Artificial combination module and use thereof in improving stress resistance and herbicide resistance of plants
By constructing artificial composite modules using synthetic biology methods, the problems of insufficient plant resistance to drought, salt, high temperature, and herbicides have been solved. This has enabled plants to achieve growth advantages and herbicide resistance in extreme environments, thereby reducing agricultural production costs.
Patent Information
- Authority / Receiving Office
- WO · WO
- Patent Type
- Applications
- Current Assignee / Owner
- THE INST OF BIOTECHNOLOGY OF THE CHINESE ACAD OF AGRI SCI
- Filing Date
- 2025-05-09
- Publication Date
- 2026-07-02
AI Technical Summary
Existing technologies cannot simultaneously improve plants' resistance to drought, salt, high temperatures, and herbicides. The use of herbicides increases agricultural production costs and fails to fundamentally solve the problem of competition between weeds and crops.
An artificial composite module containing stress resistance and herbicide resistance functional modules was designed and constructed. It was then inserted into plants using synthetic biology techniques to form a functional module with dual functions of stress resistance and herbicide resistance. This involved specific nucleotide sequences and promoter designs.
It significantly improves the drought resistance, salt tolerance, and high temperature tolerance of plants, while also enabling them to resist herbicide stress and reducing agricultural production costs.
Smart Images
Figure CN2025093981_02072026_PF_FP_ABST
Abstract
Description
An artificial modular design and its application in improving plant stress resistance and herbicide resistance.
[0001] Cross-references to related applications
[0002] This application claims priority to Chinese Patent Application No. 202411919496.2, filed on December 25, 2024, the entire contents of which are incorporated herein by reference. Technical Field
[0003] This disclosure relates to the field of biotechnology, and more specifically, to an artificial composite module and its application in improving plant stress resistance and herbicide resistance. Background Technology
[0004] Soil salinization, frequent droughts, and prolonged periods of high temperatures are among the most destructive abiotic stresses on global agriculture, significantly reducing agricultural productivity through adverse effects on seed germination, plant growth and development, plant vigor, and crop yield. Conservative estimates suggest that years with severe adverse weather conditions, such as early spring and late autumn droughts and frequent, sustained high temperatures in summer, can even lead to total crop failure. However, weeds are highly adaptable, and extreme weather provides them with more space and resources to grow. Weeds with good tolerance to extreme weather can further encroach on crop space.
[0005] Currently, the main approach to addressing the impact of extreme weather on agriculture is through physical or chemical methods to increase crop yields. This includes applying organic fertilizers, improving soil structure, spraying phytostimulants to enhance plant tolerance, and using herbicides to control weed growth. However, the application of herbicides can also negatively impact crop growth. The extensive use of herbicides increases agricultural production costs and fails to fundamentally solve the existing problems.
[0006] Using synthetic biology techniques to construct artificial modular units to enhance plant stress resistance is a widely used strategy in agriculture. Currently, artificial modular units only target drought resistance, salt tolerance, and high temperature tolerance. To further promote agricultural development, it is an urgent problem to be solved to provide an artificial modular unit that can resist herbicides, as well as drought, salt, and high temperature. Summary of the Invention
[0007] The purpose of this disclosure is to improve the stress resistance and herbicide resistance of plants.
[0008] To achieve the above objectives, the first aspect of this disclosure provides an artificial combination module, the nucleotide sequence of which is shown in SEQ ID NO.1.
[0009] Optionally, the artificial combination module is equipped with a stress resistance module and a herbicide resistance module;
[0010] The nucleotide sequence of the stress-resistant functional module is positions 1-6674 of SEQ ID NO.1;
[0011] The nucleotide sequence of the herbicide-resistant functional module is positions 6710-11673 of SEQ ID NO.1.
[0012] Optionally, the stress-resistant functional module includes a first promoter, an RBS binding site, a first spacer sequence, a SipA protein-coding gene, a second spacer sequence, a DosH protein-coding gene, and a first terminator connected sequentially from upstream to downstream.
[0013] Wherein, the first promoter is the Gmubi promoter; the RBS binding site is rpiL.
[0014] Optionally, the herbicide-resistant functional module includes a second promoter, a third spacer sequence, a GR79 gene, a fourth spacer sequence, a GAT gene, a fifth spacer sequence, a second terminator, a sixth spacer sequence, a third promoter, a seventh spacer sequence, a DICX4 gene, an eighth spacer sequence, and a third terminator connected sequentially from upstream to downstream.
[0015] The second promoter is the 2xpCaMV35S promoter; the third promoter is the p-FMV promoter.
[0016] The second aspect of this disclosure provides the application of the artificial combination module described in the first aspect in improving the stress resistance and herbicide resistance of plants.
[0017] Optionally, improving the plant's stress resistance and herbicide resistance means improving the plant's stress resistance and herbicide resistance during plant variety selection.
[0018] Optionally, the improvement of plant stress resistance can be achieved by enhancing the plant cells' ability to resist drought, tolerate high salt, and withstand high temperatures.
[0019] The third aspect of this disclosure provides a recombinant expression vector, wherein the recombinant expression vector is an artificial combination module as described in the first aspect inserted into a backbone vector.
[0020] The fourth aspect of this disclosure provides a transformant in which the artificial combination module described in the first aspect or the recombinant expression vector described in the third aspect is inserted.
[0021] The fifth aspect of this disclosure provides a method for improving the stress resistance and herbicide resistance of plants, the method comprising:
[0022] The recombinant expression vector described in the third aspect is introduced into the target plant and / or the seed of the target plant;
[0023] Alternatively, the transformant described in the fourth aspect may be used to infect the target plant and / or the target plant seeds.
[0024] Through the above technical solution, the artificial composite module disclosed herein includes a stress-resistance functional module and a herbicide-resistance functional module. This disclosure utilizes biosynthesis technology to combine the stress-resistance functional module and the herbicide-resistance functional module to assemble a functional module with both stress-resistance and herbicide-resistance functions. It shows promising application prospects in cultivating plants with both stress-resistance and herbicide-resistance capabilities.
[0025] Other features and advantages of this disclosure will be described in detail in the following detailed description section. Attached Figure Description
[0026] The accompanying drawings are provided to further illustrate the present disclosure and form part of the specification. They are used together with the following detailed description to explain the present disclosure, but do not constitute a limitation thereof. In the drawings:
[0027] Figure 1 is a diagram of the carrier construction of the artificial combination module disclosed herein.
[0028] Figure 2 is a construction diagram of the artificial combination module disclosed herein.
[0029] Figure 3 shows a picture of the genetically modified rapeseed plant that is tolerant to high salt and drought.
[0030] Figure 4 is a picture of the herbicide-resistant genetically modified rapeseed plant disclosed in this paper.
[0031] Figure 5 is a picture of the transgenic maize plant that is resistant to high salt and drought disclosed in this paper. Detailed Implementation
[0032] The specific embodiments of this disclosure will be described in detail below with reference to the accompanying drawings. It should be understood that the specific embodiments described herein are for illustration and explanation only and are not intended to limit this disclosure.
[0033] The first aspect of this disclosure provides an artificial combinatorial module, the nucleotide sequence of which is shown in SEQ ID NO.1.
[0034] The artificially combined module disclosed herein incorporates a stress-resistance functional module and a herbicide-resistance functional module; wherein the nucleotide sequence of the stress-resistance functional module is positions 1-6674 of SEQ ID NO.1; and the nucleotide sequence of the herbicide-resistance functional module is positions 6710-11673 of SEQ ID NO.1. The stress-resistance functional module and the herbicide-resistance functional module are linked by the sequence shown in positions 6675-6709 of SEQ ID NO.1.
[0035] This disclosure utilizes modern synthetic biology design methods to optimize and modify stress-resistance elements. Through the artificial design of protein functional elements, tissue-specific promoter design, and stress response design, herbicide-resistant modules that specifically respond to different types of herbicides and stress-resistance modules that respond to different abiotic stresses are artificially constructed. These two modules are assembled into an intelligent, responsive, and directed artificial composite module with dual functions of stress resistance and herbicide resistance, named EDDICW. The total length of the artificial composite module disclosed in this disclosure is 11673 bp. It has good application prospects in cultivating plants with both stress resistance and herbicide resistance. Stress resistance refers to the ability to withstand drought, high salinity, and high temperatures.
[0036] In one embodiment of this disclosure, the stress-resistant functional module includes a first promoter, an RBS binding site, a first spacer sequence, a SipA protein-coding gene, a second spacer sequence, a DosH protein-coding gene, and a first terminator connected sequentially from upstream to downstream.
[0037] Wherein, the first promoter is the Gmubi promoter; the RBS binding site is rpiL.
[0038] According to this disclosure, the nucleotide sequence of the Gmubi promoter is positions 1-1416 of SEQ ID NO.1, the nucleotide sequence of the RBS binding site rpiL is positions 1417-1785 of SEQ ID NO.1, the nucleotide sequence of the first spacer sequence is positions 1786-1851 of SEQ ID NO.1, the nucleotide sequence of the SipA protein-coding gene is positions 1852-5355 of SEQ ID NO.1, the nucleotide sequence of the second spacer sequence is positions 5356-5421 of SEQ ID NO.1, the nucleotide sequence of the DosH protein-coding gene is positions 5422-6318 of SEQ ID NO.1, and the nucleotide sequence of the first terminator is positions 6319-6674 of SEQ ID NO.1.
[0039] In one embodiment of this disclosure, the herbicide-resistant functional module includes a second promoter, a third spacer sequence, a GR79 gene, a fourth spacer sequence, a GAT gene, a fifth spacer sequence, a second terminator, a sixth spacer sequence, a third promoter, a seventh spacer sequence, a DICX4 gene, an eighth spacer sequence, and a third terminator connected sequentially from upstream to downstream.
[0040] The second promoter is the 2xpCaMV35S promoter; the third promoter is the p-FMV promoter.
[0041] According to this disclosure, the nucleotide sequence of the 2xpCaMV35S promoter is positions 6710-7489 of SEQ ID NO.1, the nucleotide sequence of the third spacer sequence is positions 7490-7496 of SEQ ID NO.1, the nucleotide sequence of the GR79 gene is positions 7497-9065 of SEQ ID NO.1, the nucleotide sequence of the fourth spacer sequence is positions 9066-9128 of SEQ ID NO.1, the nucleotide sequence of the GAT gene is positions 9129-9569 of SEQ ID NO.1, the nucleotide sequence of the fifth spacer sequence is positions 9570-9575 of SEQ ID NO.1, the nucleotide sequence of the second terminator is positions 9576-9750 of SEQ ID NO.1, the nucleotide sequence of the sixth spacer sequence is positions 9751-9758 of SEQ ID NO.1, and the nucleotide sequence of the p-FMV promoter is... The nucleotide sequence of the seventh spacer sequence is SEQ ID NO. 1, positions 9759-10322; the nucleotide sequence of the DICX4 gene sequence is SEQ ID NO. 1, positions 10323-10326; the nucleotide sequence of the eighth spacer sequence is SEQ ID NO. 1, positions 11413-11420; and the nucleotide sequence of the third terminator sequence is SEQ ID NO. 1, positions 11421-11673.
[0042] The second aspect of this disclosure provides the application of the artificial combination module described in the first aspect in improving the stress resistance and herbicide resistance of plants.
[0043] In one embodiment of this disclosure, improving the plant's stress resistance and herbicide resistance involves enhancing these abilities during plant variety selection. The plant can be selected from corn, rapeseed, etc., preferably corn and / or rapeseed.
[0044] In one embodiment of this disclosure, the improvement of plant stress resistance refers to improving the drought resistance, high salt tolerance, and high temperature tolerance of plant cells.
[0045] In this disclosure, the herbicides included in the herbicide resistance list are glyphosate and / or dicamba. High salt tolerance refers to a salt concentration of 300 mM; high temperature tolerance refers to a temperature range of 30°C to 45°C.
[0046] The third aspect of this disclosure provides a recombinant expression vector, wherein the recombinant expression vector is an artificial combination module as described in the first aspect inserted into a backbone vector.
[0047] According to this disclosure, the skeleton carrier can be pJET, pBI-121, or PLUS.
[0048] In this disclosure, a recombinant expression vector is constructed to form expression frames for stress-resistance genes (i.e., SipA protein-coding genes and DosH protein-coding genes) and herbicide-resistance genes (i.e., GR79 genes, GAT genes and DICX4 genes), thereby enabling the expression of stress-resistance genes and herbicide-resistance genes in target plants, and thus enabling the target plants to possess stress-resistance and herbicide-resistance functions.
[0049] The fourth aspect of this disclosure is a transformant in which the artificial combination module described in the first aspect or the recombinant expression vector described in the third aspect is inserted.
[0050] According to this disclosure, the host of the transformant is Agrobacterium tumefaciens EHA105.
[0051] The fifth aspect of this disclosure provides a method for improving the stress resistance and herbicide resistance of plants, the method comprising:
[0052] The recombinant expression vector described in the third aspect is introduced into the target plant and / or the seed of the target plant;
[0053] Alternatively, the transformant described in the fourth aspect may be used to infect the target plant and / or the target plant seeds.
[0054] In this disclosure, the target plant can be infected with the callus tissue of the target plant.
[0055] The present disclosure will be further illustrated by the following examples, but the present disclosure is not limited thereto.
[0056] Unless otherwise specified, the raw materials, reagents, instruments and equipment involved in the embodiments of this disclosure can all be obtained by purchase.
[0057] Unless otherwise specified, the experimental conditions in this disclosure are conventional conditions well known to those skilled in the art, such as those described in Sambrook et al. Molecular Cloning: A Laboratory Manual (New York: Cold Spring Harbor Laboratory Press, 1989), or those recommended by the manufacturer.
[0058] Cloning vector pJET: a commercially available product from ThermoFisher.
[0059] Shuttle carrier: pBI-121: preserved in this laboratory;
[0060] Agrobacterium tumefaciens EHA105: preserved in this laboratory.
[0061] Example 1
[0062] This embodiment illustrates the preparation of recombinant Agrobacterium tumefaciens using an artificially combined module:
[0063] I. Experimental Methods:
[0064] As shown in Figures 1 and 2, different types of herbicide-resistant functional modules were designed using synthetic biology. These modules, which specifically respond to different abiotic stresses, were then assembled to form a novel stress-resistance functional circuit with intelligent, directed expression, named EGDICW (i.e., the artificially synthesized module). The full-length nucleic acid sequence of the stress-resistance functional circuit EGDICW, measuring 11673 bp, was obtained using artificial chemical synthesis. It was cloned into the vector pJET, constructing the recombinant cloning plasmid pJET-EGDICW containing the complete stress-resistance functional circuit, and sequenced for verification.
[0065] Then, the stress resistance circuit EGDICW fragment containing sticky ends and the shuttle vector pBI-121 fragment were obtained by double digestion with EcoRI and HindIII. The stress resistance circuit EGDICW was ligated into the pBI-121 vector to construct the plant expression vector pBI-EGDICW. This expression vector was transformed into Agrobacterium tumefaciens EHA105. Positive recombinant strains were screened for kanamycin antibiotic resistance and verified by colony PCR sequencing.
[0066] II. Experimental Results:
[0067] The full-length nucleic acid sequence of the stress-resistant functional circuit EDDICW, with a length of 11673 bp, was obtained by artificial chemical synthesis.
[0068] The plant expression vector pBI-EGDICW containing the functional circuit EGDICW was successfully constructed and transformed into Agrobacterium tumefaciens EHA105.
[0069] After PCR, enzyme digestion, and sequencing verification confirmed the correctness of the inserted sequence, the strain was named EHA-EGDICW.
[0070] III. Experimental Conclusions:
[0071] The recombinant Agrobacterium tumefaciens EHA-EGDICW expressing the stress resistance functional circuit EGDICW was successfully constructed.
[0072] Example 2
[0073] This example illustrates the construction of genetically modified rapeseed.
[0074] The target plant used in this embodiment is Brassica napus, and the rapeseed seeds 84100-18 were preserved in our laboratory.
[0075] I. Experimental Methods:
[0076] Rapeseed seeds were disinfected by soaking in 75% ethanol and 0.1% HgCl2, respectively, and then evenly placed on plant tissue culture medium and cultured in a tissue culture room at 24℃ for one week. Hypocotyls of rapeseed seedlings were surgically cut with sterilization and placed on pre-culture medium, and cultured under light for 2-3 days to pre-culture explants.
[0077] The recombinant Agrobacterium strain EHA-EGDICW, which activates the stress resistance circuitry, was transferred, and the strain was collected by centrifugation and resuspended in OD240. 600 =1.0. The pre-cultured explants were immersed in Agrobacterium bacterial suspension for 90 seconds, air-dried, and then transferred to a co-culture medium for dark incubation for 2-3 days. Subsequently, well-grown explants were transferred to induction medium for further culture.
[0078] Explants with good callus growth were selected and transferred to MS medium supplemented with KANA antibiotic for selection. After 45-50 days of light cultivation, shoots differentiated. The differentiated callus was then transferred to MS rooting medium and cultured under light for 2 weeks. Once roots appeared and stems reached 4-5 cm in length, the seedlings were transferred to potting soil for hardening-off. After acclimatization, they were transplanted to a greenhouse, and PCR testing confirmed positive rapeseed seedlings.
[0079] II. Experimental Results:
[0080] Using Agrobacterium-mediated explant co-culture, the stress-resistance functional circuit EGDICW was transformed into rapeseed. After infecting rapeseed explants and undergoing induction culture, screening culture, rooting culture and hardening-off transplanting, the transgenic rapeseed Bn-EGDICW expressing the stress-resistance functional circuit was finally obtained through PCR verification. It can be used for subsequent stress resistance performance research.
[0081] III. Experimental Conclusions:
[0082] Through Agrobacterium-mediated transformation, the stress-resistant functional circuit EGDICW was finally obtained in rapeseed Bn-EGDICW.
[0083] Example 3
[0084] This embodiment is used to illustrate the stress resistance and herbicide resistance of rapeseed Bn-EGDICW in Example 2.
[0085] The experimental group used the transgenic rapeseed (i.e., transgenic rapeseed seeds) obtained in Example 2; the control group used non-transgenic rapeseed (i.e., wild-type seeds).
[0086] I. Experimental Methods:
[0087] Salt stress was simulated by adding NaCl; drought stress was simulated by adding polyethylene glycol PEG-6000; and herbicide stress was simulated by a mixture of glyphosate and dicamba.
[0088] The obtained transgenic rapeseed seeds that have been identified as positive were cultured with wild-type seeds in MS solid culture. After the seedlings grew true leaves, they were transplanted into plastic pots containing substrate and watered with MS nutrient solution. When the seedlings grew 5-6 true leaves, they were subjected to stress treatment.
[0089] The high salt stress treatment included watering the plants with 20 mL of salt solution at a concentration of 300 mM every day.
[0090] Drought stress treatment included watering the plants daily with 20 mL of 15% polyethylene glycol.
[0091] Herbicide stress includes spraying a commercially available mixture of glyphosate and dicamba (purchased from Weifang Zhongnong United Chemical Co., Ltd.) at the four-leaf stage of rapeseed plants, using the concentration as directed in the instructions.
[0092] II. Experimental Results:
[0093] The stress results are shown in Figures 3 and 4. The results show that under normal growth conditions before treatment, the transgenic rapeseed Bn-EGDICW had no difference in growth status compared with wild-type rapeseed, and its agronomic traits were not affected.
[0094] In Figure 3, the left side shows the growth status of non-GMO rapeseed in saline-alkali soil on day 14, and the right side shows the growth status of GMO rapeseed in saline-alkali soil on day 14. Under 15% severe drought stress for 7 days, wild-type rapeseed began to show yellowing, leaf drop, and wilting phenotypes, while the growth rate of GMO rapeseed Bn-EGDICW slowed down, and leaf and stem growth were not significantly affected. After 14 days of drought treatment, wild-type rapeseed had completely died, while GMO rapeseed began to show some wilting. In the high salt stress experiment, after 7 days of 300mM NaCl stress treatment, wild-type rapeseed showed severe dehydration and drying, while some leaves of GMO rapeseed Bn-EGDICW turned yellow, and its growth was significantly better than that of wild-type. After 14 days of high salt treatment, wild-type rapeseed had basically dried up and died, while GMO rapeseed Bn-EGDICW showed leaf curling, stem wilting, and slowed growth.
[0095] In Figure 4, the left side shows the growth status of non-GMO rapeseed 14 days after herbicide spraying, and the right side shows the growth status of GMO rapeseed 14 days after herbicide spraying. When treated with a mixture of glyphosate and dicamba, wild-type rapeseed completely died, while the GMO rapeseed Bn-EGDICW was resistant to both herbicides.
[0096] III. Experimental Conclusions:
[0097] The stress-resistance genes in the artificially combined modules disclosed herein were successfully expressed in rapeseed, significantly improving the rapeseed plants' tolerance to high salt, drought, and herbicide resistance.
[0098] Example 4
[0099] This example illustrates the construction of genetically modified maize.
[0100] I. Experimental Methods:
[0101] Corn seeds were peeled, sterilized by soaking in 75% ethanol and 0.1% HgCl2, and then evenly placed on plant tissue culture medium and cultured in a tissue culture room at 24°C for 2 weeks. Rice callus tissue was surgically extracted using sterile cuttings, placed on pre-culture medium, and cultured in the dark for 2 weeks.
[0102] The recombinant Agrobacterium strain EHA-EGDICW, which activates the expression of the stress resistance circuit, was transferred and collected by centrifugation, then resuspended to OD600 = 1.0. The pre-cultured explants were immersed in Agrobacterium bacterial suspension for 30 min, air-dried, and then transferred to co-culture medium and incubated in the dark for 2-3 days. Subsequently, they were transferred to induction medium for further culture.
[0103] Callus tissue was selected and transferred to MS selection medium supplemented with KANA antibiotic, and cultured in the dark for 2 weeks. A second screening was performed, followed by another 2 weeks of dark culture, and then differentiation culture for 1 week. The differentiated callus tissue was transferred to MS rooting medium. Once roots appeared and the stems grew to 4-5 cm, the seedlings were transferred to a greenhouse, and PCR testing was conducted on positive maize seedlings.
[0104] II. Experimental Results:
[0105] Using Agrobacterium-mediated callus co-culture, the stress-resistance functional circuit EGDICW was transformed into maize. After infecting maize callus and undergoing induction culture, resistance screening culture, rooting culture, and seedling transplantation, the transgenic maize Co-EGDICW expressing the stress-resistance functional circuit was finally obtained through PCR verification. It can be used for subsequent stress resistance performance research.
[0106] III. Experimental Conclusions:
[0107] Through Agrobacterium-mediated transformation, Co-EGDICW maize with the stress-resistant functional line EGDICW was finally obtained.
[0108] Example 5
[0109] This example illustrates the stress resistance and herbicide resistance of the corn Co-EGDICW from Example 4.
[0110] The experimental group used transgenic maize (i.e., transgenic rapeseed) obtained in Example 4; the control group used non-transgenic maize (i.e., wild-type seeds).
[0111] I. Experimental Methods:
[0112] Salt stress was simulated by adding NaCl; drought stress was simulated by adding polyethylene glycol PEG-6000; and herbicide stress was simulated by a mixture of glyphosate and dicamba.
[0113] Stress treatment was applied by irrigation. Wild-type maize and positive transgenic maize Co-EGDICW seeds were germinated and cultured into seedlings in MS medium. When the maize seedlings reached the stage of two leaves and one heart, stress treatment was applied approximately 12 days later, and the plant growth status was observed.
[0114] The high salt stress treatment included watering the plants with 20 mL of salt solution at a concentration of 300 mM every day.
[0115] Drought stress treatment included watering the plants daily with 20 mL of 15% polyethylene glycol.
[0116] II. Experimental Results:
[0117] The stress results are shown in Figure 5. The study showed that under no stress conditions, the emergence and growth of transgenic maize Co-EGDICW were no different from those of wild-type rice.
[0118] In Figure 5, the right side shows the growth status of non-GMO corn in saline-alkali soil on the 7th day, and the left side shows the growth status of GMO corn in saline-alkali soil on the 7th day.
[0119] Under 15% severe drought stress for 7 days, wild-type maize began to show signs of yellowing, leaf drop, and wilting. The growth rate of transgenic maize Co-EGDICW slowed, but leaf and stem growth were not significantly affected. In the high salt stress experiment, after 7 days of treatment with 300mM NaCl, wild-type maize showed severe dehydration and drying, while some leaves of the transgenic maize Co-EGDICW turned yellow, and its growth was significantly better than that of the wild type.
[0120] III. Experimental Conclusions:
[0121] The stress-resistance genes in the artificially combined modules disclosed herein were successfully expressed in maize, significantly improving the high salt tolerance and drought resistance of maize plants.
[0122] The preferred embodiments of this disclosure have been described in detail above with reference to the accompanying drawings. However, this disclosure is not limited to the specific details of the above embodiments. Within the scope of the technical concept of this disclosure, various simple modifications can be made to the technical solutions of this disclosure, and these simple modifications all fall within the protection scope of this disclosure.
[0123] It should also be noted that the various specific technical features described in the above specific embodiments can be combined in any suitable manner without contradiction. In order to avoid unnecessary repetition, this disclosure will not describe the various possible combinations separately.
[0124] Furthermore, various different embodiments of this disclosure can be combined in any way, as long as they do not violate the spirit of this disclosure, they should also be regarded as the content disclosed in this disclosure.
Claims
1. An artificial assembly module, characterized in that, The nucleotide sequence of the artificially combined module is shown in SEQ ID NO.
1.
2. The artificial assembly module according to claim 1, wherein, The artificial combination module is equipped with a stress-resistance module and a herbicide-resistance module. The nucleotide sequence of the stress-resistant functional module is positions 1-6674 of SEQ ID NO.1; The nucleotide sequence of the herbicide-resistant functional module is positions 6710-11673 of SEQ ID NO.
1.
3. The artificial assembly module according to claim 2, wherein, The stress-resistant functional module includes a first promoter, an RBS binding site, a first spacer sequence, a SipA protein-coding gene, a second spacer sequence, a DosH protein-coding gene, and a first terminator, connected sequentially from upstream to downstream. Wherein, the first promoter is the Gmubi promoter; the RBS binding site is rpiL.
4. The artificial assembly module according to claim 2, wherein, The herbicide-resistant functional module includes, from upstream to downstream, a second promoter, a third spacer sequence, a GR79 gene, a fourth spacer sequence, a GAT gene, a fifth spacer sequence, a second terminator, a sixth spacer sequence, a third promoter, a seventh spacer sequence, a DICX4 gene, an eighth spacer sequence, and a third terminator. The second promoter is the 2xpCaMV35S promoter; the third promoter is the p-FMV promoter.
5. The application of the artificial combination module according to any one of claims 1-4 in improving the stress resistance and herbicide resistance of plants.
6. The application according to claim 5, wherein, The improvement of plant stress resistance and herbicide resistance refers to enhancing the plant's stress resistance and herbicide resistance during plant variety selection.
7. The application according to claim 6, wherein, The improvement of plant stress resistance refers to enhancing the plant cells' ability to resist drought, tolerate high salt, and withstand high temperatures.
8. A recombinant expression vector, characterized in that, The recombinant expression vector is an artificial combination module as described in any one of claims 1-4 inserted into a backbone vector.
9. A transformant, characterized in that, The transformant contains an artificial combination module as described in any one of claims 1-4 or a recombinant expression vector as described in claim 8.
10. A method for improving the stress resistance and herbicide resistance of plants, characterized in that, The method includes: The recombinant expression vector according to claim 8 is introduced into the target plant and / or the seed of the target plant; Alternatively, the transformant as described in claim 9 may be used to infect the target plant and / or the target plant seeds.