A method for breeding a high-yield drought-resistant space-bred millet variety

Abstract

The application discloses a high-yield drought-resistant space breeding millet variety breeding method, and relates to the technical field of crop genetic breeding.The application comprises the following steps: step 1, parent selection; step 2, M1 generation planting and mutation population construction; step 3, M2 generation mutant screening; step 4, M3-M4 generation strain identification and molecular marker assisted selection; step 5, whole growth period dynamic drought resistance precision identification and water use efficiency evaluation; and step 6, multi-point regional test and variety approval.The application creates a rich genetic variation population through space mutagenesis, combines a stress and non-stress double-environment two-step screening method, simultaneously investigates the drought resistance phenotype and yield potential of a single plant in the M2 generation, significantly improves the pertinence and efficiency of early selection, further utilizes molecular marker assisted selection to perform directional polymerization on high-yield and drought-resistant related genes in the M3 generation, realizes the change from random screening to directional screening, greatly improves the selection accuracy of favorable genotypes, and reduces the subsequent breeding workload.

Description

Technical Field

[0001] This invention belongs to the field of crop genetics and breeding technology, specifically a method for breeding high-yield, drought-resistant millet varieties through space breeding. Background Technology

[0002] Millet is an important food and forage crop in the dryland farming areas of northern my country, characterized by its drought resistance, tolerance to poor soil, and high water use efficiency. However, traditional millet breeding faces bottlenecks such as a narrow genetic base, difficulty in synergistically improving high yield and drought resistance traits, and long breeding cycles.

[0003] Space breeding is an important method for creating new germplasm resources by inducing extensive variations in crop genomes using the unique environment of space. Compared with traditional radiation-induced mutagenesis, space-induced mutagenesis has the advantages of a larger range of variations, more beneficial variations, and a relatively shorter breeding cycle.

[0004] Existing methods for space breeding millet typically include: sending materials into the space environment, planting and observing variations after return to Earth, selecting individual plants across multiple generations based on agronomic traits, and finally assessing yield and drought resistance. However, these methods suffer from the blindness of mutagenesis and selection. Space-induced mutations produce random variations, and traditional methods rely solely on field phenotypes for selection, resulting in low efficiency and difficulty in accurately identifying individual plants with both high yield and drought resistance potential from a large population of variants. Furthermore, existing methods often employ single drought stress or simple field drought greenhouse assessments, failing to comprehensively and quantitatively evaluate a variety's drought resistance, water use efficiency, and yield stability throughout its growth cycle. This leads to varieties with incomplete drought resistance or a disconnect from actual production needs. Moreover, high yield and drought resistance often present physiological and genetic contradictions. Traditional phenotypic selection struggles to effectively break the negative correlation between the two in early generations, often resulting in neglecting one aspect and causing repeated iterations in the breeding process.

[0005] Therefore, this invention provides a method for breeding high-yield, drought-resistant millet varieties through space breeding, in order to solve the problems of low screening efficiency, inaccurate drought resistance identification, and difficulty in synergistic improvement of high yield and drought resistance traits in the existing technology.

[0006] The information disclosed above in this background section is only for enhancing the understanding of the background section of this invention, and therefore may include prior art that is not known to those skilled in the art. Summary of the Invention

[0007] This invention aims to establish an efficient, precise, and targeted breeding system for high-yield and drought-resistant millet varieties by integrating space-induced mutagenesis, molecular marker-assisted selection, and dynamic drought resistance identification technologies across multiple ecological zones and the entire growth period. This system will address the problems of low screening efficiency, inaccurate drought resistance identification, and difficulty in synergistically improving high yield and drought resistance traits in existing technologies.

[0008] To achieve the above objectives, the present invention provides the following technical solution: a method for breeding high-yield, drought-resistant space-bred millet varieties, comprising the following steps:

[0009] Step 1. Parental selection: Select seeds of high-yielding but moderately drought-resistant backbone millet varieties for space-induced mutation.

[0010] Step 2. M1 generation planting and mutant population construction: Plant the M0 generation seeds recovered after spaceflight to obtain M1 generation plants. Harvest all the seeds of the M1 generation plants to obtain the basic mutant population, i.e., M1 mixed seed harvest.

[0011] Step 3. Screening of M2 generation mutants: The M1 mixed-harvest seeds were planted in two separate areas under drought stress and non-stress conditions to obtain the M2 generation mutant population. First, individual plants with outstanding drought resistance were screened in the drought stress area. Then, individual plants with significantly higher yield components than the population average were screened in the non-stress area. Only the seeds of the corresponding individual plants that performed well in both environments were retained.

[0012] Step 4. Identification of M3-M4 generation lines and molecular marker-assisted selection: The seeds of the single plants selected in Step 3 were planted according to the lines. In the M3 generation, the genotype of the lines was tested using molecular markers related to high yield and drought resistance of millet. Stable lines that carry both high-yield and drought-resistant dominant alleles and have excellent agronomic traits were screened out.

[0013] Step 5. Precise identification of dynamic drought resistance and assessment of water use efficiency throughout the entire growth period: The stable lines selected in Step 4 were placed at at least 2 representative ecological sites with different water treatments, and physiological indicators were dynamically monitored. The yield was measured and the drought resistance coefficient and water use efficiency were calculated. Lines with drought resistance coefficient greater than 0.7, high water use efficiency and no decline in grain yield potential under moderate and severe drought stress were screened.

[0014] Step 6. Multi-location regional trials and variety approval: The high-yielding and drought-resistant superior lines selected in Step 5 will be subjected to multi-location and multi-year regional trials and production trials. After meeting the standards, an application for variety approval will be submitted.

[0015] Preferably, in step 1, the key high-yield millet variety has a thousand-grain weight ≥3.0g, a yield potential of ≥400kg per mu, and a drought resistance rating of medium or below.

[0016] Preferably, in step 3, the screening indicators for individual plants with outstanding drought resistance include light leaf curling, good leaf color retention, and strong stem water retention; the yield components include single plant ear weight, ear grain weight, and thousand grain weight, and significantly higher than means that the trait value is greater than the average value of the M2 generation population plus one standard deviation.

[0017] Preferably, in step 4, the molecular markers associated with high yield and drought resistance include at least two of the following: SiGN1 gene marker associated with grain number per ear, GW2 homologous gene marker associated with grain weight, DRO1 homologous gene marker associated with root architecture, and P5CS gene marker associated with proline synthesis.

[0018] Preferably, in step 5, the different water treatments include full irrigation, moderate drought stress, and severe drought stress; the dynamically monitored physiological indicators include leaf photosynthetic rate, stomatal conductance, canopy temperature, and chlorophyll fluorescence parameter Fv / Fm.

[0019] Preferably, in step 5, the screening criteria are: under moderate and severe drought stress, the drought resistance coefficient is greater than 0.7, the water use efficiency is significantly higher than the control parent, and the grain yield is not less than 95% of the control parent under abundant water conditions.

[0020] Preferably, in step 5, the water use efficiency is determined by a weighing lysimeter or a method based on the soil moisture balance equation.

[0021] Preferably, in step 2, the M1 generation is planted in sparse, large-hole cultivation with a row spacing of 40cm × 15cm.

[0022] Compared with the prior art, the present invention has the following beneficial effects:

[0023] 1. This invention creates a rich population of genetic variations through space-induced mutagenesis and combines a two-step screening method with both stress and non-stress environments. In the M2 generation, the drought resistance phenotype and yield potential of individual plants are examined simultaneously, which significantly improves the targeting and efficiency of early selection. At the same time, molecular marker-assisted selection is further used to target and aggregate high-yield and drought-resistant genes in the M3 generation, realizing the transformation from random selection to targeted selection, greatly improving the accuracy of selecting favorable genotypes and reducing the workload of subsequent breeding.

[0024] 2. This invention integrates molecular marker-assisted selection and dynamic drought resistance identification across multiple ecological sites throughout the entire growth period. It can accurately break the common genetic negative correlation between high yield and drought resistance traits. In drought resistance identification, it not only assesses the drought resistance coefficient and water use efficiency of varieties under drought stress, but also ensures that their yield potential does not decrease under abundant water conditions. Thus, it successfully breeds varieties that are good at both water and drought conditions, solving the problem of difficulty in balancing yield and stress resistance in traditional breeding.

[0025] 3. Compared with traditional single-period or qualitative identification methods, this invention sets up multi-gradient water treatment at at least two representative ecological sites and dynamically monitors key physiological indicators. Combined with precise water use efficiency measurement using a weighing lysimeter, it can comprehensively and quantitatively evaluate the drought resistance response and yield stability of varieties under different drought intensities and different growth stages. This makes the selected varieties more suitable for the complex water conditions in actual production, and their drought resistance is more comprehensive and reliable.

[0026] 4. This invention broadens the genetic basis through space-induced mutagenesis, early MAS directional selection, and precise and efficient phenotypic identification, effectively reducing blind breeding and repeated experiments, accelerating the stabilization and aggregation of superior traits. Compared with traditional breeding methods, it can obtain superior lines with stable traits, high yield and drought resistance in a shorter generation, significantly improving the overall efficiency and success rate of breeding. Attached Figure Description

[0027] To more clearly illustrate the technical solutions in the embodiments of this application or the prior art, the drawings used in the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments recorded in this invention. For those skilled in the art, other drawings can be obtained based on these drawings.

[0028] Figure 1 This is a schematic diagram of the breeding process of the present invention. Detailed Implementation

[0029] To enable those skilled in the art to better understand the technical solutions of the present invention, the present invention will be further described in detail below with reference to the accompanying drawings. All other embodiments obtained by those skilled in the art based on the embodiments of the present invention without creative effort are within the scope of protection of the present invention.

[0030] A method for breeding a high-yielding, drought-resistant millet variety through space breeding includes the following steps:

[0031] Step 1: Parent Selection

[0032] Select 2-3 high-yielding but moderately drought-resistant backbone high-yielding millet varieties as basic parent lines; the backbone high-yielding millet varieties have a thousand-grain weight ≥3.0g, a yield potential of ≥400kg per mu, and a drought resistance rating of moderate or below; the selected parent seeds will be carried on a recoverable satellite, spacecraft, or space station, and returned to Earth after staying in the space environment for a certain period of time.

[0033] Step 2: M1 generation planting and mutant population construction

[0034] M0 generation seeds recovered after spaceflight were planted in the same year and on the same plot of land as ground control seeds to obtain M1 generation plants. During planting, sparse planting with large holes was adopted, with a row spacing of 40cm×15cm to ensure full development of individual plants. Only basic traits were observed and recorded, and no culling was carried out. After maturity, the seeds of all M1 generation plants were harvested by mixing plants on a single ear basis to obtain the basic mutant population, namely M1 mixed-harvest seeds.

[0035] Step 3: Screening of M2 generation mutants

[0036] The M1 mixed-harvest seeds were planted in separate areas under both drought stress and non-stress conditions to obtain the M2 generation mutant population.

[0037] The first step is to conduct preliminary phenotypic screening. During the critical growth period from jointing to heading, individual plants with outstanding drought resistance are selected in drought-stressed areas, and their plant numbers are recorded. Screening indicators include: light leaf curling, good leaf color retention, and strong stem water holding capacity.

[0038] The second step involves a re-screening of yield potential. At maturity, in a non-stressed area, the yield components of the selected individual plants from the previous step are measured. The screening criteria are: ear weight, ear grain weight, and thousand-grain weight per plant are significantly higher than the average of the M2 generation population and not lower than the control parent. Simultaneously, in the drought-stressed area, seeds from the corresponding plant with the best drought resistance are harvested. "Significantly higher than" means that the trait value is greater than the average of the M2 generation population plus one standard deviation.

[0039] Only the seeds of the corresponding individual plants that performed well in both the first and second steps were retained to enter the next generation.

[0040] Step 4: Identification of M3-M4 generation lines and marker-assisted selection (MAS)

[0041] The selected individual seeds from step 3 were planted according to their lineages for lineage comparison experiments.

[0042] In the M3 generation, genotyping of the lines was performed using molecular markers associated with high millet yield and drought resistance. The molecular markers included at least two of the following: SiGN1 gene marker associated with grain number per ear, GW2 homologous gene marker associated with grain weight, DRO1 homologous gene marker associated with root architecture, and P5CS gene marker associated with proline synthesis.

[0043] Lines carrying both high-yield and drought-resistant alleles were screened out, and selected based on field agronomic traits to harvest superior individual plants; field agronomic traits included plant height, ear type, and lodging resistance.

[0044] In the M4 generation, MAS verification and trait stability observation were continued for the superior lines to obtain stable lines with basically uniform agronomic traits and aggregated target genes.

[0045] Step 5: Precise identification of dynamic drought resistance and assessment of water use efficiency throughout the entire growth period

[0046] The selected stable lines, substituted with M4, were subjected to a combination of precise water control experiments under rain shelters and field natural drought assessment at at least two representative ecological sites, such as semi-arid and drought-prone areas.

[0047] Three water treatments were set up: full irrigation: 75%-80% of field capacity, moderate drought stress: 50%-55% of field capacity, and severe drought stress: 35%-40% of field capacity.

[0048] Dynamic monitoring of physiological indicators: Leaf photosynthetic rate, stomatal conductance, canopy temperature, and chlorophyll fluorescence parameter Fv / Fm were measured at different growth stages, including seedling stage, jointing stage, and heading and grain-filling stage.

[0049] Grain yield, biomass, and harvest index were measured at harvest time; drought resistance coefficient and water use efficiency were calculated. Water use efficiency (WUE) was accurately measured using a weighing lysimeter or a method based on the soil moisture balance equation. WUE = grain yield / water consumption.

[0050] The lines that exhibited a drought resistance coefficient greater than 0.7 under moderate and severe drought stress, significantly higher water use efficiency than the control parents, and grain yields no less than 95% of the control parents under abundant water conditions were selected and designated as high-yielding and drought-resistant superior lines.

[0051] Step 6: Multi-regional trials and variety approval

[0052] The high-yield and drought-resistant superior lines selected in step 5 are upgraded to varieties and participate in provincial or national regional and production trials of millet varieties. Trials are conducted in at least three different ecological zones for 2-3 years to comprehensively evaluate their yield stability, adaptability, drought resistance, and quality. After meeting the approval standards, an application for variety approval is submitted.

[0053] Example 1:

[0054] Step 1. Parent selection: Select 300 grams of homozygous seeds of the high-yielding but drought-resistant variety 'Jigu 19', carry them on a recoverable scientific experimental satellite, and retrieve them after about 15 days in orbit.

[0055] Step 2. M1-M2 generation screening: In the following spring, at the experimental station, the recovered M0 seeds and ground control seeds were sown simultaneously to obtain the M1 generation. During planting, sparse planting with large holes was adopted, with a row spacing of 40cm × 15cm. After maturity, the ears were harvested from individual plants, and the grains were mixed and threshed to obtain the M2 generation population seeds.

[0056] Step 3. In the third year, establish a drought-stressed zone and a normally irrigated zone, planting 5000 M2 generation plants. In the drought-stressed zone, initially screen out 150 individual plants with light leaf curling, good leaf color retention, and strong stem water-holding capacity, and mark them. In the normally irrigated zone, yield is measured on the corresponding plants of these marked individual plants, and 50 individual plants with ear weight, ear grain weight, and thousand-grain weight that are at least 15% higher than the control average are selected. Finally, harvest the corresponding ears from these 50 individual plants in the drought-stressed zone and enter the M3 generation.

[0057] Step 4. M3-M4 Generation MAS: Fifty M3 lines were planted sequentially. Leaves were taken at the seedling stage, and the favorable alleles of SiGN1, SiDRO1, and SiP5CS were detected using KAM genotyping technology. Twelve lines containing all three favorable alleles were selected. Three lines with poor agronomical traits were eliminated in the field, retaining nine lines. These nine lines were then planted in the M4 generation. Genotyping was consistent, and field traits were uniform and stable. Five of the best lines were selected for the next stage of identification.

[0058] Step 5. Precise drought resistance assessment: Five superior lines, along with the control 'Jigu 19' and the drought-resistant control 'Jingu 21', were subjected to precise water control experiments under rain shelters at semi-arid and arid test sites. Three treatments were set up: full irrigation throughout the entire growth period (CK), moderate stress after jointing (MS), and moderate stress throughout the entire growth period (SS). Soil moisture content was monitored using soil moisture sensors. The results showed that, under the SS treatment at the arid test site, superior line A achieved a yield drought resistance coefficient of 0.75, with water use efficiency 25% higher than 'Jigu 19' and 8% higher than 'Jingu 21'; under the CK treatment at the semi-arid test site, its yield was on par with 'Jigu 19'. Superior line A showed the best overall performance.

[0059] Step 6. Regional Trials and Approval: The superior line A will participate in regional trials in the summer valley region of North China. Its yield stability, adaptability, drought resistance, and quality will be comprehensively evaluated. Once it meets the approval standards, an application for variety approval will be submitted.

[0060] This invention creates mutations through space-induced mutagenesis, performs two-step screening under both stress and non-stress conditions, uses molecular markers to directionally aggregate beneficial genes, and conducts dynamic and precise drought resistance identification across multiple ecological sites throughout the entire growth period to breed high-yield and drought-resistant superior lines. This achieves efficient synergistic improvement of high-yield and drought-resistant traits, significantly improves breeding efficiency and selection accuracy, overcomes the blindness of traditional methods, and shortens the breeding cycle.

[0061] The foregoing has only described certain exemplary embodiments of the present invention by way of illustration. Undoubtedly, those skilled in the art can modify the described embodiments in various ways without departing from the spirit and scope of the present invention. Therefore, the foregoing drawings and descriptions are illustrative in nature and should not be construed as limiting the scope of protection of the claims of the present invention.

Claims

1. A method for breeding high-yielding, drought-resistant millet varieties through space breeding, characterized in that: Includes the following steps: Step 1. Parental selection: Select seeds of high-yielding but moderately drought-resistant backbone millet varieties for space-induced mutation. Step 2. M1 generation planting and mutant population construction: Plant the M0 generation seeds recovered after spaceflight to obtain M1 generation plants. Harvest all the seeds of the M1 generation plants to obtain the basic mutant population, i.e., M1 mixed seed harvest. Step 3. Screening of M2 generation mutants: The M1 mixed-harvest seeds were planted in two separate areas under drought stress and non-stress conditions to obtain the M2 generation mutant population. First, individual plants with outstanding drought resistance were screened in the drought stress area. Then, individual plants with significantly higher yield components than the population average were screened in the non-stress area. Only the seeds of the corresponding individual plants that performed well in both environments were retained. Step 4. Identification of M3-M4 generation lines and molecular marker-assisted selection: The seeds of the single plants selected in Step 3 were planted according to the lines. In the M3 generation, the genotype of the lines was tested using molecular markers related to high yield and drought resistance of millet. Stable lines that carry both high-yield and drought-resistant dominant alleles and have excellent agronomic traits were screened out. Step 5. Precise identification of dynamic drought resistance and assessment of water use efficiency throughout the entire growth period: The stable lines selected in Step 4 were placed at at least 2 representative ecological sites with different water treatments, and physiological indicators were dynamically monitored. The yield was measured and the drought resistance coefficient and water use efficiency were calculated. Lines with drought resistance coefficient greater than 0.7, high water use efficiency and no decline in grain yield potential under moderate and severe drought stress were screened. Step 6. Multi-location regional trials and variety approval: The high-yielding and drought-resistant superior lines selected in Step 5 will be subject to multi-location and multi-year regional trials and production trials. After meeting the standards, an application for variety approval will be submitted.

2. The method for breeding high-yield, drought-resistant space-bred millet varieties according to claim 1, characterized in that: In step 1, the key high-yield millet varieties have a thousand-grain weight ≥3.0g, a yield potential of ≥400kg per mu, and a drought resistance rating of medium or below.

3. The method for breeding high-yield, drought-resistant space-bred millet varieties according to claim 1, characterized in that: In step 3, the screening criteria for individual plants with outstanding drought resistance include light leaf curling, good leaf color retention, and strong stem water retention; the yield components include single plant ear weight, ear grain weight, and thousand grain weight, and significantly higher than means that the trait value is greater than the average value of the M2 generation population plus one standard deviation.

4. The method for breeding high-yield, drought-resistant space-bred millet varieties according to claim 1, characterized in that: In step 4, the molecular markers associated with high yield and drought resistance include at least two of the following: SiGN1 gene marker associated with grain number per ear, GW2 homologous gene marker associated with grain weight, DRO1 homologous gene marker associated with root architecture, and P5CS gene marker associated with proline synthesis.

5. The method for breeding high-yield, drought-resistant millet varieties through space breeding according to claim 1, characterized in that: In step 5, the different water treatments include full irrigation, moderate drought stress, and severe drought stress; the dynamically monitored physiological indicators include leaf photosynthetic rate, stomatal conductance, canopy temperature, and chlorophyll fluorescence parameter Fv / Fm.

6. A method for breeding high-yield, drought-resistant millet varieties through space breeding according to claim 1 or 5, characterized in that: In step 5, the screening criteria are: under moderate and severe drought stress, the drought resistance coefficient is greater than 0.7, the water use efficiency is significantly higher than the control parent, and the grain yield is not less than 95% of the control parent under abundant water conditions.

7. The method for breeding high-yield, drought-resistant space-bred millet varieties according to claim 1, characterized in that: In step 5, the water use efficiency is determined by a weighing lysimeter or by a method based on the soil moisture balance equation.

8. The method for breeding high-yield, drought-resistant millet varieties through space breeding according to claim 1, characterized in that: In step 2, the M1 generation is planted in sparse, large-hole cultivation with a row spacing of 40cm × 15cm.

Images