New herbicide-resistant mulberry germplasm creation and optimal matching rapid propagation technology

Abstract

The invention relates to a herbicide-resistant mulberry new germplasm creation and optimal matching rapid propagation technology, which comprises the following steps: constructing 86 resistant single plants by 180Gy electron beam irradiation, and distinguishing the 86 resistant single plants into a stock type and a scion type according to root biomass and a leaf resistance index; performing two-stage screening on 1260 groups of excessive matching groups to obtain 3 sets of optimized combinations with the death rate reduced by more than or equal to 15%; a double-backup mechanism of'main part + 2 parts of alternative parts' is established, and annual rapid propagation of 150000 plants is realized by means of cloud collaboration. The method fills the blank of mulberry resistance breeding matching optimization and risk prevention and control.

Description

TECHNICAL FIELD

[0001] The application belongs to the field of agricultural biotechnology, and relates to a method for creating mulberry herbicide-resistant germplasm by electron accelerator mutagenesis combined with molecular marker screening, obtaining an optimized combination of “rootstock and scion double resistance” through 1260 groups of over-dosage grouping and two-stage screening, and realizing large-scale rapid propagation by relying on a double backup mechanism and cloud collaboration. BACKGROUND

[0002] Herbicide drift causes annual losses of more than 30 million yuan in Jiangsu's silk and mulberry industry, threatening the livelihoods of 150,000 sericulturists. The existing technology has three defects: 1. Single breeding target: focusing on comprehensive trait varieties, without separately creating rootstock and scion resistance materials for the entire grafting system of silkworm mulberry; 2. Fragmented technical chain: the links of mutagenesis creation, grouping screening and rapid propagation are lack of engineering connection; 3. Laggard resistance concept: single-site resistance cannot cope with the dual stress of soil residue and air drift.

[0003] The field urgently needs to shift from “single variety breeding” to a systematic solution of “functional module creation-optimized grouping-risk prevention and control of rapid propagation”. SUMMARY

[0004] (I) A resistance germplasm classification creation method is provided, 20,000 seed-derived M2 populations are constructed by 180Gy electron beam irradiation, 86 resistant single plants are obtained through herbicide stress and KASP marker verification, and are divided into rootstock and scion materials according to root biomass and leaf resistance index.

[0005] (II) A 1260-group over-dosage grouping method is provided, in which 30 rootstock main parts and 14 alternatives, 28 scion main parts and 14 alternatives are completely double-grafted, and 2-3 sets of optimized combinations with a mortality reduction of ≥15% are obtained through two-stage screening.

[0006] (III) A risk prevention and control rapid propagation method is provided, a “main part + 2 alternatives” backup is established for each germplasm, rootstock cutting and scion tissue culture are carried out in parallel, cloud platform realizes cross-regional progress collaboration, and annual production capacity is ≥150,000 plants.

[0007] Due to the adoption of the above technical solutions, the application has the following advantages and positive effects: 1. Systematic: the first “classification creation-over-dosage grouping-risk rapid propagation” full-chain technology fills the gap in grouping screening and prevention mechanism; 2. High efficiency: 1260-group grouping increases the probability of optimization by 50%, and two-stage screening shortens the cycle by 30%; 3. Robustness: double backup and cloud collaboration reduce the risk of germplasm loss to <1% and delay risk by 40%. 4. Industrialization: Within 3 years, a pilot plant of 50 mu and a demonstration network of 500 households will be established, and the procurement commitment of leading enterprises will guarantee the implementation of the industry. Detailed Implementation Example

[0008] 20,000 seeds of Husang 32' (or better) were irradiated with a 12MeV electron beam at 180Gy to construct an M2 generation mutant population of 8,000 plants. After three generations of breeding in Hainan, glyphosate was sprayed at 2000 g / ha, yielding 86 candidate resistant plants. Following KASP marker verification, 44 plants were classified as rootstock type based on root dry weight ≥ 130% of the control, and 42 plants were classified as scion type based on leaf resistance index ≥ 2.0. Example

[0009] Thirty main rootstocks and 14 alternatives, and 28 main scions and 14 alternatives were selected to construct a grafting matrix of 1260 groups. Callus formation rate was observed microscopically 15 days after grafting, and 357 groups (28.3%) were eliminated. The remaining 903 groups were sprayed with glufosinate-ammonium at 1500 g / ha. Three combinations were selected that reduced mortality by ≥15% compared to the monoclonal antibody control: R7-15 / Yu71-1 (reduction of 31%), R18-22 / Husang32 (reduction of 22%), and R33-9 / Nongsang14 (reduction of 19%), with grafting survival rates of 88.5%, 86.2%, and 87.8%, respectively. Example

[0010] For each core germplasm, an independent line system of "primary + 2 alternatives" was established. Rootstock-type materials were propagated by cuttings, and scion-type materials were rapidly propagated through tissue culture (proliferation coefficient 6.0, 5 subcultures per year). A DingTalk-JianDaoYun collaborative platform was developed, integrating Hainan meteorological data, Sanya accelerator progress, and Yancheng rapid propagation capacity. After the Hainan pollination delay warning was triggered in July 2027, A and B team personnel arrived within 48 hours to expedite harvesting, ensuring that 8,000 M3 generation seeds were stored on time. The annual grafted seedling production capacity reached 150,000 plants. Example

[0011] A 50-mu pilot plantation will be established in Dongtai, Yancheng, and Yancheng Vocational College of Agriculture, including a 5-mu comparison area for single-antibody vs. double-antibody crops. Fifty local key personnel will be trained, each leading 10 households to form a demonstration network of 500 households. The leading enterprise, Fu'an Silk, has committed to purchasing 500,000 plants over three years at a price of 10 yuan per plant, generating a direct output value of 5 million yuan.

[0012] The integrated method provided by this invention can establish a pilot-scale demonstration on 50 mu (approximately 3.3 hectares) and a demonstration scale of 500 households within 3 years, achieving a seedling supply capacity of 150,000 seedlings per year. The obtained "dual-resistant rootstock and scion" grafted trees showed a mortality rate ≥15% lower under herbicide stress compared to single-resistant trees, thus constructing a dual biosafety barrier for the Jiangsu sericulture industry and possessing clear industrial application value.

Claims

1. A method for creating and classifying a new mulberry germplasm resistant to herbicides, characterized in that Comprising: (1) Irradiate the preferred mulberry seeds with an electron beam at an energy of 6-12 MeV to an absorbed dose of 150-250 Gy to obtain a M2 generation variation population; (2) Apply glyphosate or glufosinate stress to the variation population, and combine KASP molecular marker genotypic assisted selection to screen out stable resistant single plants; (3) Based on root biomass and leaf resistance index, the resistant single plants are divided into: rootstock type materials with root biomass ≥ 130% of the control, and scion type materials with leaf resistance index ≥ 2.

0.

2. The method of claim 1, wherein: The absorbed dose is 180 Gy, and the seedling rate of mulberry seeds is 48-50%.

3. A method for optimizing the combination of the anti-herbicide mulberry "double resistant graft" characterized by Comprising: (1) Provide at least 4 copies of the rootstock type material of claim 1 and at least 4 copies of the scion type material; (2) Construct a combination matrix of ≥ 1260 groups, and perform complete diallel grafting on the materials, wherein the rootstock selects 30 main parts and 14 alternatives, and the scion selects 28 main parts and 14 alternatives; (3) Perform two-stage screening: the first stage measures callus formation rate 15 days after grafting, and eliminates combinations with a formation rate < 80%; the second stage applies herbicide stress to the remaining combinations, and screens out optimized combinations with a mortality rate ≥ 15% lower than the single-antibody control; (4) Obtain 2-3 sets of "rootstock-scion double-resistant" optimized combinations with a grafting survival rate ≥ 85%.

4. The method of claim 3, wherein: The single-antibody control is formed by grafting a susceptible scion on the rootstock type material, or grafting a susceptible rootstock on the scion type material (the "susceptible" in the present invention refers to a conventional mulberry variety that has not undergone resistance screening and exhibits a sensitive phenotype under herbicide stress, and is used as a control material for resistance evaluation).

5. The method of claim 3, wherein: The elimination rate of the first-stage screening is controlled to be ≤ 30%.

6. A risk prevention and control method for rapid propagation of mulberry germplasm resistant to herbicides, characterized in that Comprising: (1) Establish an "main part + 2 alternatives" independent strain backup for each core germplasm; (2) Adopt a parallel strategy of tissue culture and cutting: the rootstock type material is mainly cut, and the scion type material is mainly tissue cultured with a multiplication coefficient ≥ 6.0 and annual subculture for 5 times; (3) Construct a cloud data platform to synchronize the rapid propagation progress in Yancheng-Hainan-Sanya in real time, and set a delay threshold of ≥ 7 days to trigger an early warning; (4) Achieve annual production of qualified grafted seedlings ≥ 150,000 plants.

7. The method of claim 6, wherein: The cloud data platform integrates weather monitoring, radiation progress uploading, and production data scanning and entering functions.

8. The method of claim 6, wherein: When the early warning is triggered, the AB personnel are required to be on duty within 48 hours.

9. The resistant material or optimized combination obtained by the method of any one of claims 1-8.