturer's protocols. RNA quality was assessed with an Agilent BioAnalyzer RNA 6000 Nano Kit.  Fresh-frozen RNA ribo-zero library was prepared with rRNA removed from total RNA using Epicentre's Ribo-Zero rRNA Removal kit (Cat# RZH11042). 30-100 ng Ribo-Zero RNA was used for the construction of the library using the Illumina TruSeq™ RNA Sample Prep Kit (Cat# RS-122-2001) and the manufacturer's instructions were followed, except for omitting the purification step before fragmentation. FFPE RNA was prepared with the Illumina TruSeq FFPE RiboZero Gold protocol according to the manufacturer's instructions.