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Application of DCM1 Protein and Its Coding Gene in Regulating Plant Male Fertility

A technology for male fertility and male sterility, which can be used in applications, plant peptides, plant products, etc.

Active Publication Date: 2019-01-15
INST OF GENETICS & DEVELOPMENTAL BIOLOGY CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there are very few natural male sterile plants in nature, and identification is difficult, requiring a lot of manpower and material resources

Method used

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  • Application of DCM1 Protein and Its Coding Gene in Regulating Plant Male Fertility
  • Application of DCM1 Protein and Its Coding Gene in Regulating Plant Male Fertility
  • Application of DCM1 Protein and Its Coding Gene in Regulating Plant Male Fertility

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0053] Embodiment 1, the acquisition of DCM1 gene

[0054] 1. Mutant acquisition and phenotype analysis

[0055] Carry out Guanglu Ai 4 60 Co ~ γ radiation mutagenesis to obtain a series of mutants.

[0056] The dcm1 mutant had no significant difference with Guangluai 4 in the vegetative growth stage, but it showed no fruiting ( figure 1 A). The ears of most dcm1 mutants do not set seeds at all, and a small number of ears will have several seeds ( figure 1 B). Pollen I2-KI staining found that compared with the plump and dyeable pollen of Guangluai 4 ( figure 1 C), the pollen coloring of most dcm1 mutants is light or uncolored with shrunken pollen ( figure 1 D).

[0057] The pollen of Guanglu'ai 4 was given to the female gametes of the dcm1 mutant, and they could bear fruit normally, which indicated that the female gametes of the dcm1 mutant were fertile.

[0058] 2. Image clone

[0059] A family with fertility segregation was selected, and the ratio of fertile plants ...

Embodiment 2

[0065] Embodiment 2, the expression pattern analysis of DCM1 gene

[0066] 1. Real-time quantitative PCR

[0067] By real-time quantitative PCR, the expression of DCM1 gene in the root (R), internode (IN), leaf (L) and young panicle (YP) of Guangluai 4 was detected, and the expression of DCM1 gene in the dcm1 mutant was detected. The expression level in the young panicle (dcm1). Ubiquitin gene was used as an internal reference.

[0068] see results figure 2 a. The results showed that DCM1 gene was expressed in these tissues.

[0069] 2. GUS staining

[0070] A recombinant plasmid carrying the promoter of the DCM1 gene was constructed and named pDCM1::GUS vector. The pDCM1::GUS vector was introduced into Yandao No. 8 to obtain transgenic plants. GUS staining was performed on the florets.

[0071] see results figure 2 B (ruler, 1 mm). The results showed that the promoter of DCM1 gene promoted the specific expression of GUS gene in anthers.

[0072] 3. In situ hybrid...

Embodiment 3

[0075] Example 3, cytological phenotype analysis of dcm1 mutants

[0076] 1. Semi-thin section observation

[0077] In order to determine the cause of the mutant's abortion, semi-thin section observations were made on the anthers of Guangluai 4 and the dcm1 mutant at different developmental stages.

[0078] see results image 3 . image 3 Middle: A-E corresponds to Guangluai 4, F-J corresponds to the dcm1 mutant; A and F correspond to the pachytene stage, B and G correspond to the tetrad stage, C and H correspond to the microspore stage, D and I correspond to the pollen mitosis stage, E and J correspond to mature pollen stage; scale bar, 20 μm; AP represents aborted pollen, ML represents middle layer, MP represents mature pollen, Msp represents microspore, PMC represents pollen mother cell, T represents tapetum, Tds represents tetrad, MMC stands for microspore mother cell.

[0079] During early meiosis, the anther compartment of the anther is surrounded by four layers of s...

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Abstract

The invention discloses the application of DCM1 protein and coding gene thereof in regulating male fertility of plants. The invention provides a method for cultivating a male sterile plant, comprisingthe following steps: inhibiting the expression of DCM1 gene in a target plant to obtain a male sterile plant; DCM1 gene encode DCM1 protein. The DCM1 protein is a protein composed of an amino acid sequence shown in SEQ ID NO: 1 in the sequence table. The inventors of the invention, through CRISPR-Knockout of Cas9 gene in wild-type rice can cause pollen abortion and produce male sterile lines. DCM1 protein regulates callose metabolism in rice pollen mother cells, thereby affecting the formation of pollen exine and pollen fertility. The invention provides a way for artificially controlling ricepollen fertility. It has important application value in the field of rice hybrid breeding.

Description

technical field [0001] The invention belongs to the field of biotechnology, and specifically relates to the application of DCM1 protein and its coding gene in regulating the male fertility of plants. Background technique [0002] Rice is one of the most important food crops worldwide. Nearly half of the world's population, including the vast majority of East and Southeast Asia, eat rice. With the increase of the world's population, the reduction of arable land and the deterioration of climatic conditions and other factors, the demand for increased rice production has become increasingly prominent. Dwarf breeding set off a "green revolution" in modern grain production in the 1960s. With the popularization of semi-dwarf varieties, the yield of rice has achieved breakthrough growth. Subsequently, the promotion of hybrid rice with heterosis has greatly increased the yield of rice and created huge social and economic benefits. [0003] In my country, the three-line hybrid ric...

Claims

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Application Information

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IPC IPC(8): C12N15/82C12N15/29A01H5/00A01H6/46C12N15/113C07K14/415
CPCC07K14/415C12N15/113C12N15/8213C12N15/8289C12N2310/10C12N2310/20
Inventor 程祝宽张超李亚非唐丁沈懿杜桂杰
Owner INST OF GENETICS & DEVELOPMENTAL BIOLOGY CHINESE ACAD OF SCI
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