title: Illumina HiSeq 1500 sequencing; RNAseq of ovarian cancer ascites cells
design: RNAseq of ovarian cancer ascites cells
library_construction_protocol: Ascites was collected from patients with high-grade serous ovarian carcinoma undergoing primary surgery at the University Hospital in Marburg. Informed consent was obtained from all patients according to the protocols approved by the local ethical committe. Macrophage cells were extract by CD14+ MACS. T cells were extracted by CD3+ MACS. Tumor cells were selected by size (>30um). After Ficoll gradient centrifugation, TAMs were purified by CD14 MACS and lysed in Peqgold (TRIzol-like solution based on phenol and chloroform). RNA was digested with DNase (MN) and purified on a Qiagen Minelute column. Illumina-compatible libraries were made using the Epicentre Scriptseq human/mouse/rat low input Gold kit. Libraries were created using the Illumina Truseq mRNA kit v2 used according to the manufacturer's instructions.