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LR2 gene regulating stem thickness and breaking-resistant strength of rice and application thereof in lodging resistance

A technology of lodging resistance and genetics, applied in the fields of application, genetic engineering, plant genetic improvement, etc., can solve the problems of reducing rice quality, increasing the difficulty of harvesting, increasing production costs, etc., and achieve the effect of excellent genetic resources

Active Publication Date: 2019-07-30
YANGTZE UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Lodging not only directly leads to a serious loss of rice yield, but also reduces the quality of rice and increases production costs due to increased difficulty in harvesting

Method used

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  • LR2 gene regulating stem thickness and breaking-resistant strength of rice and application thereof in lodging resistance
  • LR2 gene regulating stem thickness and breaking-resistant strength of rice and application thereof in lodging resistance
  • LR2 gene regulating stem thickness and breaking-resistant strength of rice and application thereof in lodging resistance

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] Embodiment 1: Comparison of lodging resistance of mutant M18 and "93-11" wild type

[0029]The material of the test for inducing mutants came from the indica rice variety "93-11". After the seeds were treated with 0.015% (volume ratio) ethyl methanesulfonate (ie EMS) for 12 hours, the M1 generation was obtained and selfed The M2 generation was obtained, and the mutant M18 with thickened culm wall was found. Selfing multiple generations showed that the mutant trait could be genetically planted stably. Rice mutants M18 and “93-11” were planted, and the bending resistance of the stem base was investigated. The measured values ​​are shown in Table 1.

[0030] Table 1 Measurement of phenotypic values ​​of rice mutant M18 and rice variety "93-11"

[0031]

[0032] **, extremely significant difference, P value ≤0.01.

Embodiment 2

[0033] Example 2: The positioning of LR2 and the determination of candidate genes

[0034] 1. Separation population construction and localization of LR2

[0035] The rice mutant M18, which was stably inherited through multiple generations of self-crossing, was used as the female parent to backcross the rice variety "93-11" to obtain F 1 Positive individual plants, select one of the F 1 F 2 Separate groups. 248 F. 2 The segregated populations included 189 plants with normal phenotype and 59 plants with mutant phenotype. After chi-square (χ 2 ) test, the separation ratio is 3:1(χ 2 (3:1) =0.132 0.05，1 =3.84), indicating that the mutant is controlled by a single recessive nuclear gene. Using the strategy of MutMap, the F 2 The mixed DNA samples of 30 wild-type and mutant strains in the population were sequenced separately, the SNP frequency calculation of the offspring and the screening of candidate sites were performed, and the candidate interval was locked as Chr2: 31,...

Embodiment 3

[0043] Example 3: Verification of transgenic function of LR2

[0044] Using the sequence of the rice material "Nipponbare" (the genome sequence of which has been published and is a conventional variety) as a reference, the LR2 gene was designed using the online tool CRISPR-P (http: / / cbi.hzau.edu.cn / CRISPR2 / ). Knockout targets and primers (Table 3). The sgRNAs expression cassette was constructed using the Overlapping PCR method, and then the sgRNAs expression cassette and the pYLCRISPR Cas9Pubi-H expression vector were assembled using the Golden Gatecloning strategy (see Figure 4 ). The specific vector construction process can refer to the method reported by Ma et al. (Ma X, Zhang Q, Zhu Q, et al. A robust CRISPR / Cas9 system for convenient, high-efficiency multiplex genome editing in monocot and dicot plants[J]. Molecular plant, 2015, 8(8):1274-1284). After the primers "SPL" and "SPR" were sequenced and found that the sgRNAs encoded by the constructed CRISPR vector were cor...

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Abstract

The invention belongs to the technical field of plant genetic engineering, and particularly relates to the application of LR2 gene in regulating stem thickness and breaking-resistant strength of riceand application thereof in lodging resistance. The LR2 gene and an allele of the LR2 gene are isolated, wherein the LR2 gene and the allele of the LR2 gene affect the lodging resistance of the rice. The sequence of the LR2 gene is shown as SEQ ID NO: 1, and the sequence of the allele of the LR2 is shown as SEQ ID NO: 3. In the '93-11' of rice wild type and '93-11' mutant M18, the corresponding genome of the LR2 is performing comparative sequencing, and it is found that one base difference exists between the two materials, wherein the base is located on the fourth exon of a coding region, and the base substitution from C to T occurs, thereby leading to the change of one amino acid. The rice plants with LR2 gene knockout are obtained by transgenosis. The transgenic plants show the phenotypeof thickening of the stem wall and increasing of the basal internode breaking-resistant strength, thereby improving the lodging resistance. These character changes are in good agreement with the '93-11' mutants M18.

Description

technical field [0001] The invention relates to the technical field of plant genetic engineering. It specifically relates to the regulation of rice stem thickness, bending resistance and its application in lodging resistance by LR2 gene. Background technique [0002] As the world population continues to grow, it is particularly important to maintain stable and increased food production. Lodging is one of the main limiting factors affecting the stable and high yield of rice. Lodging is mostly caused by external factors. It refers to the phenomenon that the stems of plants change from a natural upright state to a permanent dislocation. It is a common problem in crop production. With the breeding of ultra-high-yielding rice varieties, the promotion of direct seeding and close planting techniques, and the increase in chemical fertilizer application, the problem of lodging has become increasingly serious. The stem is the support of the plant, and its physical properties are on...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/29C07K14/415C12N15/82A01H5/00A01H6/46
CPCC07K14/415C12N15/8213
Inventor 李志新邢永忠周想春张一林刘华
Owner YANGTZE UNIVERSITY
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