GSM7596430: TCDCA-2 H-Treg; Homo sapiens; RNA-Seq
Post culture cells were snap frozen prior to RNA isolation, further cells were lysed with lysis/binding buffer. Total RNA was extracted according to the protocol and was eluted with 60 µl Elution Buffer. RNA concentration was measured by Nanodrop and its integrity was determined by Bioanalyzer. NEBNext Poly(A) mRNA magnetic isolation module was used for polyA RNA purification with a total of 50 ng to 1 µg of good quality total RNA as input. ployA RNA isolation was done using SMARTer Apollo NGS library prep system. Libraries for RNA-seq was prepared by using NEBNext Ultra II Directional RNA Library Prep kit as per manusfacturers protocol. To study differential gene expression, individually indexed and compatible libraries were proportionally pooled (~25 million reads per sample in general) for clustering in cBot system (Illumina, San Diego, CA). Libraries at the final concentration of 15 pM were clustered onto a single read (SR) flow cell v3 using Illumina TruSeq SR Cluster kit v3 and sequenced to 51 bp using TruSeq SBS kit v3 on Illumina HiSeq system