Premo FUCCI Cell Cycle Sensor
Developed by Miyawaki and colleagues , the Invitrogen Premo FUCCI (fluorescence ubiquitination cell cycle indicator) Cell Cycle Sensor is a genetically-encoded, two-color (red and green) indicator that lets you follow cell division within a cell population. We have incorporated the FUCCI genetic constructs into the powerful BacMam gene delivery system, creating a simple and efficient method for labeling cells and following their division.
How Premo FUCCI Cell Cycle Sensor Works
With Premo FUCCI sensor you can:
- Monitor changes in cell proliferation with direct visual readout of cell cycle phase (Figure 1) for every cell in the population
- Efficient BacMam transfection in wide variety of cultured, primary, and stem cell types
- Independently control expression levels of each protein component to optimize balance
The Premo FUCCI Cell Cycle Sensor consists of a fluorescent protein-based system that employs both a red (RFP) and a green (GFP) fluorescent protein fused to different regulators of the cell cycle: cdt1 and geminin.
During the cell cycle, these two proteins are ubiquitinated by specific ubiquitin E3 ligases, which target them to the proteasome for degradation. The E3 ligases display temporal regulation of their activity which results in the biphasic cycling of geminin and cdt1levels during the cell cycle. In the G1 phase of the cell cycle, geminin is degraded; therefore, only cdt1 tagged with RFP is present and appears as red fluorescence within the nuclei. In the S, G2, and M phases, cdt1 is degraded and only geminin tagged with GFP remains, resulting in cells with green fluorescent nuclei.
During the G1/S transition, when cdt1 levels are decreasing and geminin levels increasing, both proteins are present, and when the green and red images are overlaid, nuclei appear yellow fluorescent. This dynamic color change, from red-through-yellow-to-green, represents the progression through cell cycle and division (Figures 2 and 3).
Figure 1. Time-lapse series of transduced U2OS cells. U2OS cells were transduced with Premo FUCCI Cell Cycle Sensor and allowed to divide. Images were collected over a 15-hour timeframe.
Figure 3. Imaging cell cycle progression in live cells with Premo FUCCI Cell Cycle Sensor. U2OS cells were transduced with Premo FUCCI Cell Cycle Sensor and allowed to divide. Images were collected over a 15-hour timeframe.
Table 1. Premo FUCCI Cell Cycle Sensor specifications
|Readout||As a cell progresses through the cell cycle, fluorescence of the Premo FUCCI sensor changes from red to green|
|Common filter set||RFP||FITC|
|Stains fixed cells||No|
|Format||2 x 200 μL||2 x 1,000 μL|
The BacMam system uses a modified insect cell virus (baculovirus) as a vehicle to efficiently deliver and express genes in mammalian cells with minimum effort and toxicity.
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For Research Use Only. Not for use in diagnostic procedures.