| ```markdown | |
| # Goal/Experiment: | |
| To acquire photosynthetic efficiency (F<sub>v</sub>/F<sub>m</sub>) and quantum yield of photosystem II (Y(II)) of Microcystis aeruginosa cultures using far-red acclimation with the PHYO-PAM-II Compact Version. | |
| # Application of PHYTO-PAM-II (Compact Version) For Running Rapid Light Curves on Cyanobacterial Samples | |
| **Forked from**: [Application of PHYTO-PAM-II (Compact Version) on Aureococcus anophagefferens cultures for photosynthetic efficiency and quantum yield of PSII](https://dx.doi.org/10.17504/protocols.io.eq2ly7jqelx9/v1) | |
| ### Citation: | |
| Gwen Stark, Emily E. Chase, Steven W. Wilhelm 2022. Application of PHYTO-PAM-II (Compact Version) For Running Rapid light curves on Cyanobacterial samples. protocols.io https://dx.doi.org/10.17504/protocols.io.eq2ly7jqelx9/v1 | |
| ### Contributors: | |
| Gwen Stark<sup>1</sup>, Emily E. Chase<sup>2</sup>, Steven W. Wilhelm<sup>1</sup> | |
| 1. The University of Tennessee, Knoxville | |
| 2. University of Tennessee, Knoxville | |
| --- | |
| ## Abstract | |
| A protocol to acquire photosynthetic efficiency (F<sub>v</sub>/F<sub>m</sub>) and quantum yield of photosystem II (Y(II)) of Microcystis aeruginosa cultures using far-red acclimation. | |
| ## Keywords | |
| - Photosynthetic efficiency | |
| - Quantum yield | |
| - F<sub>v</sub>/F<sub>m</sub> | |
| - HAB | |
| - Chlorophyll fluorescence | |
| - Cyanobacteria | |
| ## License | |
| This is an open access protocol distributed under the terms of the [Creative Commons Attribution License](https://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. | |
| --- | |
| ## Guidelines | |
| The collection of quantum yield and photosynthetic efficiency is highly sensitive to modifications in sampling protocol. | |
| ## Materials | |
| - PHYTO-PAM-II Compact Version and components | |
| - Laptop with a USB port, and PhytoWin_3 installed | |
| - Measurement cuvettes | |
| - At least 3 mL of culture material for each sample | |
| - 70% Ethanol | |
| - KimWipes | |
| - Necessary materials and setup for both dark adapting cultures, and a no light or low light environment for sampling | |
| ## Before Starting | |
| Familiarize yourself with the PHYTO-PAM-II equipment, manufacturer’s provided manual, and the basics of chlorophyll fluorescence parameters for full use of data collected and accuracy of results. | |
| --- | |
| ## Equipment Preparation | |
| 1. **Setting Up the PHYTO-PAM-II Compact Version:** | |
| - Toggle the power and connect to a laptop via USB. | |
| - Open the PhytoWin_3 program and select "Phyto Compact Unit." | |
| - Ensure the program is in "MEASURE" mode, and that the "ML" light is green (indicating it is ready). | |
| - Ensure that "AL" and "FR" lights are not selected. | |
|  | |
| 2. **Adjusting Settings:** | |
| - Modify Phyto-Pam settings based on the species being tested. | |
| - Use the "View Pulse" check box to observe the saturation pulse kinetics, ensuring a distinct plateau is seen. | |
| --- | |
| ## Sample Preparation | |
| 3. **Sample Transfer:** | |
| - Transfer 3 mL of culture to a quartz cuvette and minimize exposure time. | |
| - Place the cuvette in the optical port and cover with the darkening hood. | |
| 4. **Auto-Adjust Gain:** | |
| - Perform a zero-offset function using Zoff-determination to blank the PHYTO-PAM-II before runs. | |
| 5. **Far-Red Acclimation:** | |
| - Apply far-red light acclimation to induce photosystem I and oxidize the PQ pool. | |
| - Dark acclimation is not recommended for cyanobacteria. | |
| > **Note:** Evaluate the necessity between no acclimation and weak far-red acclimation for consistent results. | |
| --- | |
| ## Measurement Acquisition | |
| 6. **Experimental Measurements:** | |
| - Measurements can now be collected. | |
| 7. **Running Far-Red Light Acclimation:** | |
| - Transfer 3 mL of culture into the cuvette, auto-adjust gain, and run acclimation. | |
| - Turn on the "ML" button and wait for the light to turn green. | |
| 8. **Taking Measurements:** | |
| - Choose F<sub>v</sub>/F<sub>m</sub> measurements or run a rapid light curve. | |
| - Adjust PAR and exposure times appropriately. | |
| - Aim for the ETR curve to hit a maximum peak and level off. | |
| 9. **Recording Data:** | |
| - Data will be recorded automatically. | |
| - Use 70% ethanol to clean the cuvette between different treatments. | |
| 10. **Data Storage:** | |
| - Save and export data as a .csv file. | |
| - Unplug the equipment and charge fully for long-term storage. | |
| > **Recommendation:** Regularly charge the equipment if stored unused for extended periods. | |
| --- | |
| ### endofoutput | |
| ``` |