Planting method suitable for popularization of camelina sativa in saline-alkali soil

Abstract

The invention belongs to the technical field of camelina sativa planting, and particularly relates to a camelina sativa planting method suitable for being popularized in saline-alkali soil. The invention provides a planting method suitable for popularization of camelina sativa in saline-alkali soil. The planting method comprises the following steps: preparation before camelina sativa planting; seed variety selection; seed treatment: coating the camelina sativa seeds with a saline-alkaline tolerant rhizobium inoculant or a humate solution to obtain coated seeds; sowing the seeds; after sowing, cultivating to obtain camelina sativa plants. According to the planting method, the salt content of the surface layer of the soil can be remarkably reduced, the pH value of the soil is adjusted to be in the suitable range of 6.5-7.5, meanwhile, the germination rate, the seedling rate and the yield of the camelina sativa are increased, operation is easy and convenient, the cost is controllable, and the planting method is suitable for large-scale popularization in moderate and light saline-alkali areas and has wide application prospects. And a feasible technical scheme is provided for efficient utilization of saline-alkali land resources and regional expansion of the camelina sativa industry.

Description

Technical Field

[0001] This invention belongs to the field of flaxseed cultivation technology, specifically relating to a cultivation method suitable for promoting flaxseed cultivation in saline-alkali land. Background Technology

[0002] Flaxseed is an annual herbaceous plant belonging to the genus *Capsella* in the family Brassicaceae. Flaxseed is drought- and cold-resistant, has strong lodging resistance, a short growing season, and a high oil content. A prominent advantage of flaxseed oil is its high content of various beneficial unsaturated fatty acids, with alpha-linolenic acid (ω-3 fatty acid) accounting for 28%-43% of the total fatty acid content.

[0003] The current traditional method for cultivating flaxseed in saline-alkali land involves sowing seeds of salt-tolerant or conventional varieties in the soil. However, the high salt and alkalinity of the soil hinders seed imbibition, resulting in low germination rates, stunted growth, and low yields. Therefore, new cultivation strategies are needed to address these issues. Summary of the Invention

[0004] To address the problems of low seed germination rates, stunted growth, and low yields in existing traditional methods of cultivating flaxseed in saline-alkali land, this invention aims to provide a cultivation method suitable for promoting flaxseed in saline-alkali land. To achieve the above objective, this invention adopts the following technical solution.

[0005] This invention provides a method for promoting the cultivation of flax in saline-alkali land, comprising the following steps: Preparation before planting flaxseed: Before sowing, apply well-rotted organic fertilizer to the saline-alkali land to improve the soil properties.

[0006] Seed variety selection: Select salt-tolerant varieties of flax seeds.

[0007] Seed treatment: Flax seeds were coated with a coating solution to obtain coated seeds; the coating solution was a salt-tolerant functional bacterial solution or a humate solution, wherein the effective components of the salt-tolerant functional bacterial solution were Pseudomonas PfP-1 and Bacillus amyloliquefaciens SQR9.

[0008] Sowing: Sow the coated seeds from late March to early April each year when the soil temperature is ≥5℃.

[0009] After sowing, the plants are cultivated to obtain flax seedlings.

[0010] The planting method for promoting *Capsella bursa-pastoris* in saline-alkali land provided by this invention first involves applying well-rotted organic fertilizer to the saline-alkali land to be planted before sowing to improve the soil properties; then, selecting salt-tolerant *Capsella bursa-pastoris* seeds; next, coating the *Capsella bursa-pastoris* seeds with a coating solution to obtain coated seeds; the coating solution is a salt-tolerant functional bacterial solution or a humic acid solution, wherein the effective components of the salt-tolerant functional bacterial solution are *Pseudomonas aeruginosa* PfP-1 and *Bacillus amyloliquefaciens* SQR9; the coated seeds are sown from late March to early April each year when the soil temperature is ≥5℃; finally, the *Capsella bursa-pastoris* is harvested from mid-July to early August each year. The planting method for promoting flaxseed in saline-alkali land provided by the present invention, through the coordinated operation of the above steps, achieves efficient planting of flaxseed in saline-alkali land, and solves the problems of low seed germination rate, hindered growth and low yield in the existing traditional planting methods for flaxseed in saline-alkali land.

[0011] Firstly, applying a sufficient amount of well-rotted organic fertilizer before sowing can significantly reduce soil pH, salt concentration, and ESP (exchangeable sodium percentage), while improving soil aggregate structure and mitigating the physical-chemical resistance of salt to seed imbibition. Furthermore, selecting truly salt-tolerant varieties, whose radicle cells have enhanced osmotic regulation capabilities and sodium content... + / / K + It exhibits superior selectivity and antioxidant enzyme activity compared to conventional varieties, maintaining a germination rate of ≥85% even at higher salt concentrations. Simultaneously, the combination of salt-tolerant rhizobium inoculant and humic acid coating creates a "biochemical buffer zone" around the seed—the rhizobia reduce rhizosphere sodium levels through nitrogen fixation and extracellular polysaccharide secretion. + Concentration; humic acid through chelation of Na + Ca 2+ It replaces and stimulates root auxin synthesis, further alleviating salt stress. The three factors work synergistically to increase germination rate from below 60% to over 80%. Secondly, well-rotted organic fertilizer and humic acid continuously decompose and release slow-release NPK and trace elements, avoiding nutrient imbalances or secondary salt spikes caused by traditional "one-shot" fertilization. Humic acid increases soil CEC and reduces Na+. + For K + Ca 2+ The competitive inhibition of salt damage maintains cellular ion homeostasis and significantly alleviates growth inhibition caused by salt stress. Finally, humic acid promotes a simultaneous increase in silique number and thousand-seed weight, while maintaining the oil content of salt-tolerant varieties, ultimately increasing yield from the traditional 50-70 kg / mu to 90-110 kg / mu. Therefore, the planting method for promoting *Capsella bursa-pastoris* in saline-alkali land provided by this invention systematically weakens the adverse effects of salt-alkali stress on seed germination, seedling growth, and later grain filling through a three-pronged strategy of "soil improvement - salt-tolerant varieties - seed coating," thereby significantly improving the germination rate, growth vigor, and yield of *Capsella bursa-pastoris* in saline-alkali land.

[0012] To address the problems of high salinity, pH imbalance, and insufficient fertility in saline-alkali soils, the following measures are taken in combination: increasing the application of well-rotted organic fertilizer to improve the saline-alkali soil, selecting salt-tolerant varieties, treating seeds with humic acid coating, and employing scientific sowing techniques.

[0013] Furthermore, the salt- and alkali-tolerant functional bacterial solution is prepared by uniformly mixing the bacterial solution of *Pseudomonas* PfP-1 and the bacterial solution of *Bacillus amyloliquefaciens* SQR9 at a volume ratio of 1:0.8~1.2; the OD of the bacterial solution of *Pseudomonas* PfP-1... 600 The OD value of the bacterial culture of *Bacillus amyloliquefaciens* SQR9 was 1.4~1.6. 600 It is 1.4~1.6.

[0014] Furthermore, the coating process is as follows: The flax seeds are mixed with the coating solution at a mass ratio of 9 to 11:1, stirred to ensure that the flax seeds are evenly coated with the coating solution, and then dried to obtain the coated seeds.

[0015] Furthermore, the drying process is carried out by air drying or low-temperature air drying. Low temperature refers to a temperature of 15℃ to 25℃.

[0016] Furthermore, the amount of the decomposed organic fertilizer used is 800 kg / mu to 1000 kg / mu.

[0017] Furthermore, the flaxseeds are salt-tolerant varieties with an oil content of 35% to 42%.

[0018] Furthermore, the varieties of the flaxseed include "SNC104" or "SC-N1".

[0019] Furthermore, the sowing method is row sowing, with a row spacing of 24cm~26cm, a plant spacing of 3cm~5cm, and a sowing depth of 1cm~2cm.

[0020] Furthermore, the sowing rate is 1.5 kg / mu to 1.7 kg / mu.

[0021] The present invention also provides a flaxseed, which is obtained by the planting method described above.

[0022] Compared with the prior art, the present invention has the following beneficial effects: 1. This invention provides a method for promoting the cultivation of *Capsella bursa-pastoris* in saline-alkali land. The method involves first applying well-rotted organic fertilizer to the saline-alkali land before sowing to improve soil properties; then selecting salt-tolerant *Capsella bursa-pastoris* seeds; next, coating the seeds with a salt-tolerant rhizobium agent, humic acid, or humate to increase germination rate; sowing the coated seeds from late March to early April each year, when the soil temperature is ≥5℃; and finally, harvesting the *Capsella bursa-pastoris* from mid-July to early August each year. This method, through the coordinated steps described above, achieves efficient cultivation of *Capsella bursa-pastoris* in saline-alkali land, solving the problems of low seed germination rate, stunted growth, and low yield associated with existing traditional *Capsella bursa-pastoris* cultivation methods in saline-alkali land.

[0023] Firstly, applying a sufficient amount of well-rotted organic fertilizer before sowing can significantly reduce soil pH, salt concentration, and ESP (exchangeable sodium percentage), while improving soil aggregate structure and mitigating the physical-chemical resistance of salt to seed imbibition. Furthermore, selecting truly salt-tolerant varieties, whose radicle cells have enhanced osmotic regulation capabilities and sodium content... + / / K + It exhibits superior selectivity and antioxidant enzyme activity compared to conventional varieties, maintaining a germination rate of ≥85% even at higher salt concentrations. Simultaneously, the combination of salt-tolerant rhizobium inoculant and humic acid coating creates a "biochemical buffer zone" around the seed—the rhizobia reduce rhizosphere sodium levels through nitrogen fixation and extracellular polysaccharide secretion. + Concentration; humic acid through chelation of Na + Ca 2+ It replaces and stimulates root auxin synthesis, further alleviating salt stress. The three factors work synergistically to increase the germination rate from below 60% to over 72%. Secondly, well-rotted organic fertilizer and humic acid continuously decompose and release slow-release NPK and trace elements, avoiding nutrient imbalances or secondary salt spikes caused by traditional "one-shot" fertilization. Humic acid increases soil CEC and reduces Na+. + For K + Ca 2+ The competitive inhibition of salt damage helps maintain cellular ion homeostasis and significantly alleviates growth inhibition caused by salt stress. Finally, humic acid promotes a simultaneous increase in silique number and thousand-seed weight, while maintaining the oil content of salt-tolerant varieties, ultimately increasing yield from the traditional 50-70 kg / mu to 64-85 kg / mu. Therefore, the planting method for promoting *Capsella bursa-pastoris* in saline-alkali land provided by this invention systematically weakens the adverse effects of salt stress on seed germination, seedling growth, and later grain filling through a three-pronged strategy of "soil improvement - salt-tolerant varieties - seed coating," thereby significantly improving the germination rate, growth vigor, and yield of *Capsella bursa-pastoris* in saline-alkali land.

[0024] 2. The planting method provided by this invention is simple to operate and has controllable costs, making it suitable for large-scale promotion in moderately and mildly saline-alkali areas, and providing effective technical support for the development of agricultural resources in saline-alkali land and the sesame industry.

[0025] 3. The planting method provided by this invention addresses the problems of high salinity, pH imbalance, and insufficient fertility in saline-alkali soils. It achieves efficient cultivation of flaxseed in saline-alkali land through a combination of methods including applying well-rotted organic fertilizer to improve the soil, selecting salt-tolerant varieties, treating seeds with humic acid coating, and employing scientific sowing techniques. Practice shows that this planting method significantly reduces the surface soil salinity (by 17%–32%), adjusts the soil pH to a suitable range of 6.5–7.5, and simultaneously increases the germination rate (≥85%), seedling rate (≥80%), and yield (14%–28% higher than traditional planting methods). Furthermore, it is simple to operate, cost-effective, and suitable for large-scale promotion in moderately and mildly saline-alkali areas, providing a practical technical solution for the efficient utilization of saline-alkali land resources and the regional expansion of the flaxseed industry. Attached Figure Description

[0026] Figure 1 These are seeds and siliques of *Capsella flaxensis* at five different developmental stages in this invention. The development of *Capsella flaxensis* is divided into five stages based on seed and silique size and color, seed fresh weight, and days after flowering (DAF). Seed fresh weight is expressed as the mean ± standard error of five independent replicates.

[0027] Figure 2 This invention illustrates the effect of different treatments on the oil content of flaxseeds; where the data are the average of five independent replicates, and the error bars represent standard errors (SE).

[0028] Figure 3 The data represents the effect of different treatments on the yield of flaxseed in this invention; the data are the average of five independent replicates, and the error bars represent the standard error (SE).

[0029] Figure 4 This invention describes the changes in fatty acid composition during the development of *Capsella bursa-pastoris* seeds; wherein, (16:0): palmitic acid, (18:0): stearic acid, (18:1): oleic acid, (18:2): linoleic acid, (18:3): linolenic acid, (20:0): arachidic acid, (20:1): eicosenoic acid, and (22:1): erucic acid; the fatty acid composition of developing seeds was determined according to the description in the Materials and Methods section; the data are the average of five independent replicates, and the error bars represent standard errors (SE).

[0030] Figure 5The oil content in different tissues of *Capsella bursa-pastoris* in this invention is given; wherein the oil content is determined according to the description in the Materials and Methods section; the data is the average of five independent replicate samples, and the error bar represents the standard error (SE).

[0031] Figure 6 The fatty acid composition of different tissues of *Capsella bursa-pastoris* in this invention is shown below; wherein, (16:0): palmitic acid, (18:0): stearic acid, (18:1): oleic acid, (18:2): linoleic acid, (18:3): linolenic acid, (20:0): arachidic acid, (20:1): eicosenoic acid, and (22:1): erucic acid; the fatty acid composition of developing seeds was determined according to the description in the Materials and Methods section; the data are the average of five independent replicates, and the error bars represent standard errors (SE).

[0032] Figure 7 This invention describes the accumulation of triglycerides (TAG) in *Capsella linteus* seedlings induced by salt-alkali stress. Samples were collected at 0, 6, 12, 24, and 48 hours after salt-alkali stress treatment. The data are the average of five independent replicates, and the error bars represent standard errors (SE).

[0033] Figure 8 The differential expression of CsDGAT1s, CsDGAT2s, and CsPDATs in *Capsella bursa-pastoris* induced by salt-alkali stress (A, B, C) in this invention; wherein: A represents the differential expression of CsDGAT1s in *Capsella bursa-pastoris* induced by salt-alkali stress. B represents the differential expression of CsDGAT2s in *Capsella bursa-pastoris* induced by salt-alkali stress. C represents the differential expression of CsPDATs in *Capsella burmannii* induced by salt-alkali stress. Gene expression profiles in *Capsella burmannii* seedlings under salt-alkali stress were detected by qRT-PCR. Gene quantification was based on the expression of the endogenous housekeeping gene β-actin. The expression level of the control sample was used as the calibration value for the remaining samples (set to 1). Error bars represent the standard error (±SE) of six biological replicates. Detailed Implementation

[0034] The present invention will now be described in detail with reference to the accompanying drawings and specific embodiments, but this should not be construed as limiting the invention. Unless otherwise specified, the technical means used in the following embodiments are conventional means well known to those skilled in the art, and the materials, reagents, etc. used in the following embodiments are commercially available unless otherwise specified.

[0035] Example 1 A method for promoting the cultivation of flax in saline-alkali land includes the following steps: 1) Preparations before planting flaxseed: (1)Remove debris from the saline-alkali land for planting, eradicate weeds, stones, and dead branches and leaves to avoid the breeding of pests and diseases. Deeply plow and loosen the soil, plow the soil to a depth of 20 cm two weeks before sowing to break up the hardpan and increase soil aeration and water retention.

[0036] (2)Before sowing, spread the decomposed organic fertilizer evenly on the soil surface of the saline-alkali land for planting at a rate of 900 kg / mu, and then plow deeply by 25 cm to fully mix the decomposed organic fertilizer with the soil of the saline-alkali land to be planted.

[0037] Among them, the decomposed organic fertilizer is purchased from Zhejiang Fertilizer Co., Ltd., and the product number is NY / T525.

[0038] 2) Selection of seed varieties: Select Camelina sativa seeds of salt-tolerant varieties with strong adaptability and high oil content (40%): Select "SNC104".

[0039] Among them, the seeds of salt-tolerant Camelina sativa varieties: "SNC104" and "SC-N1" are systematically cultivated by the former research group of Molecular Agriculture and Bioenergy of Shanxi Agricultural University. The applicant holds the seeds of these two varieties and guarantees to provide them to the public within 20 years from the application date. If the public needs them, Shanxi Agricultural University can provide them externally. "SNC104" is published in the literature: Yuan Lixia, Mao Xue, Gao Changyong, Zhang Li, Xue Jinai, Yang Zhirong, Li Runzhi. Seed-specific expression of diacylglycerol acyltransferase (VgDGAT1) improves oil accumulation in Camelina sativa seeds [J]. Acta Phytophysiologica Sinica, 2015c, 51 (5): 668~678; "SC-N1" is published in the literature: Lixia Yuan (Yuan Lixia), Xue Mao, Kui Zhao, Xiajie Ji, Chunli Ji, Jinai Xue, Runzhi Li*. Characterisation of phospholipid: diacylglycerol acyltransferases (PDATs) from Camelina sativa and their roles in stress responses. Biology Open. 2017. 6 (7): 1024-1034..

[0040] 3) Seed treatment: Use humic acid to coat the Camelina sativa seeds to improve the emergence rate and obtain coated seeds.

[0041] Among them, the specific method is as follows: I. Selection of humic acid: Select humic acid (humic acid content ≥ 50%), and the dosage is 0.3% of the seed weight.

[0042] The humic acid was purchased from Jiuhong Chemical Co., Ltd.

[0043] II. Coating method: Preparation of coating solution: Dissolve potassium humate in warm water (50℃) at a mass ratio of 1:20, stir until completely dissolved, and cool to room temperature to obtain the coating solution.

[0044] Seed coating: Mix flax seeds with coating solution at a ratio of 10:1 (by mass), stir until the seeds are evenly coated, and then dry: air dry at low temperature (38℃) until the surface is no longer sticky to the touch to obtain coated seeds.

[0045] 4) Sowing: (1) Sowing time: Spring sowing (recommended): March 20 (ground temperature 6℃).

[0046] (2) Sowing method: Row sowing: row spacing 25cm, plant spacing 4cm, sowing depth 3cm (shallow sowing is recommended for saline-alkali soil), sowing rate 1.6kg / mu.

[0047] (3) Reasonable close planting to maintain seedling density: 250,000 plants per mu.

[0048] 5) Gains: Flax seeds were harvested on July 18.

[0049] Example 2: A planting method for promoting flaxseed in saline-alkali land Similar to Example 1, the difference lies in the variety of seeds. In this example, the seed variety selected is "SC-N1".

[0050] Example 3: A planting method for promoting flaxseed in saline-alkali land Similar to Example 1, the difference is that the raw material used in the preparation of the coating solution is a salt- and alkali-tolerant functional bacterial agent.

[0051] Among them, the salt-alkali tolerant functional bacterial agent is prepared by mixing the bacterial suspension of Pseudomonas PfP-1 and Bacillus amyloliquefaciens SQR9 at a volume ratio of 1:1.

[0052] The method for preparing Pseudomonas PfP-1 bacterial suspension includes the following steps: The activated seed culture of Pseudomonas PfP-1 was inoculated into a new Erlenmeyer flask containing LB liquid medium; the flask was then placed in a shaker and cultured at 29°C and 175 rpm for 15 hours. When the Pseudomonas bacteria reached the OD... 600 When the concentration is 1.5, the bacterial culture of Pseudomonas PfP-1 is obtained.

[0053] Among them, Pseudomonas PfP-1 was purchased from Jiangsu Suwei Microbial Research Co., Ltd.

[0054] The method for obtaining the bacterial culture of Bacillus amyloliquefaciens SQR9 includes the following steps: The activated seed culture of *Bacillus amyloliquefaciens* SQR9 was inoculated into a new Erlenmeyer flask containing LB liquid medium; the flask was then placed in a shaker and cultured at 37°C and 200 rpm for 30 hours. When the *Pseudomonas* reached the OD... 600 When the concentration is 1.5, the bacterial solution of Bacillus amyloliquefaciens SQR9 is obtained.

[0055] Bacillus amyloliquefaciens SQR9 was purchased from the China General Microbiological Culture Collection Center (CGMCC), with accession number CGMCC No. 5808.

[0056] Example 4: A planting method for promoting flaxseed in saline-alkali land Similar to Example 1, except that the coating solution is obtained by mixing the coating solution of Example 1 and a boric acid solution with a concentration of 0.1 g / mL at a volume ratio of 1:1.

[0057] The 0.1 g / mL boric acid solution was obtained by dissolving 1 g of boric acid in 999 g of water. The boric acid was purchased from Jinan Shengwang Chemical Co., Ltd.

[0058] Example 5: A planting method for promoting flaxseed in saline-alkali land Same as Example 1, except that no well-rotted organic fertilizer was applied to the saline-alkali land to be planted before sowing the seeds.

[0059] Example 6: A planting method for promoting flaxseed in saline-alkali land Same as Example 1, except that the seeds were not coated with humic acid before sowing.

[0060] To illustrate the effectiveness of the planting method for flaxseed provided by this invention for promoting flaxseed cultivation in saline-alkali land, the following study was conducted using the above-mentioned Example 1 as an example: I. Methods 1. Plant materials, growth status, and sampling This invention uses the flax variety “SNC104”, which has been commercially cultivated for 5 years in Dongyang Town, Yuci District, Jinzhong City, Shanxi Province, China.

[0061] The test site is located in Dongyang Town, Yuci District, Jinzhong City, Shanxi Province.

[0062] This experiment covered a total area of ​​23 mu (approximately 1.5 hectares), designated as the experimental area. Two experimental groups and six control groups were designed. The control group comprised 4 mu (approximately 0.67 hectares), and the experimental groups comprised 19 mu (approximately 1.2 hectares). The *Capsella bursa-pastoris* plants in each experimental group were planted using the planting method for promoting *Capsella bursa-pastoris* in saline-alkali land, as provided in Example 1.

[0063] The experimental groupings are shown in Table 1.

[0064] Table 1 Experimental Groups Note: "+" indicates that the experiment includes this content; "-" indicates that the experiment does not include this content.

[0065] The soil type in the experimental area is weakly alkaline saline soil, and its basic properties are shown in Table 2.

[0066] Table 2 Soil properties in the experimental area (0cm~50cm) Flaxseed plants from the different experimental groups were grown in a controlled greenhouse at 23°C under natural light. Newly opened flowers were tagged during the inflorescence stage. Flower and pod samples were collected at 7, 15, 22, 29, and 36 days post-flowering (DAF) for PCR, fresh weight, fatty acid composition, and oil and protein accumulation analysis.

[0067] Fully expanded leaves, stems, and roots were collected from plants with seven stem leaves. Tissue samples were collected from at least six *Capsella bursa-pastoris* plants in each experiment. All collected samples were immediately frozen in liquid nitrogen and stored at -80°C for RNA extraction, lipid determination, and other analyses. These experiments were repeated at least six times.

[0068] 2. Lipid extraction and gas chromatography (GC) detection of fatty acids Total lipids were extracted from seeds and vegetative tissue samples of *Capsella bursa-pastoris*. The specific steps were as follows: Samples of 20-30 mg were weighed from seeds at different developmental stages and from the vegetative tissue of *Capsella bursa-pastoris* plants. The samples were freeze-dried under high vacuum for 48 hours and then weighed. The water content was calculated by subtracting the weight recorded after freeze-drying from the weight recorded before freeze-drying. The freeze-dried samples were placed in a glass tube, and then 1 mL of 5% (v / v) sulfuric acid methanol solution (freshly prepared), 25 μL of 0.2% (v / v) butylated hydroxytoluene (BHT) methanol solution, 10 µg of triheptanyl glycerol (17:0), and 300 μL of toluene as a co-solvent were added. The mixture (sample + chemical solution) was briefly vortexed and then heated at 95 °C for 1.5 hours. After the mixture cooled to room temperature, 1.5 mL of 0.9% sodium chloride (w / v) and 1 mL of hexane were added to initiate a transmethylation reaction. Phase separation was performed by centrifuging the homogenate at 2000 rpm for 5 minutes. The upper organic phase containing fatty acid methyl esters (FAMEs) was transferred to a new dry test tube, and the remaining aqueous phase was extracted with 2 mL of hexane. The phases were then separated by centrifugation again. The organic phases were combined and dried under nitrogen. The nearly dried extract was dissolved in 1 mL of hexane, and 20 μL of the sample was transferred to a sample vial specifically designed for gas chromatography (GC) analysis.

[0069] Fatty acid methyl esters (FAMEs) of the lipid samples were separated using gas chromatography (Agilent 7890B) with an HP-88 column (0.25 mm inner diameter × 0.33 µm film thickness × 10 μm) and quantitative analysis was performed using a flame ionization detector. Three biological replicates were used in the gas chromatography experiments. The fatty acid composition was identified by comparing the retention times of fatty acids with those of known standards. The oil content in the samples was quantified by comparing the fatty acid concentration with the peak area of ​​the added internal standard (of known concentration). All data were statistically analyzed, and t-tests were used to determine the significance of differences between pairwise means.

[0070] 3. RNA preparation, cDNA synthesis, and qPCR detection Total RNA was extracted from each sample using the Plant RNeasy Mini Kit (Sigma-Aldrich) according to the manufacturer's instructions. The extracted RNA samples were further treated with DNase I (Promega) to remove contaminating DNA. RNA concentration (ng / μL) and purity ratios (260mm / 280nm and 260mm / 230nm) were determined using a NanoDrop 2000 spectrophotometer (Thermo Fisher Scientific).

[0071] Five μg of total RNA was extracted from each sample as a template, and first-strand cDNA was synthesized using the First-Strand cDNA Synthesis Kit (Fermentas). Real-time PCR experiments were performed according to the manufacturer's instructions, with each cycle incubated at 42°C for 30 minutes. The cDNA level in each sample was quantified and then diluted to a final concentration of 100 ng / μL as a template for qRT-PCR analysis.

[0072] All quantitative real-time PCR (qRT-PCR) experiments were performed on an iCycleriQ detection system (Bio-Rad) using the SYBR Green I Master Mix kit (Applied Biosystems). A reaction mixture without reverse transcriptase was used as a PCR control to ensure the RNA samples were free of DNA contamination. Each PCR reaction was performed in triplicate, including triplicate biological replicates. PCR reactions were performed in MicroAmp 96-well plates (Applied Biosystems) and covered with an optically adhesive sealing film (Applied Biosystems). Two [units / parts] were used. -ΔΔCt PCR products were quantified using a computational method (Schmittgen et al., 2008). The relative amounts of the target mRNA were standardized using the β-actin gene from *Capsella bursa-pastoris* as an internal control. Error bars represent the standard error (SE) of the fold change in relative target gene expression. Gene expression levels were calculated through three independent biological replicates and three PCR replicates for each sample. Primers used for the target gene analysis are listed in Table 3.

[0073] The 25 μL reaction system included: 1 μL of forward primer and 1 μL of reverse primer (500 nM each), 12.5 μL of SYBR Green Master Mix, 5 μL of cDNA diluted 1:10 (v / v), and 5.5 μL of HPLC molecular biology grade water.

[0074] The PCR program was set as follows: initial activation at 95℃ for 10 minutes, followed by 30-40 cycles (each cycle included denaturation at 95℃ for 15 seconds, annealing and extension at 60℃ for 1 minute), and a final extension at 72℃ for 5 minutes.

[0075] Table 3 Primer sequences used for PCR analysis of target genes in *Capsella bursa-pastoris*. II. Results 1. Fatty acid composition and lipid accumulation in flaxseed and other tissues during seed development. The seed development of cruciferous plants can generally be divided into an initial "growth stage," a second "accumulation stage," and a final "dehydration stage." The "growth stage" begins after fertilization and continues until the formation of the embryo; during this stage, cells divide rapidly, but the deposition of stored substances is minimal. In the "accumulation stage," the embryo continues to grow, and stored lipids and proteins are rapidly synthesized. The "dehydration stage" is characterized by dehydration and weak synthetic activity. To better study the dynamic changes in lipid and fatty acid accumulation, this study divided the development of *Capsella bursa-pastoris* seeds into five stages based on seed and silique size, color, seed fresh weight, and days after flowering (DAF). Figure 1 ).

[0076] Oil content of flaxseeds after various treatments ( Figure 2 ) and flaxseed yield ( Figure 3 Significant differences were observed. Fatty acid composition also showed specific changes in concentration with different developmental stages. Figure 4 Eight major fatty acids were detected in *Capsella bursa-pastoris* seeds, including palmitic acid (16:0), stearic acid (18:0), oleic acid (18:1), linoleic acid (18:2), alpha-linolenic acid (18:3), arachidic acid (20:0), eicosapentaenoic acid (20:1), and erucic acid (22:1). The composition of these fatty acids was almost opposite in early seed development (up to 15 DAF) to that in mature seeds. Early seeds showed higher levels of palmitic acid (21.9% at 7 DAF), stearic acid (20.4% at 7 DAF), oleic acid (22.1% at 15 DAF), and linoleic acid (34.3% at 15 DAF), while mature seeds showed higher levels of alpha-linolenic acid and eicosapentaenoic acid (41.3% and 12.2%, respectively).

[0077] As seeds develop, the contents of polyunsaturated linolenic acid, eicosapentaenoic acid (starting at 22 DAF), and erucic acid (starting at 22 DAF) increase significantly, reaching their peak at maturity. Correspondingly, the accumulation of the three saturated fatty acids (palmitic acid, stearic acid, and arachidic acid, starting at 15 DAF) gradually decreases, reaching their lowest levels in mature seeds. Furthermore, throughout seed development, the contents of monounsaturated oleic acid (starting at 15 DAF) and polyunsaturated linoleic acid (starting at 22 DAF) also decrease significantly.

[0078] The oil content and fatty acid composition of the vegetative organs (roots, stems, and leaves) of *Capsella bursa-pastoris* plants with seven stem leaves were also analyzed. Figure 4 As shown, the total oil content in vegetative organs is extremely low compared to seeds: 5.1% in leaves, 3.8% in roots, and 4.5% in stems. All eight major fatty acids present in seeds were detected in the vegetative organs of *Capsella bursa-pastoris* plants, with varying levels in different tissues. Figure 5 and Figure 6Among the fatty acids, linolenic acid was the most abundant, accounting for 29.7% in roots, 37.3% in stems, and 42.4% in leaves; followed by linoleic acid, accounting for 19.7% in roots, 18.3% in stems, and 10.6% in leaves; saturated palmitic acid was the third most abundant fatty acid in vegetative organs, accounting for 16.4% in roots, 19.2% in stems, and 17.7% in leaves. Furthermore, a relatively high content of 16:3 (13.5%) was detected in the photosynthetic organ, leaves, while the content of 16:3 was only trace (<1%) in non-photosynthetic organs such as roots and stems. Compared with seeds, the content of very long-chain fatty acids (VLCFAs) such as arachidic acid (20:0), eicosenoic acid (20:1), and erucic acid (22:1) was lower (<5%) in vegetative organs.

[0079] 2. Differential expression of CsDGAT1, CsDGAT2 and CsPDAT members induced by salt-alkali stress To determine whether members of the CsDGAT1, CsDGAT2, and CsPDAT family are involved in the response of *Capsella lindensis* to abiotic stress, three-week-old seedlings grown under saline-alkali stress were harvested. Seedling samples were divided into two batches: one batch was used for total lipid extraction and triglyceride (TAG) determination, and the other batch was used for total RNA extraction and quantitative real-time PCR (qRT-PCR) analysis. During saline-alkali stress, the triglyceride content in seedlings gradually increased, reaching its highest level at 24 hours (1.9 times that of the control group), and showed a rapid increase between 12 and 24 hours of stress. Figure 7 Differential expression of CsDGAT1s, CsDGAT2s, and CsPDATs was observed in *Capsella bursa-pastoris* seedlings induced by salt-alkali stress. The transcriptional levels of CsDGAT1-B and CsPDAT2 were significantly upregulated, peaking at 12 and 24 hours, respectively, representing increases of 15-fold and 8.5-fold compared to the control group. Figure 8 A and Figure 8 (C in the text). However, the mRNA levels of CsPDAT3, CsDGAT1-C, and CsDGAT2-A decreased during stress (C in the text). Figure 8 A in Figure 8 B and Figure 8 (C in the original text). Other gene members tested did not show significant changes.

[0080] Overall, the dynamic analysis above indicates that the expression of CsDGAT1-B and CsPDAT2 is closely related to the increase in lipid content induced by salt-alkali stress.

[0081] In summary, this invention identifies three members belonging to the CsDGAT1, CsDGAT2, and CsPDAT protein families within the *Capsella linteus* genome. These proteins catalyze the final acylation step in triacylglycerol (TAG) biosynthesis. The expression of these members exhibits distinct spatiotemporal patterns: transcripts of CsDGAT1-A and CsDGAT2-C accumulate and are expressed in large quantities during seed development. Furthermore, these members are regulated differently by abiotic stresses, with saline-alkali stress significantly upregulating the expression of CsDGAT1-B and CsPDAT2. This demonstrates for the first time that under saline-alkali stress, plant seedlings enhance triacylglycerol accumulation by activating different members of CsDGAT1, CsDGAT2, and CsPDAT, thereby promoting their adaptation to adverse environments. Given that capsella linteus oil is an ideal resource for both human health and bioenergy, the data from this study will contribute to the cultivation of capsella linteus plants with higher biomass and greater tolerance to various environmental stresses.

[0082] The present invention provides a planting method for promoting flaxseed in saline-alkali land, entitled "A planting method for promoting flaxseed in saline-alkali land". It precisely addresses the thorny problems of high soil salinity, pH imbalance and insufficient fertility in saline-alkali land, and forms a scientific and efficient technical solution.

[0083] In terms of soil improvement, the application of well-rotted organic fertilizer played a crucial role. The addition of well-rotted organic fertilizer is like injecting "vitality factors" into saline-alkali land; its rich organic matter effectively improves soil aggregate structure, enhances soil water and fertilizer retention capacity, and gradually regulates soil pH, alleviating excessive soil salinity. This process establishes a virtuous cycle system for the soil, making the soil environment more suitable for plant growth and laying a solid foundation for the rooting and growth of flax and watercress.

[0084] In terms of variety selection, screening for salt-tolerant varieties is the core strategy for coping with the harsh environment of saline-alkali land. These carefully selected varieties, with their unique physiological characteristics, such as special ion transport mechanisms and osmotic regulation capabilities, can grow tenaciously in soils with high salinity and abnormal pH, ensuring the feasibility of planting flax in saline-alkali land from the source and greatly increasing its survival probability in adverse conditions.

[0085] Humic acid seed coating technology adds a protective layer to seed germination and growth. Humic acid itself has many excellent properties, such as improving soil physical and chemical properties and enhancing plant resistance. Through coating treatment, humic acid can create a relatively mild microenvironment around the seed, reducing the direct toxicity of salt to the seed, promoting seed water absorption and germination, increasing seed emergence rate and seedling vigor, and enabling flax to better resist the adverse factors of saline-alkali soil in the early stages of growth.

[0086] The application of scientific sowing techniques fully considers the special conditions of saline-alkali land and is an important guarantee for achieving high-efficiency planting. Rationally planning the sowing time and avoiding sowing during periods of excessively high salt concentration or unsuitable temperature can reduce the negative impact of external factors on seed germination and seedling growth. Precisely controlling the sowing depth ensures that seeds receive sufficient water and nutrients while avoiding salt damage from sowing too deep or too shallow. Optimizing the sowing density ensures that plants can fully utilize soil nutrients, water, and light resources during their growth process, reducing intraspecific competition, promoting individual growth, and thus increasing overall yield.

[0087] In summary, this invention systematically solves the technical challenges of planting flaxseed in saline-alkali land through a multi-dimensional approach, which involves the synergistic combination of increased application of well-rotted organic fertilizer, selection of salt-tolerant varieties, humic acid-coated seed treatment, and scientific sowing techniques. This achieves efficient cultivation of flaxseed in saline-alkali land, opens up new pathways for agricultural development and utilization of saline-alkali land, and has significant economic, ecological, and social benefits, with broad prospects for promotion and application.

[0088] It should be noted that when numerical ranges are involved in this invention, it should be understood that the two endpoints of each numerical range and any value between the two endpoints can be selected. To avoid redundancy, this invention describes preferred embodiments.

[0089] Although preferred embodiments of the invention have been described, those skilled in the art, upon learning the basic inventive concept, can make other changes and modifications to these embodiments, all of which fall within the scope of the invention.

Claims

1. A method for promoting the cultivation of flaxseed in saline-alkali land, characterized in that, Includes the following steps: Before sowing, apply well-rotted organic fertilizer to the saline-alkali land to be planted; Choose salt-tolerant varieties of flax seeds; Flaxseeds seeds were coated with a coating solution to obtain coated seeds. The coating solution was a salt-tolerant functional bacterial solution or a humic acid solution, wherein the effective components of the salt-tolerant functional bacterial solution were Pseudomonas PfP-1 and Bacillus amyloliquefaciens SQR9. The coated seeds are sown from late March to early April each year, when the soil temperature is ≥5℃. After sowing, the plants are cultivated to obtain flax seedlings.

2. The planting method according to claim 1, characterized in that, The salt- and alkali-tolerant functional bacterial solution is prepared by mixing the bacterial solution of *Pseudomonas* PfP-1 and the bacterial solution of *Bacillus amyloliquefaciens* SQR9 at a volume ratio of 1:0.8-1.

2. The OD of the bacterial solution of *Pseudomonas* PfP-1 is... 600 The OD value of the bacterial culture of *Bacillus amyloliquefaciens* SQR9 was 1.4~1.

6. 600 It is 1.4~1.

6.

3. The planting method according to claim 1, characterized in that, The steps for coating are as follows: The flax seeds are mixed with the coating solution at a mass ratio of 9 to 11:1, stirred to ensure that the flax seeds are evenly coated with the coating solution, and then dried to obtain the coated seeds.

4. The planting method according to claim 3, characterized in that, The drying process is either air drying or low-temperature air drying; where low temperature refers to a temperature of 15℃~25℃.

5. The planting method according to claim 1, characterized in that, The amount of the well-rotted organic fertilizer used is 800 kg / mu to 1000 kg / mu.

6. The planting method according to claim 1, characterized in that, The seeds of the flaxseed are salt-tolerant varieties with an oil content of 35% to 42%.

7. The planting method according to claim 6, characterized in that, The varieties of the flaxseed include "SNC104" or "SC-N1".

8. The planting method according to claim 1, characterized in that, The sowing method is row sowing, with a row spacing of 24cm~26cm, a plant spacing of 3cm~5cm, and a sowing depth of 1cm~2cm.

9. The planting method according to claim 1, characterized in that, The sowing rate is 1.5 kg / mu to 1.7 kg / mu.

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