Maize plant height regulation gene combination and application thereof in creating multi-gradient plant height maize material
Patent Information
- Application Number
- CN202511838945.5
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-12-08
- Publication Date
- 2026-08-28
AI Technical Summary
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[0025] 1) The three-gene aggregation families of eight different plant height genotypes obtained by this invention effectively ensure gene diversity, avoid the problem of single gene failure, and provide a possibility for the creation of ideal maize plant type.
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Abstract
Description
Technical Field
[0001] This invention belongs to the field of crop molecular breeding technology, specifically relating to a combination of maize plant height regulating genes and its application in creating multi-gradient maize materials. Background Technology
[0002] Maize (Zea mays L.) belongs to the genus Zea in the family Poaceae.
[0003] Molecular marker-assisted selection (MAS) is highly efficient and precise in maize breeding and is widely used by researchers. Combining MAS with conventional breeding techniques can achieve efficient aggregation of multiple genes in a single material. However, this is not yet possible. Therefore, a gene combination for regulating maize plant height and its application in creating maize materials with multi-gradient plant heights is urgently needed. Summary of the Invention
[0004] To address the shortcomings of existing technologies, this invention provides a combination of maize plant height regulating genes and its application in creating maize materials with multiple plant height gradients.
[0005] To solve the above-mentioned technical problems, the present invention provides the following technical solution:
[0006] The first objective of this invention is to provide a combination of maize plant height regulating genes, including genes ZmCPS1, ZmbHLH156, and ZmEMF1L1.
[0007] Preferably, the gene ZmCPS1 has an allele AA and an allele aa, the nucleotide sequence of the allele AA is shown in SEQ ID NO. 10, and the nucleotide sequence of the allele aa is shown in SEQ ID NO. 11;
[0008] The gene ZmbHLH156 has alleles BB and bb, the nucleotide sequence of which is shown in SEQ ID NO. 12 and the nucleotide sequence of which is shown in SEQ ID NO. 13.
[0009] The gene ZmEMF1L1 has an allele CC and an allele cc. The nucleotide sequence of the allele CC is shown in SEQ ID NO. 14, and the nucleotide sequence of the allele cc is shown in SEQ ID NO. 15.
[0010] The second objective of this invention is to provide an application of a combination of maize plant height regulating genes in creating maize materials with multiple plant height gradients. This application creates maize materials with different plant heights by combining any three alleles from genes ZmCPS1, ZmbHLH156, and ZmEMF1L1.
[0011] Preferably, it includes the following steps:
[0012] S1. Using homozygous near-isogenin lines carrying a single gene regulating plant height as the base material, the base materials include A-NIL, B-NIL and C-NIL. Any two base materials are crossed, and the hybrid F1 is obtained by bagging.
[0013] S2. After dividing the F1 generation into different lines, the other basic material that was not selected above is crossed with the F1 generation, and the hybrid F2 is harvested by bagging.
[0014] S3. After dividing the F2 generation into different lines, MAS molecular marker-assisted breeding technology was used to screen out single plants with heterozygous genotypes for all three target genes. The plants were then bagged and self-crossed to obtain hybrid F3.
[0015] S4. After planting F3 generations of separate lines, MAS molecular marker-assisted breeding technology was used to screen out 8 genotypes of single plants that were homozygous for all three genes, namely AABBcc, AABBCC, AAbbcc, aaBBcc, AAbbCC, aaBBCC, aabbcc, and aabbCC. These plants were bagged and harvested from cross F4, thus obtaining maize materials with plant height gradients and a background recovery rate of over 90%.
[0016] Preferably, in step S1, A-NIL represents a pair of basic materials carrying genotypes AAbbcc or aabbcc, B-NIL represents a pair of basic materials carrying genotypes aaBBcc or aabbcc, and C-NIL represents a pair of basic materials carrying genotypes aabbCC or aabbcc.
[0017] The AA represents the allele AA of the gene ZmCPS1, the aa represents the allele aa of the gene ZmCPS1, the nucleotide sequence of the allele AA is shown in SEQ ID NO. 10, and the nucleotide sequence of the allele aa is shown in SEQ ID NO. 11.
[0018] The BB represents the allele BB of the gene ZmbHLH156, the bb represents the allele bb of the gene ZmbHLH156, the nucleotide sequence of the allele BB is shown in SEQ ID NO. 12, and the nucleotide sequence of the allele bb is shown in SEQ ID NO. 13;
[0019] CC represents the allele CC of the gene ZmEMF1L1, cc represents the allele cc of the gene ZmEMF1L1, the nucleotide sequence of the allele CC is shown in SEQ ID NO. 14, and the nucleotide sequence of the allele cc is shown in SEQ ID NO. 15.
[0020] Among them, alleles aa, bb, and CC are used to reduce maize plant height, while alleles AA, BB, and cc are used to increase maize plant height.
[0021] Preferably, the following steps are also included:
[0022] S5. To further eliminate background interference and obtain materials with a high background recovery rate, after planting F3 generations of different lines, MAS molecular marker-assisted breeding technology was used to screen out single plants with heterozygous corresponding genes. These plants were then continuously self-crossed to obtain F6. In the F6 generation, MAS molecular marker-assisted breeding technology was used to screen out single plants with 8 homozygous corresponding genes, thus obtaining another group of maize materials with plant height gradients, with a background recovery rate of over 95%.
[0023] Preferably, in steps S3-S5, the MAS molecular marker-assisted breeding technology identifies the genotype using KASP. The molecular marker for gene ZmCPS1 is CP702, and the nucleotide sequence of the primer for molecular marker CP702 is shown in SEQ ID NO. 1-3; the molecular marker for gene ZmbHLH156 is bH147, and the nucleotide sequence of the primer for molecular marker bH147 is shown in SEQ ID NO. 4-6; the molecular marker for gene ZmEMF1L1 is EM1728, and the nucleotide sequence of the primer for molecular marker EM1728 is shown in SEQ ID NO. 7-9.
[0024] Compared with the prior art, the present invention has the following beneficial effects:
[0025] 1) The three-gene aggregation families of eight different plant height genotypes obtained by this invention effectively ensure gene diversity, avoid the problem of single gene failure, and provide a possibility for the creation of ideal maize plant type.
[0026] 2) This invention can improve the efficiency of auxiliary selection in conventional breeding and save costs. By detecting plant height gene loci in maize plants using molecular markers, homozygous individual plants can be quickly identified at the seedling stage, allowing for the timely elimination of heterozygous individual plants, saving production costs, improving the efficiency of material selection, and shortening the breeding cycle of maize varieties.
[0027] 3) This invention uses a method of hybridization between near-isogenic single-gene lines, which can achieve rapid aggregation of multiple beneficial genes under the same genetic background, reduce linkage redundancy and genetic background reversion detection steps, and improve the selection efficiency of aggregation breeding. Attached Figure Description
[0028] Figure 1 A represents the plant height phenotype of the homozygous near-isogenic AA and aa lines of this invention;
[0029] Figure 1 ZhongB represents the plant height phenotype of the homozygous near-isogenic lines BB and bb of this invention;
[0030] Figure 1 C represents the plant height phenotype of the homozygous near-isogenic CC and cc lines of this invention;
[0031] Figure 2 This is the plant height phenotype of the ZmEMF1L1 knockout line of this invention;
[0032] Figure 3 This is a detailed technical roadmap for the specific implementation of this invention;
[0033] Figure 4 These are the plant height phenotypes of the eight genotype materials obtained in this invention. Detailed Implementation
[0034] The preferred embodiments of the present invention will be described below with reference to the accompanying drawings. It should be understood that the preferred embodiments described herein are for illustration and explanation only and are not intended to limit the present invention.
[0035] The homozygous near-isogenic lines carrying a single gene regulating plant height, used in the following examples with maize 9782 as the background, were the basic materials. These basic materials included A-NIL, B-NIL, and C-NIL. They were obtained by using maize 9782 as the recurrent parent and introducing single plant height genes—AA and aa genes being alleles of ZmCPS1, BB and bb genes being alleles of ZmbHLH156, and CC and cc genes being alleles of ZmEMF1L1—into 9782 through hybridization and backcrossing. AA, BB, and CC all represent genotypes from the parent By4944, while aa, bb, and cc all represent genotypes from the parent 9782. The method for constructing near-isogenic lines involves crossing 9782 × By4944 to the F1 generation, using 9782 as the recurrent parent, and continuously backcrossing to obtain BC4F1. In the self-crossed offspring BC4F2, MAS molecular marker-assisted breeding technology is used to screen for the corresponding single-gene heterozygous genotypes, and bagging and self-crossing are used to obtain BC4F3. In the BC4F3 generation, MAS molecular marker-assisted breeding technology is used to screen for the corresponding single-gene homozygous genotypes, and bagging and self-crossing are used to obtain BC4F4. This self-crossed species is a near-isogenic line with three genes.
[0036] The molecular marker-assisted breeding technology used in the following examples uses the same molecular marker information as KASP: the molecular marker for ZmCPS1 is CP702, and the primers for molecular marker CP702 are SEQ ID NO. 1 ~ 3; the molecular marker for ZmbHLH156 is bH147, and the primers for molecular marker bH147 are SEQ ID NO. 4 ~ 6; the molecular marker for ZmEMF1L1 is EM1728, and the primers for molecular marker EM1728 are SEQ ID NO. 7 ~ 9.
[0037] If the target base of the molecular marker CP702 is T, it indicates that the maize sample is of the ZmCPS1 allele 'aa' genotype; if the target base is C, it indicates that the maize sample is of the ZmCPS1 allele 'AA' genotype. If the target base of the molecular marker bH147 is G, it indicates that the maize sample is of the ZmbHLH156 allele 'bb' genotype; if the target base is A, it indicates that the maize sample is of the ZmbHLH156 allele 'BB' genotype. If the target base of the molecular marker EM1728 is G, it indicates that the maize sample is of the ZmEMF1L1 allele 'cc' genotype; if the target base is A, it indicates that the maize sample is of the ZmEMF1L1 allele 'CC' genotype.
[0038] Example 1: A method for creating multi-gradient maize materials through three-gene aggregation (steps as follows) Figure 3 (As shown).
[0039] Based on homozygous near-isogenic lines carrying a single gene regulating plant height, using maize 9782 as the background, the basic materials included A-NIL, B-NIL, and C-NIL, whose plant height phenotypes were as follows: Figure 1 The knockout phenotype of the ZmEMF1L1 gene is as follows: Figure 2 The knockout line has a reduced height. In this example, aaBBcc and aabbCC were crossed, and the hybrid F1 was obtained by bagging. 1， Ensure at least 20 hybrid seeds.
[0040] After dividing the F1 generation into different lines, in this example, AAbbcc and aaBbCc were crossed, and the hybrid F2 was harvested by bagging.
[0041] After dividing the F2 generation into lines, MAS molecular marker-assisted breeding technology was used to screen out single plants with heterozygous genotypes for all three target genes, and the hybrid F3 was obtained by bagging and self-crossing.
[0042] After F3 generation of separate plant lines, MAS molecular marker-assisted breeding technology was used to screen for eight genotypes of individual plants with homozygous corresponding to three genes: AABBcc, AABBCC, AAbbcc, aaBBcc, AAbbCC, aaBBCC, aabbcc, and aabbCC. These plants were bagged and harvested from the cross F4, thus obtaining the first group of maize materials with plant height gradients, whose phenotypes are as follows: Figure 4 The background response rate for this material is over 90%.
[0043] Example 2
[0044] To further eliminate background interference and obtain maize materials with plant height gradients and higher background recovery rates, after planting F3 generations of different lines, MAS molecular marker-assisted breeding technology was used to screen out single plants with heterozygous corresponding genes. These plants were then continuously self-crossed to obtain F6. In the F6 generation, MAS molecular marker-assisted breeding technology was used to screen out single plants with eight homozygous corresponding genes, thus obtaining the second group of maize materials with plant height gradients, with a background recovery rate of over 95%.
[0045] Finally, it should be noted that the above descriptions are merely preferred embodiments of the present invention and are not intended to limit the present invention. Although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art can still modify the technical solutions described in the foregoing embodiments or make equivalent substitutions for some of the technical features. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the protection scope of the present invention.
Claims
1. A combination of maize plant height regulating genes, characterized in that, This includes the genes ZmCPS1, ZmbHLH156, and ZmEMF1L1.
2. The maize plant height regulating gene combination according to claim 1, characterized in that, The gene ZmCPS1 has alleles AA and aa, the nucleotide sequence of allele AA is shown in SEQ ID NO. 10, and the nucleotide sequence of allele aa is shown in SEQ ID NO. 11; The gene ZmbHLH156 has alleles BB and bb, the nucleotide sequence of which is shown in SEQ ID NO. 12 and the nucleotide sequence of which is shown in SEQ ID NO.
13. The gene ZmEMF1L1 has an allele CC and an allele cc. The nucleotide sequence of the allele CC is shown in SEQ ID NO. 14, and the nucleotide sequence of the allele cc is shown in SEQ ID NO.
15.
3. The application of a maize plant height regulating gene combination as described in any one of claims 1-2 in creating multi-gradient maize materials, characterized in that, The application creates maize materials with different plant heights by aggregating any three alleles from the genes ZmCPS1, ZmbHLH156, and ZmEMF1L1.
4. The application according to claim 3, characterized in that, Includes the following steps: S1. Using homozygous near-isogenin lines carrying a single gene regulating plant height as the base material, the base materials include A-NIL, B-NIL and C-NIL. Any two base materials are crossed, and the hybrid F1 is obtained by bagging. S2. After dividing the F1 generation into different lines, the other basic material that was not selected above is crossed with the F1 generation, and the hybrid F2 is harvested by bagging. S3. After dividing the F2 generation into different lines, MAS molecular marker-assisted breeding technology was used to screen out single plants with heterozygous genotypes for all three target genes. The plants were then bagged and self-crossed to obtain hybrid F3. S4. After planting F3 generations of separate lines, MAS molecular marker-assisted breeding technology was used to screen out 8 genotypes of single plants that were homozygous for all three genes, namely AABBcc, AABBCC, AAbbcc, aaBBcc, AAbbCC, aaBBCC, aabbcc, and aabbCC. These plants were bagged and harvested from cross F4, thus obtaining maize materials with plant height gradients and a background recovery rate of over 90%.
5. The application according to claim 4, characterized in that, In step S1, A-NIL represents a pair of basic materials carrying genotypes AAbbcc or aabbcc, B-NIL represents a pair of basic materials carrying genotypes aaBBcc or aabbcc, and C-NIL represents a pair of basic materials carrying genotypes aabbCC or aabbcc. The AA represents the allele AA of the gene ZmCPS1, the aa represents the allele aa of the gene ZmCPS1, the nucleotide sequence of the allele AA is shown in SEQ ID NO. 10, and the nucleotide sequence of the allele aa is shown in SEQ ID NO.
11. The BB represents the allele BB of the gene ZmbHLH156, the bb represents the allele bb of the gene ZmbHLH156, the nucleotide sequence of the allele BB is shown in SEQ ID NO. 12, and the nucleotide sequence of the allele bb is shown in SEQ ID NO. 13; CC represents the allele CC of the gene ZmEMF1L1, cc represents the allele cc of the gene ZmEMF1L1, the nucleotide sequence of the allele CC is shown in SEQ ID NO. 14, and the nucleotide sequence of the allele cc is shown in SEQ ID NO.
15. Among them, alleles aa, bb, and CC are used to reduce maize plant height, while alleles AA, BB, and cc are used to increase maize plant height.
6. The application according to claim 5, characterized in that, It also includes the following steps: S5. To further eliminate background interference and obtain materials with a high background recovery rate, after planting F3 generations of different lines, MAS molecular marker-assisted breeding technology was used to screen out single plants with heterozygous corresponding genes. These plants were then continuously self-crossed to obtain F6. In the F6 generation, MAS molecular marker-assisted breeding technology was used to screen out single plants with 8 homozygous corresponding genes, thus obtaining another group of maize materials with plant height gradients, with a background recovery rate of over 95%.
7. The application according to claim 6, characterized in that, In steps S3-S5, the MAS molecular marker-assisted breeding technology identifies genotypes using KASP. The molecular marker for gene ZmCPS1 is CP702, and the nucleotide sequences of the primers for molecular marker CP702 are shown in SEQ ID NO. 1-3; the molecular marker for gene ZmbHLH156 is bH147, and the nucleotide sequences of the primers for molecular marker bH147 are shown in SEQ ID NO. 4-6; the molecular marker for gene ZmEMF1L1 is EM1728, and the nucleotide sequences of the primers for molecular marker EM1728 are shown in SEQ ID NO. 7-9.