Single cell RNA sequencing of skin T cells from patient with mycosis fungoides

Capture and processing of single skin T cells from patient with mycosis fungoides was performed using the Fluidigm C1 Autoprep system. Cells were loaded at a concentration of 2,000 cells/μL onto C1 integrated fluidic circuits (IFC) chips for 5-10 μm cells. All C1 capture sites were microscopically inspected to determine the sites containing only a single cell. Empty sites and those with multiple cells were excluded from further analysis. ERCC (External RNA Controls Consortium) spike-in RNAs were added and served as a control. SMARTer Ultra Low RNA Kit (Clontech) was used for reverse transcription and cDNA preamplification. The cDNA products from each cell were then used to prepare Illumina sequencing libraries and then sequenced on Illumina HiSeq2500 using single-end 100-base reads. Gene expression was quantified using Kallisto and then normalized to TPM values single-cell RNA-seq investigation of a cutaneous T-cell lymphoma