Expression profiling of mouse CD4+ T cells following Klf1 retrovirus vector transfection

Krüppel-like factor 1 (Klf1) is a transcription factor that promotes ß-globin synthesis in erythroid progenitors, and its role in immunological homeostasis is unknown. The full-length mRNA of mouse Klf1(NM-010635) was cloned into the pMIG retroviral vector, which contains an IRES-GFP cassette. Splenocytes from C57BL/6 mice were cultured for 48 h in the presence of Concanavalin A (ConA) (10 µg/mL) and IL-2 (50 µg/mL), and gene transduction was conducted. CD4+ T cells that were transfected with retroviral vectors were divided into 3 fractions according to GFP fluorescence (strongly positive, weakly positive and negative) using a FACSVantage flow cytometer. Total RNA was then extracted. Sequence libraries were prepared according to standard methods using Smart-seq2 (Illumina), and they were analyzed with a next generation sequencer, a MiSeq system (Illumina).