For quantitative RT-PCR analyses of pooled cultured cells, RNA was isolated using the RNAqueous Kit (Applied Biosystems), treated with DNase (Applied Biosystems) and reverse transcribed with Superscript III (Invitrogen). mRNA levels were quantified by RT-PCR assay using the Applied Biosystems 7900HT Fast real-time PCR system and RQ analysis software. For quantitative RT-PCR analyses of single cells, single EGFP-labeled iN cells (n = 1) were sorted by FACS into individual wells of 96-well PCR plates using a protocol built into the FACSAriaII flow cytometer’s software package, and mRNA levels were measured using the Fluidigm Biomark dynamic array system as previously described39. Information on TaqMan assays used in the expression assay can be found in Supplementary Table 2. Human specific primer sets used to analyze collybistin mRNA expression included collybistin forward (5′-ATGATTGACA TTGCTATCGA-3′) and reverse (5′-CCAGTCTAGG ACAGAAGCC-3′); GAPDH forward (5′-TTGAGGTCAA TGAAGGGGTC-3′) and reverse (5′- GAAGGTGAAG GTCGGAGTCA-3′).
