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Gene editing and application of rice quality trait gene

A gene editing, rice technology, applied in the field of crops to achieve the effect of efficient gene editing

Pending Publication Date: 2022-01-18
北大荒垦丰种业股份有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, how to use gene editing methods to simultaneously edit multiple genes and multiple loci in different rice varieties to obtain homozygous edited japonica rice lines with aroma, non-transgenic, and stable genetics has rarely been reported.

Method used

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  • Gene editing and application of rice quality trait gene
  • Gene editing and application of rice quality trait gene
  • Gene editing and application of rice quality trait gene

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0086] The outstanding advantage of gene editing breeding is that the target gene can be pre-selected for editing. The technical principle is to use specific endonucleases to cut the target gene at the designed site, and through the cell's own DNA damage repair system, the broken DNA will be non-homologous end joining (NHEJ) or homology-dependent recombination (HDR) There are two main ways to fix it.

[0087] This embodiment includes: 1. Construction of genetic transformation and editing vectors, constructing molecular tools such as specific endonucleases into the transformation vectors; 2. Genetic transformation of recipient plants, placing the above-mentioned molecular tools in cells to play a role; 3. 1. Molecular biological analysis of transformed and regenerated plants, detecting the status of the target gene editing site, and confirming whether the gene editing is successful; 4. Molecular biological analysis of the offspring of the gene editing plants, confirming the edi...

Embodiment 2

[0123] 1. Construction of gene editing transformation vector:

[0124] Using the same KF50 as described in the construction of the gene editing transformation vector in Example 1 as the gene editing transformation vector, the Kenfeng Seed Industry's self-owned rice variety Longken 290 was improved by gene editing to increase the aroma and waxy quality traits.

[0125] 2. Genetic transformation of japonica rice:

[0126] Using the genetic transformation method for indica rice described in Example 1, the gene editing transformation vector KF50 was used to transform the self-owned rice variety Longken 290 of Kenfeng Seed Industry. The transformed japonica rice variety is easy to produce callus, has strong differentiation and regeneration ability, and high transformation efficiency , suitable for Agrobacterium transformation. Resistant calli were screened for about 3-4 weeks after infection with the gene editing vector, and were directly transferred to the differentiation medium ...

Embodiment 3

[0140] 1. Construction of gene editing transformation vector:

[0141] Using the same KF50 as described in the construction of the gene editing transformation vector in Example 1 as the gene editing transformation vector, Kenfeng Seed Industry's self-owned rice variety Kendao 32 was improved by gene editing to increase the aroma and waxy quality traits.

[0142] 2. Genetic transformation of japonica rice:

[0143] Using the indica rice genetic transformation method described in Example 1, the Kenfeng Seed Industry’s own rice variety Kendao 32 was transformed with the gene editing transformation vector KF50. The transformed japonica rice variety is easy to produce callus, has strong differentiation and regeneration ability, and high transformation efficiency , suitable for Agrobacterium transformation. Resistant calli were screened for about 3-4 weeks after infection with the gene editing vector, and were directly transferred to the differentiation medium for 3-4 weeks to diff...

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Abstract

The invention belongs to the field of crops, and particularly relates to an editing method and application of a japonica rice variety quality improvement breeding fragrance control and starch synthesis gene. The invention discloses a gene editing transformation vector, which takes pCambia3301 as a vector skeleton, and further comprises a ZmU6 promoter and a ZmUBI1 promoter, the vector also comprises a plurality of gRNA fragments designed for recognizable sites of different exon regions of Badh2 and Waxy genes; the gRNA fragments are located between the ZmU6 promoter and the ZmUBI1 promoter, and synchronous expression is performed through the ZmU6 promoter; and the gRNA fragments are interactively connected through a tRNA technology. According to the invention, a gene editing genetic transformation vector is constructed, an agrobacterium tumefaciens-mediated method is used for transforming a japonica rice variety, various editing of a nucleotide sequence of regenerated plant Badh2 and Waxy gene target regions are realized, and finally, a glutinous rice, non-transgenic and stably inherited homozygous edited rice strain with fragrance is obtained.

Description

technical field [0001] The invention belongs to the field of crops, and in particular relates to a gene editing method of an aroma control gene and a starch synthase gene of an improved strain of japonica rice and an application thereof. Background technique [0002] The rapid development of gene editing technology in recent years uses specific endonucleases to selectively cut specific genes in cells, through two DNA repair mechanisms inherent in cells, non-homologous end joining (NHEJ) and homologous end joining (NHEJ) Rely on Homology dependent repair (HDR), repair the fracture, and ensure the survival of cells. NHEJ has high repair efficiency, does not require a template, and occupies a dominant position, but the process of cutting point linkage will introduce insertion or deletion mutations, thereby inactivating the gene; HDR needs to follow the template, although the repair is accurate, but the proportion is very small. If provided The pre-designed DNA fragment is used...

Claims

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Application Information

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IPC IPC(8): C12N15/53C12N15/54C12N15/84C12N15/113A01H5/10A01H6/46
CPCC12N9/0008C12N9/1051C12N15/8242C12N15/8245C12N15/113C12Y102/01008C12Y204/01021C12N2310/20
Inventor 李忠森王明月马瑞朱婷刘丹徐希德李强马红友
Owner 北大荒垦丰种业股份有限公司
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