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CN-121975819-A - Application of AGO10 gene in regulation and control of soybean branch number

CN121975819ACN 121975819 ACN121975819 ACN 121975819ACN-121975819-A

Abstract

The invention belongs to the technical field of biology, and particularly discloses application of an AGO10 gene in regulating and controlling soybean branch number, and double-strand break is carried out under the mediation of endonuclease Cas9 through an effective target point for editing the soybean AGO10 gene, so that AGO10a/b different allele mutants are obtained. The Gmago a/b mutant has significantly increased branching number compared to wild type soybean, resulting in increased total pod number per plant and increased yield per plant. Therefore, the AGO10 gene has good application prospect and application value in the aspect of creating high-yield soybeans. The knocking out of the AGO10 gene provides a new strategy for increasing the branch number of soybeans and also provides a reliable means and material for cultivating new varieties and improved plant types of soybeans.

Inventors

  • LU SIJIA
  • CHEN LEI
  • LIU BAOHUI
  • KONG FANJIANG

Assignees

  • 广州大学

Dates

Publication Date
20260505
Application Date
20260403

Claims (8)

  1. The application of the AGO10 gene in regulating and controlling the branch number of soybeans is characterized in that the AGO10 gene is an AGO10a gene and an AGO10b gene, the CDS sequence of the AGO10a gene is shown as SEQ ID NO.1, the CDS sequence of the AGO10b gene is shown as SEQ ID NO.2, and the regulation and control are shown as that the branch number of the soybeans is increased after the expression of the AGO10 gene is reduced.
  2. 2. The use according to claim 1, wherein the way to reduce the expression of the AGO10 gene is to knock out the AGO10 gene.
  3. 3. The use according to claim 2, characterized in that by constructing the CRISPR-Cas9 vector of the AGO10 gene, a knockout mutant strain is obtained with a significantly higher number of soybean branches than the wild type AGO10 gene.
  4. 4. The use according to claim 3, wherein the AGO10 knockout mutant strain has a significantly increased pitch, pod number, hundred and grain weight compared to the wild type.
  5. 5. A plant breeding method characterized in that the method is (1) or (2) below: (1) Obtaining plants with more branches, nodes, single-plant pod numbers, hundred-plant grain weights and single-plant grain weights than the target plants by reducing the activity of AGO10 protein in the target plants; (2) Obtaining plants with more branches, nodes, single-plant pod numbers, hundred-plant grain weights and single-plant grain weights than the target plants by reducing the expression of the AGO10 gene in the target plants; The target plant is soybean, the AGO10 gene is AGO10a and AGO10b genes, the CDS sequence of the AGO10a gene is shown as SEQ ID NO.1, the CDS sequence of the AGO10b gene is shown as SEQ ID NO.2, the AGO10 protein is AGO10a and AGO10b protein, the amino acid sequence of the AGO10a protein is shown as SEQ ID NO.3, and the amino acid sequence of the AGO10b protein is shown as SEQ ID NO. 4.
  6. 6. The plant breeding method according to claim 5, wherein the means for reducing the expression of the AGO10 gene is knocking out the AGO10 gene.
  7. 7. A method for increasing the number of soybean branches is characterized by knocking out an AGO10 gene, wherein the nucleotide sequence of the AGO10 gene is shown as SEQ ID NO. 1.
  8. 8. The plant breeding method according to claim 6 or the method for increasing soybean branching number according to claim 7, wherein the AGO10 knockout mutant strain is obtained by using agrobacteria-mediated CRISPR-Cas9 gene editing technology.

Description

Application of AGO10 gene in regulation and control of soybean branch number Technical Field The invention belongs to the technical field of biology, and particularly relates to application of an AGO10 gene in regulating and controlling soybean branch number, and more particularly relates to application of reducing expression of the AGO10 gene in increasing soybean branch number. Background Soybean (Glycine max (l.) merr.) branch number is an important quantitative trait related to soybean plant architecture, fitness and yield, directly affecting soybean pod number and seed number per plant. Under the condition of the stalk lodging problem, the plant lodging can be relieved by relatively reducing the plant density, and meanwhile, the labor and seed cost are reduced. Under the condition, the variety with more branches is selected, so that the defect of less sowing quantity can be overcome, and the yield can be increased. Increasing the number of branches is of great importance in soybean breeding to increase yield. The Argonaute protein (AGO) was first found in Arabidopsis. The silencing complex consists of a small RNA and AGO proteins, most of which are considered to have cleavage activity. Different AGO proteins exert different biological functions. AtAGO10 and AtAGO1 are used for jointly regulating and controlling the termination of flower stem cells, atAGO is used for mediating the activity of miR172, miR172 is targeted to inhibit AP2 so as to inhibit WUS expression and terminate flower stem cells, atAGO and AtAGO1 can be combined with miR165/166, MIR165/166 is targeted to the HD Zip III gene family, the expression of stem cells is regulated, but the actions of AtAGO10 and AtAGO1 are opposite, the expression stability of the HD Zip III gene is maintained, and the flower stem cells are normally terminated. AGO protein can regulate stress resistance, nutrition reproduction and plant morphology of plants, is important to plant growth, and has no report on regulation of soybean branch number. Disclosure of Invention The invention aims to provide an application of an AGO10 gene in regulating and controlling the number of soybean branches, in particular to an application of reducing the expression of the AGO10 gene in increasing the number of soybean branches. In order to achieve the above object, the technical scheme of the present invention is as follows: Based on the soybean genome-wide sequence disclosed in phytozome, the soybean AGO10 gene (also called AGO10a/b gene) comprises an AGO10a gene and an AGO10b gene, wherein the AGO10a gene has the gene number Glyma.17G119400, AGO b and the Glyma.05G01300 gene has the gene number. The length of the coding region nucleotide of the AGO10a gene is 2712bp, the CDS sequence is shown as SEQ ID NO.1, the coded protein consists of 903 amino acids, and the sequence is shown as SEQ ID NO. 3. The coding region of the AGO10b gene has the nucleotide length of 2727bp, the CDS sequence is shown as SEQ ID NO.2, the coded protein consists of 908 amino acids, and the sequence is shown as SEQ ID NO. 4. The main aim of the invention is to provide the application of the AGO10 gene in regulating and controlling the branch number of soybeans. The method is characterized in that the soybean branch number is increased after the expression of the AGO10 gene is reduced. In addition, the number of knots, the number of pods per plant, the hundred plant grain weight and the single plant grain weight of the AGO10 gene knockout mutant strain are also significantly increased compared with the wild type. Preferably, the means of reducing the expression of the AGO10 gene is a knockout of the AGO10 gene. According to the invention, by constructing the CRISPR-Cas9 vector of the AGO10 gene, the AGO10 gene knockout mutant strain with the number of soybean branches being significantly larger than that of wild type is obtained. Specifically, a CRISPR-Cas9 recombinant vector comprising an AGO10 gene-specific target sequence is transferred into soybean by using conventional biological methods such as Ti plasmid, ri plasmid, plant viral vector, direct DNA transformation, microinjection, conductance, agrobacterium mediation, etc., and the transformed soybean is cultivated into a plant. The invention also comprises a CRISPR-Cas9 recombinant vector containing the gene and a transformed plant line, and the function of a host cell containing the vector in regulating and controlling the soybean branch number also falls into the protection scope of the invention. The gene function protected by the invention not only comprises the AGO10 gene, but also comprises the function of the homologous gene with higher homology (such as homology higher than 80%, more preferably higher than 90%, more preferably higher than 95%, more preferably higher than 98%) with the AGO10 gene in regulating and controlling the branch number of soybean. The invention also discloses a plant breeding method, which is (1) or (2) as follows: