Determination of the respiration rate of plant leaves and tissues
Experiment preparation:
Apparatus: a set of oxygen electrode system; 30ml syringe, scissors, double-sided knife (or single-sided knife), 1ml pipette, tweezers, pipette, absorbent paper, hole punch.
Reagents: (1) Determination medium 50mmol / L Tris-HCl buffer solution pH7.5. Weigh out 6.05 grams of Tris (trishydroxymethylaminomethane) 800-900ml of water, dropwise add 1mol / L HCl, adjust the pH to 7.5, and add 1000ml of water. In addition, the measurement medium can also use phosphoric acid, Hepes, Tricine and other buffers with pH7.5 50mmol / L. Only the aqueous solution with pH 7.5 can also be used as the measuring medium.
(2) 0.5mol / L KCl: Weigh 1.86g of KCl and dissolve in 50ml of distilled water.
Experimental steps:
1 Prepare the required instruments and reagents, install the aerobic electrode, and connect it to the computer for calibration (for detailed operation steps, please refer to the manufacturer's instructions or Chinese instructions).
2 Preparation of materials: take the leaves (tissues) of plants such as wheat and rice, take a certain area (or weigh a certain weight) with a puncher (generally 0.5cm2 leaf debris should be sampled per ml reaction volume),
Put the leaf pieces in a small beaker containing the measuring medium, and then put the small beaker in a vacuum to evacuate, expel the gas in the cell gap of the leaf (tissue), and make the leaf sink to the bottom. or
Pour the leaf fragments into a 30ml syringe, perform manual vacuum infiltration, remove the gas in the leaves, and let the leaves sink. Then take out the sinking leaf block and use scissors to cut it into pieces with an area of ​​about 2mm2
Piece into the oxygen electrode reaction cup.
3 Determination of respiration rate:
Add 1 ~ 2 ml of the measuring medium to the reaction cup containing the leaf fragments, and then put it into the magnetic rotor. Start the magnetic rotor to stir the solution. After equilibrating for about 1 minute, start the measurement. After 3 to 5 minutes
Stop the measurement. For details on the use of oxygen electrodes, please refer to the Chinese manual.
Note: If a transparent reaction cup is used, it is necessary to block the reaction cup when measuring the respiration rate of the blade.
4 After the reaction, wash the cuvette. Repeat step 3, after all experiments, follow the instructions to clean the cuvette, clean the electrode, and keep the electrode dry. (See notes in the instructions)
5 Calculation of respiration rate: Obtain the amount of oxygen change per unit time from the recording curve, and calculate the respiration rate of the sample based on the leaf area and the volume of the reaction medium.
The oxygen electrode models that can be used to complete the experiment are: Leaflab2 (only used to determine the blade), Chlorolab2, Chlorolab3, Oxygraph, Oxytherm.
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