Wheat fertility-restoring gene and application thereof

A fertility restoration gene and gene technology, applied in the field of plant biology, can solve the problems of narrow restoration sources, limited hybrid variety resources, and unstable fertility of sterile lines.

Active Publication Date: 2017-10-31
BEIJING NEXT GENERATION HYBRID WHEAT BIOTECHNOLOGY CO LTD +2
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The three-line method has not been widely used in production due to the difficulty in breeding sterile lines, narrow recovery sources, and cytoplasmic side effects.
At the same time, in order to solve the technical bottlenecks of the existing "three-line" and two-line" hybridization technologies of wheat, such as unstable sterile line fertility, limited hybrid variety resources, complicated seed production technology, and high seed production costs, it provides more many possibilities

Method used

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  • Wheat fertility-restoring gene and application thereof
  • Wheat fertility-restoring gene and application thereof
  • Wheat fertility-restoring gene and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1、4

[0086] Embodiment 1, Breeding of 4AgS-ms double-terminal variant addition lines

[0087] The Lanzhou nuclear male sterility mutant was found in the hybrid F4 population of the hybrid combination of spring wheat varieties. It was crossed with 9 wheat lines including China Spring, the F1 generation was self-crossed, and the segregation ratio of the sterile and fertile plants of the F2 generation was observed, which was in line with the ratio of 1:3, indicating that wheat Lanzhou nuclear male sterility mutation The body is a typical recessive mutation controlled by a single gene (Zhou Kuanji et al., 1996).

[0088] The blue-grain disomic addition line is a wheat strain with two additional 4Ag chromosomes of E. elongatus in the wheat genome. The reason for the blue grain color is that the additional 4Ag chromosomes contain the blue-grain gene Ba. The pollen of the blue-grained disomic addition line of wheat was given to the Lanzhou nuclear male sterile mutant, and the light blue-...

Embodiment 2

[0091] Embodiment 2, utilizing flow cytometry to separate 4Ag chromosome

[0092] The root tip cells were treated with cell cycle synchronization by double blocking method, and after formalin fixation, mechanical homogenization and DAPI staining, they were loaded into a flow cytometer.

[0093] First, flow cytometry karyotype analysis was performed to confirm the size of the chromosomes to be separated. The 4AgS-ms paired-end somatic addition line was used as the experimental group, and China Spring was used as the control. According to the size difference of wheat chromosomes, the karyotype map usually forms four independent peaks (Vrana et al., 2000): Peak I contains four chromosomes 1D, 4D, 6D and 7D; peak II contains 1A, 3A, 6A, There are 6 chromosomes in 2D, 3D and 5D; peak III includes 10 chromosomes in total 2A, 4A, 5A, 7A, 1B, 2B, 4B, 5B, 6B and 7B; 3B is the largest and forms a peak independently. Comparing the karyotype map of China Spring, it can be seen that in ad...

Embodiment 3、4

[0095] Example 3, High-throughput sequencing and sequence assembly of 4Ag chromosome

[0096] Chromosomes isolated by flow cytometry are highly condensed chromatin, which cannot be directly applied to high-throughput sequencing. DNA must be separated from chromosomes by proteinase K digestion. 2.2 million 4ES chromosomes were co-purified to obtain about 500ng DNA with good integrity.

[0097] Using 100ng of 4ES genome as a template, Qiagen REPL1-g Single Cell Kit was used for gene amplification, and a total of 30μg of amplification products were obtained. The amplified products were digested, cloned, and sequenced. Among the 31 clones, no Escherichia coli and human contamination were found. One clone could completely match the genome sequence of wheat. The remaining clones were probably derived from E. elongatum 4Ag. 97%.

[0098] Considering that the genomes of Triticum species have a high proportion of repetitive sequences and difficult splicing (the proportion of repetiti...

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Abstract

The invention discloses a wheat fertility-restoring gene and an application thereof, belongs to the technical field of plant biology and particularly relates to a recessive nuclear male sterility gene and cloning of a promoter thereof, and an application of the recessive nuclear male sterility gene in cross breeding. A fertility-restoring gene FRG1 is successfully cloned through a flow cytometry and a high-throughput sequencing method. The gene is capable of completely restoring the male fertility of a Lanzhou nuclear male sterility mutant or an allelic mutant thereof and a basis is laid for construction of a novel wheat hybrid breeding technology system. Meanwhile, more possibilities are provided for solving the technical bottleneck problems that an existing wheat two-line and three-line hybrid technology is unstable in sterile line fertility, limited in hybrid variety resource, complicated in seed production technology and high in seed production cost. The provided gene and a sterile line propagation and retention method have important significance and application value in wheat cross breeding work.

Description

technical field [0001] The invention belongs to the field of plant biotechnology, and in particular relates to the cloning of a plant recessive nuclear male sterility gene, its male sterile line propagation method and its application in hybrid breeding, and more specifically relates to a recessive nuclear male sterility gene The cloning of its promoter and its application in cross breeding. technical background [0002] Heterosis is a common phenomenon in the biological world. Hybrid breeding is the main way to select and breed new varieties, and is the most important method in modern breeding. Compared with rice, corn, sorghum, etc., the research on the use of heterosis in wheat is relatively lagging behind, and the growth of its average yield has also tended to stagnate in the past ten years, or even declined for many years. Wheat is a self-pollinated crop. The core issue of wheat heterosis utilization is the technical system for efficient production of wheat hybrids. A l...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/29C07K14/415C12N15/82A01H5/00
CPCC07K14/415C12N15/8287C12N15/8289C12N15/52C12N15/8231C12N15/8209C12N2830/007C12N2830/008C12N2830/65
Inventor 马力耕王峥李健何航陈少霞邓兴旺
Owner BEIJING NEXT GENERATION HYBRID WHEAT BIOTECHNOLOGY CO LTD
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