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Immunology T cell Tools

Study the balance of Tregs and Th17 cells

Expression vectors and reporters

  • Overexpress Human/Mouse Foxp3
  • Overexpress Human/Mouse RORγt
  • Foxp3, RORγt and IL-17 reporters
  • Jurkat T cell lines

Tools to study T helper 17 (Th17 cells)

   

T cell helper 17 (Th17) cells play a role in the inflammatory response. The characteristics of this subtype of T cell are that they secrete abundant amounts of the pro-inflammatory cytokine IL-17 and express the RORC/RORγt transcription factor.

Overexpress RORC/RORγt with lentivectors and piggyBac transposons

Use lentivectors or piggyBac transposons with dual GFP and Puro markers to stably overexpress human RORC or mouse RORγt. The mouse RORγt lentivector (TCL300A-1) and the mouse piggyBac transposon construct (TCL300PB-1) were transiently transfected into 293FT cells. After 24 hours, cellular lysates were prepared and Western blot analyses performed to detect the RORγt protein expression using a mouse RORγt antibody. Robust overexpression of mouse (lanes 1,2) RORγt proteins were observed when compared to the nontransfected cell lysate control (lane 3). Tubulin levels were determined using an anti-tubulin antibody to serve as loading controls. The lentivector and piggyBac plasmid maps and Western blot data for the RORγt lentivector and piggyBac transfection experiments are shown below.

Jurkat stable T cell lines overexpressing either human RORC or mouse RORγt

Human Jurkat T cells were transduced with either TCL200A-1 (human) and TCL100A-1 (mouse) packaged lentivirus and stable cell lines established using puromycin (2.5ug/ml) selection for 7 days. The resulting stable cell lines were evaluated for GFP levels using flow cytometry and cellular lysates tested using Western blots for the overexpression of RORγt. The flow cytometry and Western blot data are shown below.