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Application of cucumber photosynthetic system II oxygen evolution enhanced protein CsPSII-OEEP to resisting melon epidemic diseases

A technology of photosynthetic system and cucumber, which is applied in the field of plant molecular biology and plant genetic engineering, can solve the problem that the oxygen evolution enhancing protein of cucumber photosynthetic system II has not been found, and achieve the effect of wide application prospects

Active Publication Date: 2021-12-14
INST OF VEGETABLES GUANGDONG PROV ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Previous studies have shown that plant photosynthetic system II oxygen evolution-enhancing protein has thioredoxin activity and plays an important role in abiotic stress. However, it has not been found that cucumber photosynthetic system II oxygen evolution-enhancing protein CsPSII-OEEP plays an important role in resisting melon blight report

Method used

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  • Application of cucumber photosynthetic system II oxygen evolution enhanced protein CsPSII-OEEP to resisting melon epidemic diseases
  • Application of cucumber photosynthetic system II oxygen evolution enhanced protein CsPSII-OEEP to resisting melon epidemic diseases
  • Application of cucumber photosynthetic system II oxygen evolution enhanced protein CsPSII-OEEP to resisting melon epidemic diseases

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0092] Example 1 Construction of cucumber CsPSII-OEEP gene overexpression vector

[0093] Cucumber CsPSII-OEEP gene belongs to the member protein gene of cucumber photosynthetic system II. The cucumber CsPSII-OEEP gene CDS full-length (SEQ ID NO.2) was constructed into pGWB5 (pGWB5 plasmid map as figure 1 Shown), the specific position is between attR1 and attR2 behind the 35S promoter (the ccdB gene was replaced by gene recombination exchange), and an overexpression vector (pGWB5-CsPSII-OEEP) was obtained; the non-conserved region of the cucumber CsPSII-OEEP gene CDS Fragments (CDS 1-239bp, SEQ ID NO.4) and their complementary fragments of 239bp length were constructed on pK7GWIW (the pK7GWIW plasmid map is shown in figure 2 shown), the specific position is between the two attR1 and attR2 behind the 35S promoter (the ccdB gene was replaced by gene recombination exchange), and the gene silencing vector (pK7GWIW-CsPSII-OEEP) was obtained.

[0094] The amino acid sequence of c...

Embodiment 2

[0102] Example 2 Constructing a Cucumber Cotyledon Model of Transient Overexpression / Gene Silencing

[0103] (1) The overexpression vector and the silencing vector in Example 1 were respectively transformed into Agrobacterium GV3101, and cultured upside down on the corresponding resistant medium for 48 hours.

[0104] (2) Pick a single clone and add it to 4 mL of LB medium containing corresponding antibiotics and rifampicin, shake the bacteria at 180 rpm for 24 hours at 28°C.

[0105] (3) Add fresh LB medium containing corresponding antibiotics and rifampicin at a ratio of 1:100, shake the bacteria at 28°C and 180 rpm until the OD600 value is about 3.0.

[0106] (4) Centrifuge at 3000rpm for 5 minutes to collect the bacteria, and use the suspension (10mM MES, 10mM MgCl 2 ) to resuspend the cells, adjust the OD600 value to about 0.4, and add 200 mM acetosyringone.

[0107] (5) Stand at room temperature for 3 hours.

[0108] (6) Prick a needle hole on both sides of the main v...

Embodiment 3

[0121] Example 3 Cucumber cotyledon disease resistance experiment

[0122] For the cucumber cotyledon model with transient overexpression of CsPSII-OEEP successfully constructed in Example 2, the cucumber cotyledon model of CsPSII-OEEP gene silencing, and the wild-type cucumber cotyledon, the cucumber seedlings after injection were cultured in the dark for 12 hours and then cultured in the light at 22°C for 3 Two days later, the cucumber cotyledon was isolated in vitro, and after being inoculated with Phytophthora meloni, the cucumber cotyledon was cultured in a culture dish at 28°C for 24 hours in the dark.

[0123] The resistance function of the target gene to the disease was judged according to the size of the lesions on the isolated cucumber cotyledons of different treatments. The results are shown in the attached Figure 5 shown.

[0124] The results showed that the transient overexpression of CsPSII-OEEP gene (pGWB5-CsPSII-OEEP) in cucumber cotyledon can significantly enh...

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Abstract

The invention belongs to the field of plant molecular biology and plant genetic engineering, and discloses application of cucumber photosynthetic system II oxygen evolution enhanced protein CsPSII-OEEP to resisting melon epidemic diseases. It is the first time for the invention to find that the cucumber photosynthetic system II oxygen evolution enhanced protein CsPSII-OEEP has resistance to melon epidemic diseases: plant cotyledons that overexpress the cucumber photosynthetic system II oxygen evolution enhanced protein CsPSII-OEEP are inoculated with phytophthora melonis, so that the lesion area is obviously reduced, plant cotyledons that silence the cucumber photosynthetic system II oxygen evolution enhanced protein CsPSII-OEEP are inoculated with the phytophthora melonis, so that the lesion area is obviously increased; it can be seen that the cucumber photosynthetic system II oxygen evolution enhanced protein CsPSII-OEEP plays a very important role in resisting the melon epidemic disease of plants, can be used for resisting the melon epidemic diseases, preparing products for resisting the melon epidemic diseases and cultivating transgenic plants for resisting the melon epidemic diseases, and has a wide application prospect.

Description

technical field [0001] The invention belongs to the fields of plant molecular biology and plant genetic engineering, and specifically relates to the application of cucumber photosynthetic system II oxygen evolution enhancing protein CsPSII-OEEP in resisting melon blight. Background technique [0002] Melon blight is a worldwide oomycete disease. The pathogen is Phytophthora melonis, which causes the leaves, stems, and fruits of cucumbers, pumpkins, wax gourds, and bitter melons to turn brown and cause death or dead seedlings. The disease occurs in areas where melons are grown in my country. In the north, the occurrence of summer and autumn open field cultivation and spring protected sweet melons is heavier, and in the south, the occurrence is heavier in spring and summer and autumn with high temperature and humidity. The entire growth period and various parts of melons can be damaged. The damaged growth point of the seedlings and the base of the tender stems are water-soake...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/415C12N15/29C12N15/84A01H5/00A01H6/34
CPCC07K14/415C12N15/8282C12N15/8218C12N15/8205
Inventor 金庆敏吴廷全杜虎王瑞钟玉娟杨晓珊
Owner INST OF VEGETABLES GUANGDONG PROV ACAD OF AGRI SCI
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