Mir-X Inducible miRNA System (Red)
Mir-X Inducible miRNA System (Red)
Brand: Takara Bio.
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Mir-X Inducible miRNA System (Red)
The Mir-X Inducible miRNA System (Red) is a set of expression vectors that allows you to establish a Tet-regulated microRNA (miRNA) expression system in mammalian cells. An additional feature of the system is that a bright red fluorescent protein marker, mCherry, is coexpressed along with your miRNA, enabling you to verify miRNA expression in intact cells. Inducible expression of the mCherry/pre-miRNA transcript is accomplished by creating a double-stable cell line that expresses the Tet-On Advanced transactivator and that bears your customized pmRi-mCherry/miRNA vector, and then culturing the cell line in the presence of the tetracycline derivative, doxycycline.
Overview
- Track your inducible miRNA expression with fluorescent protein markers
- High fold-induction and low basal expression
Applications
- Tetracycline-inducible miRNA expression in mammalian cells
Tetracycline-inducible microRNA
Tetracycline-inducible microRNA. In the presence of doxycycline (Dox), the Tet-On Advanced protein binds to the inducible promoter, PTight, and elicits high levels of transcription of a composite mRNA that encodes a fluorescent protein (mCherry or ZsGreen1) and your miRNA sequence in its 3’ UTR. Cells that fluoresce also express your microRNA.
Tetracycline-inducible miRNA
Tetracycline-inducible miRNA. The RNAiMonitor System was used to create vectors that express a luciferase reporter containing miR-1 or miR-9 cognate target sequences in their 3’ UTRs (3 each). These reporters were then co-transfected with pmRi-ZsGreen1 vectors containing miR-1 or miR-9 miRNA inserts into MCF-7 Tet-On Advanced cells. The parent pmRi-ZsGreen1 vector provided a negative control for each experiment. Inducing miR-1 or miR-9 expression with Dox resulted in >90% reduction in normalized luciferase activity
Tetracycline-inducible shRNA in MCF-7 cells
Tetracycline-inducible shRNA in MCF-7 cells. MCF-7 Tet-On Advanced cells expressing an miR-1 miRNA construct from the pmRi-mCherry and pmRi-ZsGreen1 vectors, exhibit bright red and green fluorescence, respectively, but only in the presence of doxycycline.
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Tetracycline-inducible microRNA
Tetracycline-inducible microRNA. In the presence of doxycycline (Dox), the Tet-On Advanced protein binds to the inducible promoter, PTight, and elicits high levels of transcription of a composite mRNA that encodes a fluorescent protein (mCherry or ZsGreen1) and your miRNA sequence in its 3’ UTR. Cells that fluoresce also express your microRNA.
Tetracycline-inducible miRNA
Tetracycline-inducible miRNA. The RNAiMonitor System was used to create vectors that express a luciferase reporter containing miR-1 or miR-9 cognate target sequences in their 3’ UTRs (3 each). These reporters were then co-transfected with pmRi-ZsGreen1 vectors containing miR-1 or miR-9 miRNA inserts into MCF-7 Tet-On Advanced cells. The parent pmRi-ZsGreen1 vector provided a negative control for each experiment. Inducing miR-1 or miR-9 expression with Dox resulted in >90% reduction in normalized luciferase activity
Tetracycline-inducible shRNA in MCF-7 cells
Tetracycline-inducible shRNA in MCF-7 cells. MCF-7 Tet-On Advanced cells expressing an miR-1 miRNA construct from the pmRi-mCherry and pmRi-ZsGreen1 vectors, exhibit bright red and green fluorescence, respectively, but only in the presence of doxycycline.
