GSE203395
RNA-seq data from PBMC of M.tb-infected and M.tb-uninfected infants in MVA85A clinical trial
Frozen PBMC from infants 7 days before randomization (PBMC from two infant were 28 days after randomization) were stimulated in RPMI with 10% Feotal calf serum (FCS) at 5 million/ml with 1 million CFU/ml BCG (SII) or left unstimulated in a total volumn of 200ul/well in 96-well plates. Cells were incubated  for 12 hours (37C, 5% CO2). Plates were centrifuged and 200ul supernatant was removed without disrupting the pellet and transferred into clean U-bottom plates. The plates were centrifuged again, the supernatant were flicked-out  and the cells were resuspended in 200ul RLT buffer with beta-mercaptoethanol (10ul/ml). The samples were mixed with the buffer to lyse the cells and the plates were sealed and stored at -20C. At the time of RNA-extraction, plates were thawed at 37% for 10 mins. RNA extraction was performed using the Qiagen RNeasy kit following manufacturer's instructions (including optional DNase digestion) and RNA was eluted twice, 30 ul per elution. RNA was quantified by nanodrop and samples were stored at -80C. Overall design: Frozen PBMC from M.tb-infected and M.tb-uninfected infants 7 days before randomization (except that PBMC from 2 M.tb-infected infant were 28 days after randomization) were stimulated with eitehr BCG or media for 12 hours, RNA were then extracted.