GSM5416963: CAD_PAT_1; Homo sapiens; RNA-Seq
We took a sample of PAT (1.0 g) adjacent to the pericardial surface during surgery. The adipose sections collected from 5 CAD and 4 control patients were kept in 10% formalin for histological analysis. The other adipose tissues were immediately snap-frozen in liquid nitrogen (–80°C). Five CAD and 4 control adipose tissue samples underwent RNA sequencing. Paired-end libraries were synthesized using the TruSeq™ RNA Sample Preparation Kit (Illumina, USA) following the TruSeq™ RNA sample preparation guide. Briefly, the poly-A containing mRNA molecules were purified using poly-T oligo-attached magnetic beads. Thereafter, the mRNA was fragmented into small pieces using divalent cations under 94°C for 8 min. The cleaved RNA fragments were copied into first-strand cDNA using reverse transcriptase and random primers. Subsequently, the second strand cDNA was synthesized using DNA Polymerase I and RNase H. These cDNA fragments then underwent an end repair process, the addition of a single 'A' base, and ligation of the adapters. The products were purified and enriched with PCR to generate the final cDNA library. Clean libraries were quantified using a Qubit® 2.0 Fluorometer (Life Technologies, USA), and validated by Agilent 2100 bioanalyzer (Agilent Technologies, USA) to confirm the insert size and evaluate the mole concentration. Cluster was generated using cBot with the library diluted to 10 pM and then was sequenced using the Illumina NovaSeq 6000 (Illumina, USA).