GSM5752710: Participant_9_Calorie_After; Homo sapiens; RNA-Seq
Biopsy of subcutaneous adipose tissue (40 mg) was taken after local anesthesia from 21 individuals by a trained surgeon from Ribeirao Preto Medical School-USP. Systematically, all first biopsies were collected from the right side of the abdomen (first day of each nutritional intervention) and the second biopsy from the left side of the abdomen (last day of each nutritional intervention). The adipose tissue was rapidly transported to the research laboratory. Tissue samples were rinsed in phosphate-buffered saline to remove adhering blood, then were snap-frozen in liquid nitrogen and stored at -80ºC until completion of the study. Libraries were prepared for RNAseq as previously described (ref Dall&Hassing et al 2021, PMID: 34762911). Briefly, RNA was isolated using Trizol (Thermofisher) followed by purification with an RNeasy mini kit (Qiagen). Libraries were prepared using the Illumina TruSeq Stranded total RNA Gold protocol (Illumina). Total RNA was depleted of rRNA, fragmented and converted to cDNA. cDNA was subsequently adenylated to prime for adaptor ligation and cleaned using AMPure beads. DNA fragments were amplified using PCR and subject to a final cleanup. Libraries were quality controlled using a Bioanalyzer instrument (Agilent Technologies). Libraries were subjected to 51-bp paired-end sequencing on a NovaSeq 6000 system (Illumina).