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Application of Arabidopsis gene AtSDH in regulating plant stress resistance

A technology of transgenic plants, Arabidopsis thaliana, applied in the application field of cultivating stress-resistant transgenic plant varieties, can solve unclear problems

Inactive Publication Date: 2011-01-26
SHANDONG AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, it is still unclear whether ABA can improve the ability of plants to resist adversity stress by regulating sorbitol metabolism.

Method used

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  • Application of Arabidopsis gene AtSDH in regulating plant stress resistance
  • Application of Arabidopsis gene AtSDH in regulating plant stress resistance
  • Application of Arabidopsis gene AtSDH in regulating plant stress resistance

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0025] Screening, Phenotypic and Genetic Analysis of Deletion Mutant Homozygotes

[0026] The T-DNA deletion mutants SALK 035903 (sdh-1) and SALK 020855 (sdh-2) inserted into the promoter region and coding region of the AtSDH gene purchased from the Arabidopsis Biological Resource Center were PCR-based. To screen homozygous mutants, the primers are:

[0027] SALK_035903

[0028] LP: 5′-CGAATTGGGTTTTAAAAATTCG-3′

[0029] RP: 5′-TGGATGGCTTATCGAGTTTTG-3′

[0030] SALK_020855

[0031] LP: 5′-TGAAAGCTGTTGGTATTTGTGG-3′

[0032] RP: 5′-CTACCTCCCTGTCTCCAAACC-3′

[0033] T-DNA (BP) primer: 5′-TCAAACAGGATTTTCGCCTGCT-3′

[0034] RT-PCR analysis of T-DNA insertion homozygous mutants sdh-1 and sdh-2 showed that the gene was not expressed at the transcriptional level in the mutants. The growth and development of deletion homozygous mutant plants under normal and stress conditions were analyzed.

Embodiment 2

[0036] Cloning of AtSDH gene, construction of vector, screening of transgenic plants

[0037] The AtSDH promoter sequence was amplified and isolated from the Arabidopsis genome by PCR method, and connected to the GUS reporter vector of pCAMBIA1391; RNA was extracted from Arabidopsis thaliana, reverse-transcribed into cDNA, and the CDS sequence of AtSDH was amplified from the cDNA , respectively connected to the overexpression vector of pBI121 and the subcellular localization vector of pBI121-GFP; Agrobacterium-mediated transformation to obtain transgene-positive plants.

[0038] Search the entire AtSDH genome sequence from the Arabidopsis information resource tair (The Arabidopsis Information Resource), and then take the 1551bp including TATAbox and other sequences before the start codon ATG, and include the first exon and the first exon after ATG An intron sequence, a total of 1779bp, then design primers, select the SalI / BamHI restriction site according to the multiple clonin...

Embodiment 3

[0043] Expression analysis of AtSDH gene

[0044] GUS histochemical staining of the transgenic plants carrying AtSDHP-GUS showed that AtSDH was mainly expressed in the roots and young leaves of the plants. Localization analysis of GFP fusion protein in transgenic Arabidopsis carrying AtSDH-GFP by laser confocal microscope showed that the protein encoded by the gene was localized in the cytoplasm ( figure 1 ). The autofluorescence of chloroplasts, mitochondria, peroxisomes, Golgi apparatus, and endoplasmic reticulum fluorescently labeled probes were used to detect and exclude the localization of the protein in the above organelles, and confirmed that the protein encoded by the gene was localized in the cytosol of the cytoplasm.

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Abstract

The invention relates to an application of Arabidopsis gene AtSDH, in particular to an application of the gene in regulating plant stress resistance and application of the gene in culturing anti-stress transgenic plant variety. In the invention, analysis of over-expression transgenic plants and deletion mutants to sensitivity and growth of ABA under stress condition is used to determine that the role of AtSDH in Arabidopsis stress is related to the plant hormone ABA, thereby providing important basis for understanding the role of plant sorbitol metabolism in plant stress resistance. When used in agricultural production, the gene can enable the seedlings and plants to more tolerate stress and lays theoretical foundation for culturing new variety of transgenic crops. Compared with wild Arabidopsis, the sensitivity of the transgenic Arabidopsis with over-expression of AtSDH gene to ABA is reduced and the capacity of the seedlings and plants to draught tolerance, high salt tolerance and the like is improved.

Description

technical field [0001] The invention relates to the application of an Arabidopsis thaliana gene AtSDH, in particular to the application of the gene in regulating the stress resistance of plants, and the application of the gene in the cultivation of transgenic plant varieties resistant to stress, and belongs to the field of genetic engineering. Background technique [0002] In the process of plant growth and development, abnormal growth conditions, such as drought, low or high temperature, high concentration of soil salt ions, etc., will cause ion imbalance and hyperosmotic stress in plants, resulting in poor growth, slow growth, especially reduced crop yields, and even cause plant death. Therefore, with the gradual deterioration of the environment and the reduction of arable land, people pay more and more attention to increasing the yield of food by improving the stress resistance of crops. [0003] Under adversity conditions, plants can reduce the damage caused by stress b...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/53A01H5/00
Inventor 王秀玲高新起
Owner SHANDONG AGRICULTURAL UNIVERSITY
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