Method for increasing fruit setting number of macadamia nuts

Abstract

The invention belongs to the technical field of agricultural planting, and particularly relates to a method for increasing the fruit setting number of macadamia nuts, which comprises the following steps: separating selected macadamia nut female parent and male parent plants by using an insect-proof net before flowering so as to avoid the influence of insects; after the female parents enter a full-bloom stage, selecting sunny weather with the daytime air temperature of 20-30 DEG C and the wind speed of less than 1m / s, cleaning original pollen on styluses of the female parents by using a sterile writing brush, brushing the collected pollen of the male parents on the styluses by using a brush pen for pollination, keeping isolation for 14 days after pollination, and then removing the insect-proof net. The high-affinity pollination combination is screened by combining germplasm such as Guijin No.1, Guijin A4 and Guijin YM26 with pollen viability. The optimal pollination combination is screened by evaluating the affinity of 16 hybrid combinations of 10 germplasms of macadamia nuts. Results show that when A4 is used as a male parent and Guihot No.1 is used as a female parent, the affinity of cross pollination is the highest, the success rate of cross pollination is remarkably improved, the average fruit setting number is remarkably increased, and effective technical support is provided for high-yield cultivation of macadamia nuts.

Description

Technical Field

[0001] The present invention belongs to the technical field of agricultural planting, and specifically relates to a method for increasing the yield of macadamia nuts ( Macadamia integrifolia ) This method is particularly suitable for variety improvement and yield increase in commercial cultivation. Background Art

[0002] Macadamia nuts ( Macadamiaintegrifolia Maiden & Betche (Maiden & Betche) is a Proteaceae plant native to the tropical rainforests of Australia. It was selected and cultivated for its high nutritional and commercial value. It was introduced and cultivated in China in the 1970s and 1980s. Currently, the main producing areas are Guangxi, Yunnan, Guizhou, and Guangdong. Key varieties include OC, Guire No. 1, A4, and Changning No. 1. However, commercial cultivation of this crop faces significant technical bottlenecks.

[0003] Under open pollination conditions, macadamia fruit set has a low fruit set rate of only 0.1-0.3%. One reason for this low fruit set rate is a certain degree of self-incompatibility. Hybridization can significantly increase fruit set and yield. Macadamia nut has varying pollen affinity for different varieties, and selecting suitable pollinating tree varieties can increase fruit set and yield. Ito et al. found that macadamia nut production in mixed plantings averaged approximately 16 kg higher per tree than in single plantings. Therefore, research into selecting suitable macadamia and sire varieties is crucial.

[0004] The pollinated but unfertilized florets of macadamia nuts usually fall off naturally within 1-2 weeks after flowering. Fruit drop at this stage is dominated by pollination and fertilization, and the first fruit set number should be counted after 2 weeks.

[0005] Common methods for measuring pollen viability include staining and germination. The staining method is quick and easy, but can result in false positives or negatives. The germination method is free of false positives, but germination rates vary under different germination conditions. Macadamia pollen viability is related to factors such as opening time and germplasm. Under open pollination conditions, pollen viability is positively correlated with the number of initial fruits set in the maternal plant. However, the relationship between pollen viability and maternal compatibility in different macadamia germplasms remains unclear. Summary of the Invention

[0006] The present invention addresses technical problems in the prior art, such as the low natural pollination fruit setting rate (less than 0.3%) of macadamia nuts and the lack of scientific basis for variety combination screening. The invention provides an artificially controlled hybrid pollination method based on germplasm optimization, which can increase the number of fruits set to an average of more than 21.57 per sequence.

[0007] The technical solution of the present invention is achieved by the following steps: A method for increasing the number of macadamia nut fruit setting, comprising the following steps: (1) Isolation Before flowering, isolate both the macadamia macadamia plant and the macadamia macadamia plant with insect-proof nets to avoid insects. (2) Emasculation When the female plant enters the flowering period, choose a sunny day with a daytime temperature of 20-30℃ and a wind speed of less than 1m / s, and use a sterile brush to clean the pollen on the female plant's style; (3) Artificial pollination Use a sterilized brush to evenly apply the pre-collected male parent pollen to the female parent's stigma; (4) Post-processing Maintain insect-proof net isolation for 14 days after pollination.

[0008] Furthermore, the female parent is Macadamia nut Guire No. 1 ( Macadamia integrifolia 'Guire 1').

[0009] Furthermore, the male parent is Macadamia A4 ( Macadamia integrifolia 'A4').

[0010] Furthermore, the macadamia nut female parent and male parent are adult fruit trees with a tree age of 5-7 years and strong growth.

[0011] Furthermore, the insect-proof net is 80-100 mesh, made of polyethylene, and has a light transmittance of ≥85%.

[0012] Furthermore, the peak flowering period is when the number of flowers blooming on a single plant reaches 75%-85% of the total number of flowers.

[0013] Furthermore, the male parent pollen is fresh pollen collected within 12 hours after flowering, and the pollen germination rate is ≥28.57%.

[0014] Compared with the prior art, the present invention has the following beneficial effects: This study used germplasms such as Guire No. 1, A4, and YM26, combined with pollen viability, to screen for suitable pollination combinations. The optimal pollination combination was selected by evaluating the affinity of 16 hybrid combinations from 10 macadamia germplasms. Results showed that hybrid pollination with A4 as the male parent and Guire No. 1 as the female parent had the highest affinity and the highest average initial fruit set, significantly improving the hybrid pollination success rate and increasing the average fruit set, providing effective technical support for high-yield macadamia cultivation. BRIEF DESCRIPTION OF THE DRAWINGS

[0015] Figure 1 This is a composite graph of the average number of primary fruits per sequence in the hybrid pollination experiment; Figure 2 This is a composite diagram of the macadamia nut pollen viability staining method; Figure 3 This is a bar chart of macadamia nut pollen germination rate; Figure 4 This is a combined graph of the correlation analysis between pollen germination rate and average first fruit set number.

[0016] Description of main reference numerals: Figure 1 In the figure, A: Cloud-rain plot of the number of first fruits in the hybrid pollination experiment (violin plot + scatter plot + box plot), where × represents an outlier; B: Cloud-rain plot of the number of first fruits in each hybrid combination (violin plot + scatter plot + box plot), where × represents an outlier; C: Histogram of the average number of first fruits in each hybrid combination, using Duncan's new multiple range method for multiple comparisons. Different letters above the bars indicate significant differences. Figure 2 Figure A: Staining with potassium iodide starch indicator (0.5%); Figure B: Staining with TTC staining solution (2%); Figure C: Staining with acetic acid carmine staining solution; Figure D: Histogram of pollen staining rates using acetic acid carmine staining. Multiple comparisons were performed using Duncan's new multiple range method. Different letters above the bars indicate significant differences. Figure 3 In the data, the multiple comparison method was Duncan's new multiple range method. Different letters above the columns indicate significant differences. Figure 4 Middle, A: Linear regression diagram of pollen germination rate and average number of first fruit set, R is the spearman rank correlation coefficient, p ≤ 0.05 is statistically significant; B: Double Y bar graph of pollen germination rate and average number of first fruit set when Guire No. 1 is the female parent; C: Double Y bar graph of pollen germination rate and average number of first fruit set when YM26 is the female parent. DETAILED DESCRIPTION

[0017] The following is a clear and complete description of the technical solution of the present invention. Obviously, the embodiments described are part of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without making any creative efforts are within the scope of protection of the present invention.

[0018] 1. Materials and Methods 1.1 Test materials Five accessions of maternal plants and ten accessions of paternal plants were selected, and a total of 16 hybrid combinations were designed, as detailed in Table 1. The experimental plants were planted in the tropical and subtropical germplasm resource nursery of the Guangxi Institute of South Subtropical Agricultural Sciences, with a row-to-plant spacing of 5 × 5 m, and were 5 years old. They were cultivated under the same water and fertilizer management.

[0019] Table 1 Hybridization test combinations

[0020] 1.2 Test methods 1.2.1 Cross-pollination Artificial hybridization pollination method: Before flowering, use insect-proof nets to isolate the macadamia nut female and male plants at the same time to avoid insects; (2) Emasculation: When the female plant enters the peak flowering period, choose a sunny day with a daytime temperature of 20-30℃ and a wind speed of less than 1m / s, and use a sterile brush to clean the pollen on the female plant's style; (3) Artificial pollination: Use a sterilized brush to evenly apply the pre-collected male plant pollen to the female plant's stigma; (4) Post-processing: Maintain the insect-proof net isolation for 14 days after pollination.

[0021] Average number of first fruit set = total number of fruit set / total number of inflorescences 1.2.2 Pollen staining Potassium iodide starch indicator (0.5%), TTC staining liquid (0.2%) and acetic acid carmine staining liquid were dropped onto different concave slides respectively, and the pollen to be measured was shaken off and left to stand for 10 minutes before being observed under a 100x microscope.

[0022] 1.2.3 In vitro pollen germination A liquid in vitro germination method (10% sucrose, 0.05% boric acid, and 0.05% calcium chloride) was used. A sucrose solution was dropped onto a concave glass slide. The stigma containing pollen was gently brought into contact with the solution using tweezers and gently agitated to evenly distribute the pollen within the droplet. The concave glass slide was placed in a Petri dish lined with moistened filter paper and covered. After incubation at 24°C for 6 hours, the pollen was examined under a microscope at 100x magnification. Pollen tubes were considered germinated when their length was greater than twice their diameter. Germinated pollen tubes were counted and the germination rate was calculated.

[0023] Pollen germination rate = number of pollen germination / number of pollen × 100% 2. Test results 2.1 Artificial hybridization pollination experiment See the results Figure 1 .

[0024] Depend on Figure 1 It can be seen that the number of first fruits set by artificial hybridization varies greatly among different pollination combinations. The first fruits in this experiment were thicker, indicating that the data distribution was more concentrated, with a median of 10, a range of 1-35, and a mode of 8 ( Figure 1 A). The average number of first fruit set after artificial hybridization pollination ranged from 6.5 to 21.6. The violin parent of 791 (♂) ⓧ A4 (♀) was the thickest and had the most concentrated data distribution, while the opposite was true for A4 (♂) ⓧ Guire 1 (♀). The median of A4 (♂) ⓧ Guire 1 (♀) was higher than that of other combinations, and the fruit set was generally higher, while the opposite was true for 791 (♂) ⓧ A4 (♀). The medians of the combinations with YM26 as the female parent were similar, but the distribution widths varied significantly ( Figure 1.B). A4 (♂) ⓧ Guire 1 (♀) had the highest average initial fruit set number, while 791 (♂) ⓧ A4 (♀) had the lowest average initial fruit set number, representing the best and worst hybrid combinations in this experiment, respectively. The average initial fruit set number of hybrid combinations with Guire 1 as the female parent was higher, while the average initial fruit set number of hybrid combinations with YM26 as the female parent was less different, indicating that YM26 is less selective in pollination trees than other germplasms. When Guire 1 was the female parent, the average initial fruit set number of A4 as the male parent was significantly higher than that of other germplasms, while the average initial fruit set number of Longbian 9 and 791 as male parents was similar. When YM26 was the female parent, the average initial fruit set number of A4 as the male parent was also significantly higher than that of other germplasms, indicating that A4 is suitable as a pollination germplasm for YM26 and Guire 1.

[0025] 2.2 Pollen viability of macadamia germplasm See the results Figure 2 and Figure 3 .

[0026] Depend on Figure 2 It can be seen that this experiment used three pollen staining methods: potassium iodide starch indicator (0.5%) staining, TTC staining solution (2%) staining, and acetic acid carmine staining solution staining. Potassium iodide starch indicator (0.5%) and TTC staining solution (2%) could not change the color of macadamia nut pollen of all germplasms, while acetic acid carmine had good staining properties for macadamia nut pollen ( Figure 2 A- Figure 2 C). In acetic acid carmine staining, the staining rate was as high as 83.93%, with A4 having the lowest staining rate and JW having the highest staining rate of 99.32%. The staining rates of YM26 and Guire No. 1 were significantly higher than those of A4, A38, and FS2. There was no difference in the staining rates between FS3, Guire No. 1, JW, YM26, and Longbian No. 9. There was no difference in the staining rates between FS3, Guire No. 1, and 791. There was no difference in the staining rates between A4, A38, and FS2. ( Figure 2 D).

[0027] Depend on Figure 3 Significant differences in pollen germination rates were observed among different macadamia accessions. Among the nine accessions, Guire 1 had the highest germination rate, at 71.94%, followed by Longbian 9 at 65.06% and YM26 at 59.52%. FS2 had the lowest germination rate, at 17.14%. The pollen germination rates of Guire 1, Longbian 9, YM26, and 791 were significantly higher than those of the other accessions. There were no significant differences between Guire 1, Longbian 9, and YM26, nor between FS3, A4, A38, and JW. The patterns of pollen germination rates across the various accessions were similar to those observed with acetic carmine staining.

[0028] 2.3 Correlation analysis between pollen germination rate and average number of first fruit set To explore the effect of pollen viability on artificial hybridization pollination, this study analyzed the correlation between pollen germination rate and the average number of first fruits per sequence. Figure 4 .

[0029] Depend on Figure 4 It can be seen that the Spearman rank correlation coefficient is 0.09 and the P value is 0.77, indicating that there is a weak positive correlation between the pollen germination rate of macadamia nuts and the average number of first fruit set after artificial pollination, but there is insufficient evidence to reject the null hypothesis ( Figure 4 A).

[0030] When YM26 and Guire 1 were used as female parents, the results showed that the male parent pollen germination rate of the combination with a high average number of first fruit set in artificial hybridization was not necessarily high. For example, the germination rate of A4 pollen was only 28.57%, but the average number of first fruit set was as high as 21.57 when pollinated with Guire 1, and 12.95 when pollinated with YM26. Both were the best pollination varieties for Guire 1 and YM26 ( Figure 4 B. Figure 4 C). Correlation analysis between macadamia nut pollen germination rate and average initial fruit set number, as well as double-Y-axis bar chart analysis, showed that the correlation between pollen germination rate and average initial fruit set was low under artificial pollination conditions.

Claims

1. A method for increasing the number of macadamia nuts, characterized in that: The following steps are involved: (1) Isolation Before flowering, isolate both the macadamia macadamia plant and the macadamia macadamia plant with insect-proof nets to avoid insects. (2) Emasculation When the female plant enters the flowering period, choose a sunny day with a daytime temperature of 20-30℃ and a wind speed of less than 1m / s, and use a sterile brush to clean the pollen on the female plant's style; (3) Artificial pollination Use a sterilized brush to evenly apply the pre-collected male parent pollen to the female parent's stigma; (4) Post-processing Maintain insect-proof net isolation for 14 days after pollination.

2. The method according to claim 1, characterized in that The female parent is Macadamia Guire No.

1.

3. The method according to claim 1, characterized in that The male parent is Macadamia A4.

4. The method according to claim 1, wherein The female parent and the male parent are 5-7 years old, healthy and mature fruit trees.

5. The method according to claim 1, wherein The insect-proof net is 80-100 mesh, made of polyethylene, and has a light transmittance of ≥85%.

6. The method according to claim 1, characterized in that The peak flowering period in step (2) is when the number of flowers blooming on a single plant reaches 75-85% of the total number of flowers.

7. The method according to claim 1, characterized in that The male parent pollen in step (3) is fresh pollen collected within 12 hours after flowering, and the pollen germination rate is ≥28.57%.

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