A method and device for determining photosynthetic rate of plant leaves based on carbon 13 isotope labeling

Abstract

The application provides a method and device for determining photosynthetic rate of plant leaves based on carbon 13 isotope labeling, and belongs to the technical field of biology. 13 The application first places the leaves to be measured into an assimilation chamber and labels them with CO2, and then measures the change in C content of specific parts of the leaves before and after labeling by means of an elemental analyzer and a stable isotope ratio mass spectrometer. 13 The photosynthetic rate of the leaves in unit area and unit time can be calculated. 13 The amount of C, i.e. the photosynthetic rate. The method and device can simultaneously determine the photosynthetic rates of multiple parts of the leaves, and greatly improve the determination efficiency.

Description

Technical Field

[0001] This invention belongs to the field of biotechnology, and specifically relates to a method and apparatus for determining the photosynthetic rate of plant leaves based on carbon-13 isotope labeling. Background Technology

[0002] Green plant growth depends on photosynthesis in its leaves; therefore, photosynthetic rate is one of the most important traits reflecting plant growth status. However, the photosynthetic rate varies significantly between different parts of the same leaf. In practice, researchers often measure the photosynthetic rate of only one part of the leaf to represent the whole, but this single-point measurement inevitably introduces errors. In-depth research into the spatial heterogeneity of photosynthetic rate within the leaf is crucial for accurately assessing plant photosynthetic capacity and for accurate modeling of plant-level processes.

[0003] Several methods have been developed both domestically and internationally to determine the heterogeneity of photosynthetic rates in different parts of a leaf, but each has its own limitations. The leaf dry weight method and iodine staining method have large errors due to assimilate transport; the chlorophyll fluorescence method and carbon-14 isotope (radioactive) labeling method can only perform qualitative characterization but not quantitative determination; portable photosynthesis meters can only measure the photosynthetic rate at a single point each time, and the stabilization time before measurement is long, resulting in low efficiency. Summary of the Invention

[0004] To overcome the shortcomings of existing technologies, this invention provides a method based on... 13 A method and apparatus for measuring the photosynthetic rate of plant leaves using C isotope labeling can accurately and efficiently measure the photosynthetic rate of different parts of the leaf.

[0005] The technical solution of the present invention is as follows:

[0006] This invention provides a method for measuring the photosynthetic rate of plant leaves based on carbon-13 isotope labeling. The method involves inducing photosynthesis in plant leaves and then using a method containing carbon-13 isotope labeling... 13 A CO2 mixture was used to assimilate and label the photosynthetically induced leaves; the leaf mass per unit area (LMA), carbon content in dry matter, and other parameters of the assimilated and labeled leaves were measured. 13 The ratio of C to total C is used to calculate the local photosynthetic rate of the leaf according to the following formula:

[0007]

[0008] In the formula, C% is the carbon content in dry matter, and R label It is in the labeled sample 13 The ratio of C to total C, R ref In unlabeled samples 13 The ratio of C to total C, t is the assimilation labeling time, m c yes 13The molecular weight of a carbon atom.

[0009] In one implementation method, the assimilated leaf is divided into several leaf tissues, and the area 'a' of the leaf tissues is measured. leaf Calculate the leaf mass per unit area, LMA, based on the dry weight (DW).

[0010] As one implementation method, leaf tissue is dried and ground into powder, and the carbon content in the leaf dry matter is measured using an elemental analyzer, while the carbon content in the leaf dry matter is measured using a stable isotope ratio mass spectrometer. 13 The ratio of C to total C.

[0011] In one embodiment, the mixed gas comprises a concentration of 400-410 ppm. 13 CO2 and nitrogen and oxygen, the 13 The volume ratio of CO2 to nitrogen and oxygen is 1:2500, and the volume ratio of nitrogen to oxygen in the nitrogen-oxygen mixture is 4:1. The relative humidity of the mixed gas is 50-70%.

[0012] As one implementation method, the flow rate of the nitrogen and oxygen is adjusted to be no less than 40 L·min. -1 The 13 The CO2 flow rate is adjusted according to the nitrogen and oxygen flow rates.

[0013] In one implementation, the photosynthetic induction time is 20-40 min, and the assimilation labeling time is 2-3 min.

[0014] The present invention also provides an apparatus for measuring the photosynthetic rate of plant leaves based on the above method. The apparatus includes a gas source, a gas mixing chamber (3), and an assimilation chamber (4). The gas source is... 13 The CO2 unit (1) and the nitrogen-oxygen unit (2), the 13 The CO2 device (1) and the nitrogen-oxygen device (2) are connected to the gas mixing box (3) through a pipe equipped with a flow meter (8). The gas mixing box (3) is connected to the assimilation chamber (4) through a pipe. One end of the assimilation chamber (4) is connected to the tail gas absorption device (5). A light source (6) is provided above the assimilation chamber (4), and a fan (7) is provided inside.

[0015] In one implementation, the assimilation chamber (4) is made of transparent material, and several chambers are provided, all of which are connected to the gas mixing box (3) and the exhaust gas absorption device (5) through pipes.

[0016] In one embodiment, the assimilation chamber (4) is divided into an upper cover and a lower box. The lower box is provided with a flange and a gasket to seal with the upper cover. One end of the assimilation chamber is a movable opening for inserting the blade to be tested.

[0017] As one implementation method, the exhaust gas absorption device (5) is filled with NaOH solution, which can absorb CO2 in the exhaust gas.

[0018] Beneficial effects:

[0019] This invention, based on the principle of isotope labeling, proposes a novel method for determining the photosynthetic rate of plant leaves. The method involves first placing the leaf to be tested into an assimilation chamber and then using… 13 CO2 was used to label the plant leaves, and then the leaf mass per unit area (LMA), carbon content in leaf dry matter, and other parameters in dry matter were measured. 13 The ratio of C to total C is used to calculate the local photosynthetic rate of the leaf according to a formula. This invention also provides an apparatus for measuring the photosynthetic rate of plant leaves based on the above method. The method and apparatus of this invention can simultaneously measure the photosynthetic rate of multiple parts on the same leaf, significantly improving measurement efficiency and providing high accuracy. Attached Figure Description

[0020] Figure 1 This is a structural diagram of the device for measuring the photosynthetic rate of plant leaves according to the present invention; wherein, 1 is... 13 1 is a CO2 unit, 2 is a nitrogen-oxygen unit, 3 is a gas mixing chamber, 4 is an assimilation chamber, 5 is a tail gas absorption unit, 6 is a light source, 7 is a fan, 8 is a flow meter, and 9 is a blade.

[0021] Figure 2 Photosynthetic rate (A) 13C The distribution pattern of maize within the same leaf.

[0022] Figure 3 for 13 C-labeled new assimilation 2 minutes later 13 The transfer status of C.

[0023] Figure 4 The photosynthetic rate (P) measured by a portable photosynthesis meter n ) and this patent 13 Photosynthetic rate measured by CO2 labeling method (A 13C The comparison between the two lines is shown. The solid line represents the 1:1 line; the dashed line represents the regression fitted line. Detailed Implementation

[0024] Carbon-13 isotopes are important tools in plant physiology and ecology, and are commonly used for tracing. This invention provides a method for measuring the photosynthetic rate of plant leaves based on carbon-13 isotope labeling, by using... 13 CO2 was used to label the leaves, and measurements were taken at specific leaf locations before and after labeling. 13 The change in the ratio of C to total C can be used to accurately and efficiently measure the photosynthetic rate of different parts of the leaf.

[0025] The method of the present invention includes: inducing photosynthesis in plant leaves using a solution containing 400 ppm... 13 A CO2 mixture was used to assimilate and label the photosynthetically induced leaves; the leaf mass per unit area (LMA), carbon content in dry matter, and other parameters of the assimilated and labeled leaves were measured. 13 The ratio of C to total C is used to calculate the local photosynthetic rate of the leaf according to the following formula:

[0026]

[0027] In the formula, C% is the carbon content in dry matter, and R label It is in the labeled sample 13 The ratio of C to total C, R ref In unlabeled samples 13 The ratio of C to total C, t is the assimilation labeling time, m c yes 13 The molecular weight of a carbon atom.

[0028] This invention induces photosynthesis in plant leaves, bringing them to a relatively stable photosynthetic state. As one implementation method, the leaf is placed in a light assimilation chamber for photosynthetic induction. Specifically, one end of the light assimilation chamber is opened, the leaf to be tested is placed inside, and the connection between the leaf and the chamber is sealed, for example, using foam adhesive. A light source is positioned above the light assimilation chamber, and the top cover of the chamber is opened to induce photosynthesis in the leaf. The light source provides sufficient illumination for the plant's photosynthesis. As one implementation method, the light source is an LED light source. As one implementation method, the photosynthetic induction time is 20-40 minutes, more preferably 25-35 minutes, to ensure complete photosynthetic induction of the leaf.

[0029] This invention involves photosynthetically induced leaves undergoing... 13 CO2 assimilation labeling. This invention uses a label containing... 13 A mixed gas of CO2 is used to assimilate and label the blades. The mixed gas of this invention includes, in addition to, a concentration of 400-410 ppm of... 13 In addition to CO2, it also includes nitrogen and oxygen. 13 The volume ratio of CO2 to nitrogen and oxygen is 1:2500, and the volume ratio of nitrogen to oxygen in the nitrogen-oxygen mixture is 4:1. As one embodiment, the nitrogen-oxygen mixture comprises 80% N2 and 20% O2 by volume, which is close to the current atmospheric environment. 13 CO2 gas is mixed with the nitrogen and oxygen gas to obtain the above-mentioned mixed gas. As one embodiment, the... 13 The ratio of CO2 flow rate to the nitrogen and oxygen flow rate is 1:2500. As one embodiment, the injection flow rate of nitrogen and oxygen is adjusted to be no less than 40 L / min. -1 When the flow rate of nitrogen and oxygen is adjusted to 40 L·min-1 At that time, the 13 The CO2 flow rate was adjusted to 16 mL / min. -1 The mixed gas injected in this invention has a similar gas ratio to that in the current environment, thus not altering the photosynthetic state of the plant leaves. The temperature and humidity of the injected mixed gas are as close as possible to those of the current environment to reduce measurement errors during marking caused by drastic fluctuations in temperature and humidity. As one embodiment, the relative humidity of the mixed gas is 50-70%, more preferably 55-65%; the temperature of the mixed gas is 25-30℃, more preferably 27-29℃. The assimilation marking time is preferably 2-3 minutes, ensuring sufficient assimilation on the leaf markings. 13 The carbon isotope meets the measurement requirements and can also prevent the assimilate from being transported to different parts of the leaf due to excessively long labeling time, thus causing errors.

[0030] This invention involves removing assimilated and labeled leaves and dividing them into several leaf tissues according to the detection purpose, avoiding the midrib during the division. As one embodiment, the assimilated and labeled leaves are uniformly cut into several segments from top to bottom. The leaf mass per unit area (LMA), carbon content in dry matter, and other parameters are measured for each leaf tissue. 13 The ratio of C to total C.

[0031] As one implementation method, the method for measuring leaf mass per unit area (LMA) is as follows: Measuring the leaf area (a) of leaf tissue. leaf cm 2 After drying, the dry weight (DW, g) and leaf mass per unit area (LMA, g·cm³) were measured. -2 It equals the dry weight of the leaf tissue divided by the leaf area of ​​the leaf tissue.

[0032] As one implementation method, leaf tissue is dried and ground into powder, and the carbon content in the leaf dry matter is measured using an elemental analyzer, while the carbon content in the leaf dry matter is measured using a stable isotope ratio mass spectrometer. 13 The ratio of C to total C. The specific determination method should follow the instrument's instruction manual and standard operating procedures in this field.

[0033] The local photosynthetic rate of a leaf can be calculated using the following formula:

[0034]

[0035] In the formula, C% is the carbon content in dry matter (dimensionless); R label It is in the labeled sample 13 The ratio of C to total C (dimensionless); R ref In unlabeled samples 13 The ratio of C to total C (dimensionless); t is the assimilation labeling time (s); mc yes 13 The molecular weight of C atom (13 g·mol⁻¹) -1 ).

[0036] The method described above can simultaneously measure the photosynthetic rate of multiple parts on the same leaf, greatly improving the measurement efficiency and providing high accuracy of the measurement results.

[0037] The present invention also provides an apparatus for measuring the photosynthetic rate of plant leaves based on the above method. The apparatus includes a gas source, a gas mixing chamber 3, and an assimilation leaf chamber 4. The gas source is... 13 CO2 unit 1 and nitrogen-oxygen unit 2, the 13 CO2 device 1 and nitrogen-oxygen device 2 are connected to the gas mixing box 3 through a pipe equipped with a flow controller 8. The gas mixing box 3 is connected to the assimilation chamber 4 through a pipe. One end of the assimilation chamber 4 is connected to the exhaust gas absorption device 5. A light source 6 is installed above the assimilation chamber 4, and a fan 7 is installed inside.

[0038] The assimilation chamber 4 of this invention is used to process the blade. 13 The primary container for CO2 labeling is preferably made of a transparent material, such as acrylic sheet. In one embodiment, the assimilation chamber 4 is divided into an upper cover and a lower box body. The lower box body is equipped with a flange and gasket to seal it to the upper cover. One end of the assimilation chamber has a movable opening for inserting the leaf to be tested.

[0039] In one embodiment, the light source 6 of the present invention is preferably an LED light source, which is disposed directly above the assimilation chamber 4.

[0040] The assimilation chamber 4 of the present invention is equipped with a fan 7 for mixing the gas inside. Preferably, the fan 7 is located near the air inlet of the assimilation chamber 4.

[0041] In one embodiment, the assimilation chamber 4 of the present invention is connected to the gas mixing box 3 at one end and the tail gas absorption device 5 at the other end. The mixed gas in the gas mixing box 3 is injected into the assimilation chamber 4 and flows out from the pipe connected to the tail gas absorption device 5. In another embodiment, the tail gas absorption device 5 contains a NaOH solution to absorb the remaining gas. 13 CO2 is used to prevent pollution. The concentration of the NaOH solution can be selected as 0.1-0.2 mol·L⁻¹. -1 .

[0042] As one implementation method, the 13 CO2 device 1 and nitrogen-oxygen device 2 are high-pressure gas cylinders, which are pressurized to supply the gas... 13 CO2 gas and nitrogen-oxygen mixture are injected into the gas mixing chamber 3. As one embodiment, in the...13 Flow controllers 8 are installed in the gas output pipelines of CO2 unit 1 and nitrogen-oxygen unit 2 to control the flow rate. 13 The outflow rates of CO2 gas and nitrogen and oxygen are adjusted to ensure that the gas in gas mixing chamber 3 meets the gas requirements of the assimilation label.

[0043] In one embodiment, the assimilation chamber 4 of the present invention can be provided in several forms, all of which are connected to the gas mixing box 3 and the tail gas absorption device 5 through pipes, so as to meet the requirement of measuring the photosynthetic rate of different leaves at the same time and improve the measurement efficiency.

[0044] In one implementation, the pipe is a transparent PVC flexible tube, which connects various devices to form a gas flow path.

[0045] The technical solutions of the present invention will be described in detail below with reference to the embodiments, but they should not be construed as limiting the scope of protection of the present invention.

[0046] Example 1

[0047] A device for measuring the photosynthetic rate of plant leaves, the device comprising a PVC transparent flexible tube connected to... 13 The apparatus consists of a CO2 unit (1), a nitrogen-oxygen unit (2), a gas mixing chamber (3), and an assimilation chamber (4). The assimilation chamber (4) is made of high-transparency acrylic sheet, and each chamber measures 100cm long, 15cm wide, 5cm high, and 0.5cm thick. The assimilation chamber comprises an upper cover and a lower housing. The lower housing has a flange and rubber gasket for sealing with the upper cover. One end of the assimilation chamber is open for inserting the blade to be tested.

[0048] The gas connection sequence is as follows Figure 1 As shown. 13 CO2 unit 1 and nitrogen-oxygen unit 2 are connected to the gas mixing chamber 3 via a hose equipped with a flow meter 8. The gas mixing chamber 3 is connected to the assimilation chamber 4 via a hose. The other end of the assimilation chamber 4 is connected to a tail gas absorption device 5, which contains 0.1 mol·L⁻¹ gas. -1 NaOH solution. An LED light source 6 is installed directly above the assimilation chamber 4, and a fan 7 is installed at the internal air inlet.

[0049] Example 2

[0050] This example measures the photosynthetic rate of different parts of a leaf on a specific ear in potted maize at the R2 stage.

[0051] (1) According to Figure 1The apparatus shown is for measuring the photosynthetic rate of plant leaves. Four assimilating leaf chambers are prepared, each connected at one end to a mixed gas chamber and a tail gas absorption device via PVC transparent flexible tubing. A mixed gas containing 400 ppm is injected into the assimilating leaf chambers through a high-pressure gas cylinder. 13 CO2 and 80%-20% nitrogen and oxygen, 13 The volume ratio of CO2 to nitrogen and oxygen is 1:2500. The relative humidity (RH) of the mixed gas is 60-70%. The flow rate of the nitrogen and oxygen high-pressure cylinder (80% N2 and 20% O2) is adjusted to 40 L / min using a flow meter (YQB-731L-50; Provov). -1 ;pure 13 The flow rate of the CO2 high-pressure cylinder was adjusted to 16 mL / min using another flow meter (IKFD0-10SCCM; Beijing Aino). -1 Place an appropriate amount of damp paper towels in the gas mixing chamber to increase humidity. A fan with a wind speed of approximately 0.5 m / s is placed near the air inlet. -1 A mixing fan is used to ensure uniform gas composition within the blade chamber. 0.1 mol·L⁻¹ -1 NaOH solution filters out the airflow from the assimilation chamber to absorb the remaining airflow. 13 CO2 pollution can be prevented. Use air conditioning to keep the room temperature at around 29°C and use a humidifier to maintain the relative humidity at around 60%.

[0052] (2) Fully insert one ear-shaped leaf of the maize into the assimilating chamber. Seal the connection between the leaf and the assimilating chamber with foam sealant (GSLIM; Deli) to prevent air leakage. Place the chamber under light and open the top cover to induce photosynthesis for 30 minutes. The effective photosynthetic light flux density of the leaf is approximately 1300 μmol·m⁻¹. -2 ·s -1 The photosynthetic saturation point is close to that of corn leaves. The device has four assimilatory leaf chambers, allowing it to accommodate four plants simultaneously for photosynthetic induction.

[0053] (3) Cover the assimilation chamber with the top cover and seal the chamber, then introduce 400 ppm. 13 A mixture of CO2, 80% N2, and 20% O2, with a RH of 60-70%. Ventilation was stopped after 2 minutes.

[0054] (4) Cut the marked leaf into eight even sections from top to bottom. Cut approximately 4cm from each section. 2 Leaf tissue, avoiding the midrib. When measuring leaf tissue area (a leaf cm 2 After drying, the dry weight (DW, g) was measured. Leaf mass per unit area (LMA, g·cm³) was also measured. -2 ) equals DW divided by aleaf .

[0055] (5) Grind the dried leaf tissue into powder, measure the carbon content in the leaf dry matter using an elemental analyzer, and measure the carbon content in the leaf dry matter using a stable isotope ratio mass spectrometer. 13 The ratio of C to total C.

[0056] (6) Samples were also taken from the leaves of the maize ear that were not labeled with isotopes to determine the carbon content in different parts. 13 The ratio of C to total C was used as a control to calculate the local photosynthetic rate of the leaf.

[0057] Leaf local photosynthetic rate (A) 13C μmol·m -2 ·s -1 It is calculated using the following formula:

[0058]

[0059] In the formula, C% is the carbon content in dry matter (dimensionless); R label It is in the labeled sample 13 The ratio of C to total C (dimensionless); R ref In unlabeled samples 13 The ratio of C to total C (dimensionless); t is the assimilation labeling time (s); m c yes 13 The molecular weight of C atom (13 g·mol⁻¹) -1 ).

[0060] Figure 2 The results show the distribution of photosynthetic rate within the same maize leaf. The results indicate that the photosynthetic rate of maize leaves increases significantly from the base to the tip, with values ​​ranging from 13.8 to 19.3 μmol·m⁻¹. -2 ·s -1 .

[0061] Example 3

[0062] Similar to Example 3, except that an assimilating leaf chamber is used according to Figure 1 The connection shown is used to measure the photosynthetic rate of plant leaves. After stopping ventilation in step 3, the leaf is removed from the first assimilating chamber, the airway is switched to the next assimilating chamber, and the above operation is repeated for assimilation labeling.

[0063] Example 4

[0064] To verify the present invention 13 The reliability of the CO2 labeling method in the quantitative measurement of photosynthetic rate was tested using the following two methods, with the following operating procedures:

[0065] 1) Assimilate transport during assessment labeling

[0066] The assimilation marker site and surrounding areas were measured after 2 minutes. 13 The ratio of C to total C is used to assess assimilate transport, after labeling. 13 C will introduce measurement errors if it is transferred to other parts of the blade.

[0067] Ten potted maize plants were used: five were used to assess assimilate translocation at the leaf tip (70% of leaf length from the ligule), and the other five were used to assess assimilate translocation at the leaf base (30% of leaf length from the ligule). The assimilation labeling procedure was the same as in Example 2. Two minutes after labeling, leaf tissue samples were collected from the labeled area and at 0-2 cm and 2-4 cm above and below it. After drying, the samples were measured. 13 The ratio of C to total C. Before labeling, five control samples should be collected from both the leaf tip and base to determine the control. 13 The ratio of C to total C. Assimilation of marker sites. 13 C mass (M) label (g) and its proportion of the total (P) label , %) are calculated using the following formulas respectively:

[0068] M label =DW×C%×(R) label -R ref (2)

[0069]

[0070] In the formula, M -2 M -1 M1 and M2 represent leaf tissue assimilation at locations 2-4 cm below and 0-2 cm above the marked area, respectively. 13 C quality. M -2 M -1 M1 and M2 are also calculated based on formula (2).

[0071] See results Figure 3 When performing [a procedure] on the portion of the leaf extending 1 / 3 of the way from the ligule... 13 Two minutes after CO2 labeling, 99.3% of the newly assimilated cells... 13 C remained at the marked site, while less than 0.7% of the newly assimilated 13C was transported to adjacent leaf tissues. Similar results were observed when marking was performed at two-thirds of the leaf length from the ligule.

[0072] 2) Portable photosynthesis instrument and the present invention 13 Comparison of photosynthetic measurement results using CO2 labeling method

[0073] This comparison was conducted using 13 leaves from 5 potted corn plants. The plants were dehydrated to different levels of drought, and gas exchange in the mid-region of each leaf was measured using a portable photosynthesis system (Licor-6800; LI-COR, USA). The photon density was set to 1300 μmol·m⁻² in the leaf chamber of the portable photosynthesis system. -2 ·s -1 The air temperature, relative humidity, and CO2 concentration were set to 27℃, 60%, and 400ppm, respectively. The net photosynthetic rate (P0.05) measured by the portable photosynthesis meter was... n μmol·m -2 ·s -1 After the stomatal conductance stabilized, the results were recorded. Then, the gas supply channel of the portable photosynthesis system was immediately switched from ambient air to the pre-mixed gas mixture (400 ppm). 13 (CO2, 80%-20% nitrogen and oxygen) The temperature, light quantum density, and relative humidity of the leaf chamber in the portable photosynthesis apparatus remained constant. After 2.5 minutes of ventilation, leaf samples were cut from the chamber and their leaf mass per unit area (LMA), carbon content in dry matter, and other parameters were measured. 13 The ratio of C to total C, and the photosynthesis rate A obtained according to the calculation method in Example 2. 13C .

[0074] Using the Licor-6800 portable photosynthesis system and the present invention 13 The CO2 labeling method was used to compare and analyze the photosynthetic rate at the middle position of the same leaf. The results showed a strong correlation between the photosynthetic rate values ​​measured by the two methods. Figure 4 ;R 2 =0.95, P<0.0001), and the slope is close to 1.

[0075] The above experimental results show that the method for determining the photosynthetic rate of plant leaves in this invention is reliable.

[0076] The above description is only a preferred embodiment of the present invention. It should be noted that for those skilled in the art, several improvements and modifications can be made without departing from the principle of the present invention, and these improvements and modifications should also be considered within the scope of protection of the present invention.

Claims

1. A method for measuring photosynthetic rate of a plant leaf based on carbon 13 isotope labeling, characterized by, The plant leaves are photosynthetically induced with a light source, and a mixed gas containing 13 CO2 is used to assimilate mark the leaves after the photosynthetic induction; the leaf mass per unit area LMA, the carbon content in dry matter, and the ratio of C to total C in dry matter of the leaves after the assimilation marking are measured; and the leaf local photosynthetic rate is calculated according to the following formula: 13 A = (LMA - LMA0) / (t - t0) where C% is the carbon content in dry matter, R label is the ratio of C to total C in the labeled sample 13 R ref is the ratio of C to total C in the unlabeled sample 13 t is the assimilation labeling time, m c is 13 the molecular weight of the C atom.

2. The method of claim 1, wherein, The assimilation labeled leaves were divided into several leaf tissues, and the area a of the leaf tissues was measured leaf and the dry weight DW, and the leaf mass per area LMA was calculated.

3. The method of claim 1, wherein, Leaf tissues were oven-dried and ground into powder, and the carbon content in the dry matter of the leaves was measured using an elemental analyzer, and the ratio of C to total C in the dry matter of the leaves was measured using a stable isotope ratio mass spectrometer. 13 C to total C.

4. The method of claim 1, wherein, The mixed gas includes CO2 at a concentration of 400-410 ppm 13 CO2 and nitrogen-oxygen gas, the 13 The volume ratio of CO2 to nitrogen-oxygen gas is 1:2500, and the volume ratio of nitrogen to oxygen in the nitrogen-oxygen gas is 4:

1.

5. The method of claim 4, wherein, The flow rate of the nitrous gas is adjusted to be not less than 40 L·min -1 , the 13 The flow rate of the CO2 is adjusted according to the flow rate of the nitrous gas.

6. The method of claim 1, wherein, The photosynthesis induction time is 20-40 min, and the assimilation labeling time is 2-3 min.

7. A device for determining the photosynthetic rate of a plant leaf based on the method according to any one of claims 1 to 6, characterized in that It comprises a gas source, a gas mixing box and a assimilation leaf chamber 13 CO2 device and nitrogen oxygen device, the 13 CO2 device and nitrogen oxygen device are connected with the gas mixing box through pipelines provided with flow control meters, and the gas mixing box is connected with the assimilation leaf chamber through a pipeline; one end of the assimilation leaf chamber is connected with a tail gas absorption device; a light source is arranged above the assimilation leaf chamber, and a fan is arranged in the assimilation leaf chamber.

8. The apparatus of claim 7, wherein, The assimilation leaf chambers are made of transparent material, are provided with several chambers, and are connected with the gas mixing box and the tail gas absorption device through pipes.

9. The apparatus of claim 7, wherein, The assimilation leaf chamber is divided into an upper cover and a lower box body, the lower box body is provided with a flange and a gasket to seal the upper cover, and one end of the assimilation leaf chamber is an opening for placing the leaves to be tested.

10. The apparatus of claim 7, wherein, The tail gas absorption device is provided with NaOH solution.

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