Abstract
Traditionally in vivo pH measurements in plants are carried out using different fluorescent probes such as BCECF and SNARFs by injecting plant cells or incubating tissues with the probes. This approach, however, carries several problems, the most important one being the handling of the plant cells or tissues. Stress and wounding of the plant can possibly influence the outcome of the pH measurements. Other problems are the difficulties of getting the probes more than a few cell layers into the plant tissue and to get an even distribution of the probe.
Increasing numbers of different variants of the green fluorescent protein, GFP, have become available, some of which are pH sensitive. This offers the opportunity to simply express the pH sensitive protein in different plant tissues, making it possible to conduct in vivo pH measurements in a non-invasive manner.
One sensor being characterized consists of a pH sensitive GFP variant fused to the pH insensitive mRFP. This allows for quantitative ratiometric pH measurements.
Another pH sensor is based on FRET. It consists of the pH insensitive cyan fluorescent protein, CFP, fused to a pH sensitive YFP (yellow). FRET efficiency between the FRET donor, CFP, and the FRET acceptor, YFP, will change as YFP fluorescence is quenched with decreasing pH.
These sensors are being characterized with respect to their sensitivity towards other physiologically relevant ions such as Cl-, K+, Na+, Mg2+, and Ca2+, as well of their ability to react quickly towards changes in pH.
Plants and plant tissues expressing these sensors in different tissues and in different cellular compartments will offer an opportunity to investigate and quantify the physiological role of pH and proton currents and gradients in different situations, for example with respect to transport mechanism across the plasma membrane.
Increasing numbers of different variants of the green fluorescent protein, GFP, have become available, some of which are pH sensitive. This offers the opportunity to simply express the pH sensitive protein in different plant tissues, making it possible to conduct in vivo pH measurements in a non-invasive manner.
One sensor being characterized consists of a pH sensitive GFP variant fused to the pH insensitive mRFP. This allows for quantitative ratiometric pH measurements.
Another pH sensor is based on FRET. It consists of the pH insensitive cyan fluorescent protein, CFP, fused to a pH sensitive YFP (yellow). FRET efficiency between the FRET donor, CFP, and the FRET acceptor, YFP, will change as YFP fluorescence is quenched with decreasing pH.
These sensors are being characterized with respect to their sensitivity towards other physiologically relevant ions such as Cl-, K+, Na+, Mg2+, and Ca2+, as well of their ability to react quickly towards changes in pH.
Plants and plant tissues expressing these sensors in different tissues and in different cellular compartments will offer an opportunity to investigate and quantify the physiological role of pH and proton currents and gradients in different situations, for example with respect to transport mechanism across the plasma membrane.
| Originalsprog | Engelsk |
|---|---|
| Publikationsdato | 2007 |
| Antal sider | 1 |
| Status | Udgivet - 2007 |
| Begivenhed | Bioimaging Workshop - Frederiksberg, Danmark Varighed: 6 jun. 2007 → 7 jun. 2007 |
Konference
| Konference | Bioimaging Workshop |
|---|---|
| Land/Område | Danmark |
| By | Frederiksberg |
| Periode | 06/06/2007 → 07/06/2007 |