Part:BBa_K2492003
GEM-GECO, a calcium indicator. When bound with calcium, GEMgeco has blue shift of its emission.
GEM-GECO is a kind of calmodulin-GFP chimeras, and it fluoresces green without Ca2+ but blue with Ca2+ [1] .GEM-GECO is used as a calcium ion indicator, indicating the visualization of neuron activity( AWA for instance). In recent advances, calcium is a universal indicator for neuron excitation, because calcium ions are able to influx at action potential. GEM-GECO is capable of monitoring and responding to the concentration of calcium ions due to the existence of fluorescence resonance energy transfer between two chromophores inside of the functional domain. The reason why GEM-GECO enables the response of disparate calcium ion concentration is that the chromophore of blue-green emission ratiometric undergoes excited-state proton transfer and emits from the low-energy (green fluorescent) anionic form in the calcium ions free state[2], while in the calcium ions bound state,two chromophores are less compact and photon transfer is blocked, resulting the blue fluorescent form. Although we utilise CoCHR in AWA for efficient manipulation, the excited or inhibited state of neuron is not able to be obtained timely. In order to visualise whether the channelrhodopsin-CoCHR is excited or not, we fuse GEM-GECO after CoCHR for the visualisation of neuron activity. As soon as we activate or suppress the channelrhodopsin to manipulate the condition of neuron, the change of calcium ion concentration tigers emission of GEM-GECO from one fluorescent type to another type. For example, we use 405nm wavelength to excite GEM-GECO, and receive light at 511nm and 462nm. If calcium bound to GEM-GECO, the received wavelength is close to 462nm, indicating the neuron excitation. As for further researches, if we intend to observe the response of AWA, that is, investigating whether C.elegans prefer or repel to the certain substance, the change of fluorescent reveals the condition of AWA. Moreover, GEM-GECO is also benefit for the large radiometric change and it decreases the cross talk between channelrodpsin-CoCHR. Meanwhile, the fusion protein mcherry which is a marker for CoCHR position is also able to mark GEM-GECO due to expression linkage.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI site found at 1064
Illegal SapI.rc site found at 970
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