A method for screening low-temperature-resistant garden chrysanthemum varieties in plateau areas

Abstract

The invention discloses a method for screening low-temperature resistant garden chrysanthemum varieties. The method can evaluate the low-temperature adaptability of garden chrysanthemums by measuring two root indexes and one photosynthetic pigment index of garden chrysanthemums, thereby screening low-temperature resistant garden chrysanthemum varieties. The method of the invention can identify low-temperature resistant garden chrysanthemum varieties by measuring only three indexes, which not only has low identification cost, but also is simple to operate and is very easy to promote.

Description

Technical Field

[0001] The invention relates to a method for screening garden chrysanthemums, in particular to a method for screening low-temperature-resistant garden chrysanthemum varieties in plateau areas, and belongs to the technical field of chrysanthemum variety screening. Background Art

[0002] Chrysanthemum (Chrysanthemum morifolium) is a perennial herbaceous plant of the genus Chrysanthemum in the family Asteraceae. Among them, garden chrysanthemum has the characteristics of short and compact plant shape, wide variety and strong ornamental value, making it a good ground cover material for urban greening. Asexual cuttings are used in production. Because of its fast growth and high reproduction coefficient, it occupies a very important position in the flower market.

[0003] In recent years, with the construction of the environment and the breeding of new varieties, many chrysanthemum varieties have been introduced in various regions. However, the temperature requirements for chrysanthemum regrowth are relatively strict. As one of the key factors affecting cuttings, too high or too low temperature is not conducive to the germination and growth of new roots. Even low temperature can cause slow rooting, poor development of the root system and cuttings in chrysanthemum cuttings (Tsipouridis et al., 2005; Li Xiaowei et al., 2014).

[0004] The Qinghai region where the inventor is located has also introduced a lot of chrysanthemum varieties. However, because Qinghai is located in the Qinghai-Tibet Plateau, the altitude is high and the temperature is low, which seriously affects the growth and development of cutting seedlings, greatly reducing the survival rate of cutting seedlings, resulting in long rooting time of cutting seedlings, poor root quality and poor growth and development of cuttings. Although most of the production has adopted solar greenhouses, due to the weak overall foundation, its illumination, temperature and controllability are poor, and the cost of heating system seedling cultivation is high, the spring low temperature has become a constraint factor for chrysanthemum facility cultivation in plateau areas, causing the time required for cuttings to take root to be longer than other areas. Therefore, when introducing chrysanthemum varieties in Qinghai, rooted seedlings are mostly introduced, and their high price and large volume seriously affect the application of chrysanthemums in Qinghai. There is no report on the research on the screening of low-temperature resistant varieties of garden chrysanthemums and the evaluation of adaptability, which further causes the blindness when introducing chrysanthemum varieties in Qinghai. Summary of the invention

[0005] In view of the above-mentioned defects of the prior art, the present invention aims to provide a method for screening low-temperature-resistant varieties of garden chrysanthemums that is simple and meets the actual production requirements.

[0006] The method for screening low-temperature-resistant garden chrysanthemum varieties in plateau areas of the present invention comprises the following steps:

[0007] (1) Cut garden chrysanthemum cuttings and plant them in plug trays. Set two temperature treatments for cuttings: the stress group was 15℃ / 15℃ day / night and the control group was 25℃ / 20℃;

[0008] (2) Determination of the total root surface area and number of root tips of garden chrysanthemum cuttings: During the growth period of garden chrysanthemums, the total root surface area and number of root tips were measured, respectively in cm 2 The sum root is the unit;

[0009] (3) Determination of the total chlorophyll content of garden chrysanthemum cuttings: Take a fresh sample of the garden chrysanthemum to be tested and extract chlorophyll with ethanol. Pour the chlorophyll extract into a cuvette, use the ethanol solution as a control and adjust to zero, and measure the absorbance at wavelengths of 665nm and 649nm; calculate the concentrations of chlorophyll a, chlorophyll b and total chlorophyll according to the formula to obtain the total chlorophyll content in the tissue; where: chlorophyll a concentration = 13.95A 665 -6.88A 649 ; Chlorophyll b concentration = 24.96A 649 -7.32A 665 ; Total chlorophyll concentration = chlorophyll a concentration + chlorophyll b concentration; Chloroplast pigment content = (pigment concentration * extract volume) / sample fresh weight;

[0010] (4) Calculate the low temperature resistance coefficients of the indicators measured in step (2) and step (3): the low temperature resistance coefficient of the total root surface area X1 = (measured value after stress / measured value of the control) * 100%; the low temperature resistance coefficient of the total chlorophyll content X2 = (measured value after stress / measured value of the control) * 100%; the low temperature resistance coefficient of the number of root tips X3 = (measured value after stress / measured value of the control) * 100%;

[0011] (5) Calculate the comprehensive evaluation value of low temperature resistance = 0.220 + 0.090X1 + 0.580X2 + 0.095X3; the higher the comprehensive evaluation value of low temperature resistance is, and when X1, X2 and X3 are not zero, the more low temperature resistant the variety is.

[0012] The step (1) of the present invention of cutting the garden chrysanthemum cuttings can be carried out according to conventional methods in the art. In a specific example, the top of the garden chrysanthemum with a length of 8 to 10 cm is cut as the cuttings, and the top 2 to 3 leaves are left, and the cuttings are inserted into the plug tray.

[0013] The growing period of the garden chrysanthemum in step (2) of the present invention refers to 26 to 28 days after cutting.

[0014] The total root surface area and the number of root tips of garden chrysanthemum cuttings in step (2) of the present invention can be determined according to a root scanning system commonly used in the art, such as the root phenotyping analysis system GXY-A, the root phenotyping analysis software Win-RHIZO and other root scanning analysis software.

[0015] The total chlorophyll content of garden chrysanthemum cuttings in step (3) of the present invention can be determined according to conventional methods in the art. In order to make the determination results more unified and accurate, the present application provides a specific determination method: take fresh samples of the garden chrysanthemum to be tested from the stress group and the control group, wash the surface dirt, cut them into pieces and mix them evenly, weigh 0.1g of the cut fresh samples, a total of 3 parts, and place them in 10mL centrifuge tubes respectively; add 5mL of 95% ethanol to each centrifuge tube to completely immerse the sample in the ethanol solution; after immersing in a dark place for 24 hours, pour the chlorophyll extract into a colorimetric dish, use 95% ethanol solution as a control and adjust to zero, and measure the absorbance at wavelengths of 665nm and 649nm.

[0016] The higher the comprehensive evaluation value of low temperature resistance in step (5) of the present invention, the more low temperature resistant the variety is. Generally, when the score is ≥0.85, it is a low temperature resistant garden chrysanthemum variety, that is, a variety that can grow by cutting at a temperature of 10°C to 25°C.

[0017] In some embodiments, the garden chrysanthemum described in the present invention is specifically Jinling Pink Magnolia, Jinling Red Lotus, Jinling Fireworks, Jinling Smile, Jinling Poinsettia or Jinling Lianyan.

[0018] The beneficial effects of the present invention compared to the prior art are as follows:

[0019] The present invention provides a simple and effective method for screening low-temperature resistant varieties of garden chrysanthemums in plateau areas. By measuring two root indexes and one photosynthetic pigment index of garden chrysanthemums, low-temperature adaptability of garden chrysanthemums can be evaluated, thereby screening low-temperature resistant varieties of garden chrysanthemums. The method of the present invention can identify low-temperature resistant varieties of garden chrysanthemums by measuring only three indexes, which not only has low identification cost, but also is simple to operate and is very easy to promote.

[0020] The method described in the present invention establishes a relatively reliable comprehensive evaluation method, which can effectively screen out varieties that are resistant to low temperatures during the cutting period, and is of great significance for the introduction and industry of garden chrysanthemums in plateau areas. DETAILED DESCRIPTION

[0021] The following examples are provided for better understanding of the present invention, but are not intended to limit the present invention. The experimental methods in the following examples are conventional methods unless otherwise specified. The experimental materials used in the following examples are purchased from conventional biochemical reagent stores unless otherwise specified.

[0022] Unless otherwise specified, the experimental materials of the following embodiments are as follows: 6 garden chrysanthemum varieties introduced by Nanjing Agricultural University are used as test materials, namely Jinling Lianyan (pink purple), Jinling Xiaoyan (purple), Jinling Poinsettia (red), Jinling Lihua (multicolor), Jinling Fenyulan (pink purple), and Jinling Honghe (red). The flowering period is from mid-August to late September, which overlaps with the tourist season in Qinghai. The cutting seedlings of the 6 garden chrysanthemum varieties were planted in the greenhouse of the experimental site of the Horticulture Institute of Qinghai University, using random blocks, a plant spacing of 40×40 cm, and the same conventional water and fertilizer management methods.

[0023] Example 1

[0024] The present invention screens low-temperature-resistant varieties of 6 garden chrysanthemum varieties introduced from Nanjing Agricultural University. Specific test materials are shown in Table 1.

[0025] Table 1 Information on garden chrysanthemum varieties

[0026]

[0027] The test materials were planted in the experimental field of the Horticulture Institute of Qinghai Academy of Agricultural and Forestry Sciences in May 2021, with a planting spacing of 40 cm and a row spacing of 40 cm, randomly arranged, and the same conventional water and fertilizer management methods were adopted. The test selected healthy and pest-free mother plants and cut 8-10 cm of the top as cuttings, leaving 2-3 leaves on the upper part, and inserted them into 50-hole trays. There were 2 temperature treatments: day / night 15℃ / 15℃ and 25℃ / 20℃. The materials were placed under different temperature conditions, with 25℃ as the control, and 50 cuttings were used for each treatment.

[0028] The selection of low temperature resistant garden chrysanthemum varieties includes the following steps:

[0029] (1) Determination of the total root surface area and root tip number of garden chrysanthemum:

[0030] During the growth period of garden chrysanthemums, the total root surface area and the number of root tips were measured by the root phenotyping system GXY-A. The total root surface area is expressed in cm 2 The unit is root, and the number of root tips is root.

[0031] (2) Determination of the total chlorophyll content of garden chrysanthemums: During the growth period of garden chrysanthemums, the total chlorophyll content in their leaves was determined: the tops of each variety treated at different temperatures were selected, the dirt on the surface of the tops was washed, and after being cut into pieces and mixed, 0.1g of the cut fresh samples were weighed, a total of 3 portions, and placed in 10mL centrifuge tubes. 5mL of 95% ethanol was added to each centrifuge tube to completely immerse the sample in the ethanol solution. After soaking in the dark for 24 hours, the chlorophyll extract was poured into a cuvette, and a 95% ethanol solution was used as a control and zeroed, and the absorbance at wavelengths of 665nm and 649nm was measured. The concentrations of chlorophyll a, chlorophyll b and total chlorophyll were calculated according to the formula to obtain the pigment content in the tissue. (Chlorophyll a concentration = 13.95A 665 -6.88A 649 ; Chlorophyll b concentration = 24.96A 649 -7.32A 665 ; Total chlorophyll concentration = chlorophyll a concentration + chlorophyll b concentration; Chloroplast pigment content = (pigment concentration * extract volume) / sample fresh weight)

[0032] (3) Calculate the low temperature resistance coefficient α of the indicators measured in step (1) and step (2): the low temperature resistance coefficient of the total root surface area X1 = (measured value after stress / measured value of the control) * 100%; the low temperature resistance coefficient of the total chlorophyll content X2 = (measured value after stress / measured value of the control) * 100%; the low temperature resistance coefficient of the number of root tips X3 = (measured value after stress / measured value of the control) * 100%.

[0033] (4) Evaluation of low temperature adaptability of garden chrysanthemum: The low temperature adaptability of garden chrysanthemum was evaluated by establishing an optimal regression equation with the low temperature resistance coefficient of each individual index of garden chrysanthemum as the independent variable, and the calculation was performed according to the following formula:

[0034] Comprehensive evaluation value of low temperature resistance = 0.220 + 0.090X1 + 0.580X2 + 0.095X3.

[0035] The screening results of the low-temperature resistance traits of garden chrysanthemums are shown in Table 2. The higher the comprehensive evaluation value of low-temperature resistance (D value), the stronger the low-temperature resistance of the material. Among them, 'Jinling Lianyan' has the largest D value, indicating that this variety has the strongest low-temperature resistance. It has been verified that the score ranking is consistent with the actual low-temperature resistance (Sun Donglei et al., 2021). Among them, 'Jinling Lianyan' and 'Jinling Honghe' are of the same type, which are highly low-temperature resistant types and are varieties that can grow by cuttings at temperatures of 10℃~25℃.

[0036] Table 2 Comprehensive evaluation results of low temperature resistance of garden chrysanthemum

[0037]

[0038]

[0039] The inventors of the present application also studied the morphological differences of six garden chrysanthemums at different temperatures, further verifying the reliability of the scheme described in the present application.

[0040] The six rural chrysanthemum varieties in Example 1 were selected from healthy and pest-free mother plants and 8-10 cm top tips were cut as cuttings, leaving 2-3 leaves on the upper part, and inserted into 50-hole plug trays, with a total of 4 temperature treatments: 5°C, 10°C, 15°C, and 25°C. The materials were placed under different temperature conditions, 50 cuttings were taken for each treatment, and samples were taken 28 days after cutting to observe rooting and development.

[0041] The results showed that the roots of all varieties grew normally under normal culture (25°C); at 15°C, all varieties except 'Jinling Lihua' took root; at 10°C, only 'Jinling Honghe' and 'Jinling Lianyan' had roots, and the other varieties did not take root; this is also consistent with the results of the screening method described in Example 1, further proving the reliability of the method described in the present application.

Claims

1. A method for screening low-temperature resistant garden chrysanthemum varieties in plateau areas, characterized in that: The following steps are involved: (1) Cut garden chrysanthemum cuttings and plant them in plug trays. Set two temperature treatments for cuttings: the stress group was 15℃ / 15℃ day / night and the control group was 25℃ / 20℃; (2) Determination of the total root surface area and number of root tips of garden chrysanthemum cuttings: During the growth period of garden chrysanthemums, the total root surface area and number of root tips were measured, respectively in cm 2 The sum root is the unit; (3) Determination of the total chlorophyll content of garden chrysanthemum cuttings: Take a fresh sample of the garden chrysanthemum to be tested and extract chlorophyll with ethanol. Pour the chlorophyll extract into a cuvette, use the ethanol solution as a control and adjust to zero, and measure the absorbance at wavelengths of 665nm and 649nm; calculate the concentrations of chlorophyll a, chlorophyll b and total chlorophyll according to the formula to obtain the total chlorophyll content in the tissue; where: chlorophyll a concentration = 13.95A 665 -6.88A 649 ; Chlorophyll b concentration = 24.96A 649 -7.32A 665 ; Total chlorophyll concentration = chlorophyll a concentration + chlorophyll b concentration; Chloroplast pigment content = (pigment concentration * extract volume) / sample fresh weight; (4) Calculate the low temperature resistance coefficients of the indicators measured in step (2) and step (3): the low temperature resistance coefficient of the total root surface area X1 = (the measured value of the total root surface after stress / the measured value of the total root surface of the control) * 100%; the low temperature resistance coefficient of the total chlorophyll content X2 = (the measured value of the total chlorophyll after stress / the measured value of the total chlorophyll of the control) * 100%; the low temperature resistance coefficient of the number of root tips X3 = (the measured value of the number of root tips after stress / the measured value of the number of root tips of the control) * 100%; (5) Calculate the comprehensive evaluation value of low temperature resistance = 0.220 + 0.090X1 + 0.580X2 + 0.095X3; the higher the comprehensive evaluation value of low temperature resistance is, and when X1, X2 and X3 are not zero, the more low temperature resistant the variety is.

2. The method for screening low-temperature resistant garden chrysanthemum varieties according to claim 1, characterized in that: Step (1) Cut the top of a garden chrysanthemum with a length of 8 to 10 cm as a cutting, leaving 2 to 3 leaves on the top, and plant them in a plug tray.

3. The method for screening low-temperature resistant garden chrysanthemum varieties according to claim 1, characterized in that: The garden chrysanthemum growth period in step (2) refers to 26 to 28 days after cutting.

4. The method for screening low-temperature resistant garden chrysanthemum varieties according to claim 1, characterized in that: Step (2) The total root surface area and the number of root tips of garden chrysanthemum cuttings are determined using a root scanning system.

5. The method for screening low-temperature resistant garden chrysanthemum varieties according to claim 4, characterized in that: The root scanning system is the root phenotyping analysis system GXY-A or the root phenotyping analysis software Win-RHIZO.

6. The method for screening low-temperature resistant garden chrysanthemum varieties according to claim 1, characterized in that: The method for determining the total chlorophyll content of garden chrysanthemum cuttings in step (3) is specifically as follows: fresh samples of the garden chrysanthemum to be tested from the stress group and the control group are taken respectively, the surface dirt is washed off, the samples are chopped and mixed evenly, 0.1 g of the chopped fresh samples are weighed for each, a total of 3 portions, and the samples are placed in 10 mL centrifuge tubes respectively; 5 mL of 95% ethanol is added to each centrifuge tube to completely immerse the samples in the ethanol solution; after immersing in a dark place for 24 hours, the chlorophyll extract is poured into a cuvette, 95% ethanol solution is used as a control and zeroed, and the absorbance at wavelengths of 665 nm and 649 nm is measured.

7. The method for screening low-temperature resistant garden chrysanthemum varieties according to claim 1, characterized in that: When the score is ≥0.85, it is a low-temperature-resistant garden chrysanthemum variety.

8. The method for screening low-temperature resistant garden chrysanthemum varieties according to claim 7, characterized in that: Low-temperature-resistant garden chrysanthemum varieties refer to varieties that can grow by cuttings at temperatures between 10℃ and 25℃.

9. The method for screening low-temperature resistant garden chrysanthemum varieties according to claim 1, characterized in that: The garden chrysanthemum is specifically Jinling pink magnolia, Jinling red lotus, Jinling fireworks, Jinling smile, Jinling poinsettia or Jinling Lianyan.

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