Skip to main content
. 2013 Sep 26;8(9):e75984. doi: 10.1371/journal.pone.0075984

Figure 1. Map of the Tet-Off construct and induction of LDH-A shRNA.

Figure 1

(A) PMVECs were infected with a retrovirus, enabling reverse tetracycline-controlled transactivator protein (tTA) expression. Cells were selected to homogeneity using blasticidin, reinfected with a lentivirus [shown in (B)], and selected to homogeneity using puromycin. The resulting double-transfection enabled the expression of a LDH-A short hairpin RNA (shRNA) in the absence of doxycycline. Bsr, blasticidin resistance gene; EGFP, enhanced green fluorescent protein; HIV RRE, human immunodeficiency virus Rev response element; IRES EMV, encephalomyocarditis virus internal ribosome entry site; LTR, retro/lentiviral long terminal repeat; PAC, puromycin resistance gene; PSV40, simian virus 40 promoter; PTet, doxycycline-regulated promoter; wPRE, woodchuck hepatitis virus posttranscriptional regulatory element; mir, 5’–3’ flanking sequence derived from the murine mir (micro-RNA gene)-15. (C) Doxycycline retrieval promoted expression of the red fluorescent protein mCherry in infected PMVECs.