title: Early gene expression after YF17D vaccination that is associated with YF17D-specific T cell responses
design: Early gene expression after YF17D vaccination that is associated with YF17D-specific T cell responses
library_construction_protocol: Blood was drawn from subjects into Tempus tubes and RNA extraction performed. RNA isolation from whole blood was performed using the Tempus Spin RNA Isolation Kit, eluted in 90uL elution buffer and stored at -80C. 1 Î¼g total RNA was used for library preparation by Azenta Life Sciences. Poly(A) mRNA isolation was performed using Oligo(dT) beads and mRNA fragmentation was performed using divalent cations and high temperature. Priming was performed using random primers followed by synthesis of first strand and the second-strand cDNA. Purified cDNA was then treated to repair both ends and dA-tailing added, followed by a T-A ligation to add adaptors to both ends. Size selection of adaptor-ligated DNA was then performed using DNA Clean Beads. Each sample was then amplified by PCR using P5 and P7 primers and the PCR products were validated.