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Application of RAF49 protein and coding gene thereof in regulating and controlling drought resistance of plants

A technology of RAF49, 1.RAF49, applied in application, plant products, genetic engineering, etc., can solve problems such as accumulation of organic matter and decrease of seed setting rate, inhibition of crop nutrition and reproductive growth, impact on crop yield and quality, and improve breeding efficiency , shorten the breeding cycle, and reduce the degree of leaf wilting

Active Publication Date: 2020-03-24
CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Water stress will inhibit crop nutrition and reproductive growth, resulting in shorter plant height, wilting leaves, reduced photosynthesis, decreased organic matter accumulation and seed setting rate, and insufficient grain or fruit, which seriously affects crop yield and quality

Method used

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  • Application of RAF49 protein and coding gene thereof in regulating and controlling drought resistance of plants
  • Application of RAF49 protein and coding gene thereof in regulating and controlling drought resistance of plants
  • Application of RAF49 protein and coding gene thereof in regulating and controlling drought resistance of plants

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0045] Example 1 Construction and detection of RAF49 gene overexpression vector

[0046] Extract total RNA from corn (Zea mays L.), reverse transcribe to obtain cDNA, use cDNA as a template, and F and R as primers to amplify the RAF49 gene. on the carrier. The specific construction method is as follows:

[0047] (1) The total RNA of corn was extracted with the RNA extraction kit of Magen Company, and the specific steps were referred to the instructions of the kit.

[0048] (2) Use the reverse transcription kit from Thermo Company to reverse transcribe the RNA into cDNA, and refer to the kit instructions for specific steps.

[0049] (3) Using cDNA as a template and F and R as primers, amplify the cDNA of RAF49 gene, run electrophoresis and cut the gel to recover the amplified product, and the recovery method refers to the kit of Tiangen Company.

[0050] The primer sequences used for RAF49 gene amplification are as follows:

[0051] SEQ ID NO.3: Upstream primer F: GCTCTAGAG...

Embodiment 2

[0056] SEQ ID NO. 6: NosR-seq: AGACCGGCAACAGGATTCAATC. Construction and detection of embodiment 2 RAF49 gene overexpression plant

[0057] The pBCXUN-RAF49 overexpression plasmid constructed in Example 1 was transformed into competent Agrobacterium strain EHA105 by heat shock method, and positive clones were identified by colony PCR. Inoculate a single colony of Agrobacterium correctly identified in 2-3 mL of liquid medium containing 100 μg / mL kanamycin and 50 μg / mL rifampicin, culture with shaking at 28 °C overnight, and transfer a large amount of liquid containing antibiotics the next day Shake culture in the medium, collect the cells after several transfers, and resuspend to OD 600 Between 0.8-1.0. The obtained recombinant Agrobacterium suspension was used to infect the immature corn embryos of B73 under aseptic conditions, and the callus was induced to form seedlings. The transgenic plants were self-propagated to obtain the T3 generation for subsequent experiments.

[...

Embodiment 3

[0059] Example 3 Phenotype Detection of RAF49 Gene Overexpression Maize Drought Treatment

[0060] The RAF49 gene overexpression maize line T3 generation that embodiment 2 constructs carries out drought treatment experiment, and specific method is as follows:

[0061] Add 140g of soil to each small pot, add water to the tray, put 4 seeds in each small pot, cover with 50ml of soil, pour out the remaining water in the tray after absorbing the water, and cultivate it in a greenhouse at 25°C. After emergence, it will grow Remove a seedling that is uneven, add 1L of water to the tray, pour out the water after it is full, and start drought treatment (continue to cultivate in a greenhouse at 25°C, no water will be given during the cultivation process), dry treatment for about a week, and observe wild plants. phenotypes such as growth and leaf wilting degree of transgenic plants under drought treatment. Three pots of wild-type and transgenic plants were used as biological replicates....

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Abstract

The invention relates to the technical field of plant genetic engineering, in particular to an application of RAF49 protein and an encoding gene of the RAF49 protein to regulation and control of drought resistance of plants. According to the invention, the corn RAF49 gene is found to positively regulate the drought resistance of the plant, and the drought resistance of the plant can be effectivelyimproved by increasing the expression quantity of the RAF49 gene. The RAF49 overexpression transgenic corn plant is constructed, the growth condition of the plant under the drought condition is obviously superior to that of a wild type corn plant, the leaf wilting degree is obviously reduced, the transpiration rate and the stomatal conductance are obviously lower than those of the wild type cornplant, the water loss of the plant is effectively reduced, and the drought resistance is obviously improved. The discovery of the drought-resistant function of RAF49 provides a new gene target and resources for cultivating new varieties of drought-resistant plants, and provides a theoretical basis for illustrating the molecular mechanism of plant drought stress signal response.

Description

technical field [0001] The invention relates to the technical field of plant genetic engineering, in particular to the application of RAF49 protein and its coding gene in regulating plant drought resistance. Background technique [0002] Drought is an important abiotic stress factor that affects crop yield and quality, and is one of the major problems facing global agricultural production. Water stress will inhibit crop nutrition and reproductive growth, resulting in shorter plant height, wilting leaves, decreased photosynthesis, decreased organic matter accumulation and seed setting rate, and insufficient grain or fruit, which seriously affects crop yield and quality. Maize is one of the three major food crops in the world. It is planted in a wide range of areas. It originated in hot and humid tropical regions and is not resistant to drought. Breeding new drought-resistant varieties can effectively cope with yield loss caused by drought stress and improve water use efficie...

Claims

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Application Information

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IPC IPC(8): C12N9/12C12N15/54C12N15/82A01H5/00A01H6/46
CPCC12N9/1205C12N15/8273C12Y207/01037
Inventor 巩志忠王瑜韩祎楠
Owner CHINA AGRI UNIV
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