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Application of cucumber light-harvesting chlorophyll a/b binding protein CsPS1 to resisting phytophthora melonis

A technology for combining proteins and chlorophyll, which is applied in the field of plant molecular biology and plant genetic engineering, and can solve the problems of unsatisfactory chemical control effect and economic loss, etc.

Active Publication Date: 2021-01-05
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

Once the disease occurs, the control effect of chemical agents is not ideal, causing economic losses

Method used

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  • Application of cucumber light-harvesting chlorophyll a/b binding protein CsPS1 to resisting phytophthora melonis
  • Application of cucumber light-harvesting chlorophyll a/b binding protein CsPS1 to resisting phytophthora melonis
  • Application of cucumber light-harvesting chlorophyll a/b binding protein CsPS1 to resisting phytophthora melonis

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] Example 1 Construction of Cucumber CsPS1 Gene Overexpression Vector

[0042] Cucumber CsPS1 gene belongs to light-harvesting chlorophyll a / b binding protein gene. Previous studies have shown that light-harvesting chlorophyll a / b binding proteins play an important role in plant development, plant stress, especially abiotic stress. The full length of the cucumber CsPS1 gene CDS was constructed into pGWB5 (the plasmid map is attached figure 1 As shown), the overexpression vector was obtained; the cucumber gene CDS non-conserved segment 207bp length fragment and its complementary fragment were constructed on pK7GWIW (plasmid map as attached figure 2 shown), and obtained on the gene silencing vector.

[0043] The amino acid sequence of cucumber CsPS1 protein is (267aa):

[0044] MAASSMALSSPSFAGQAVKLSPTASDLLGEGRITMRKTAGKPKPVSSGSPWYGPDRVKYLGPFSGEPPSYLTGEFPGDYGWDTAGLSADPETFAKNRELEVIHSRWAMLGALGCVFPELLSRNGVKFGEAVWFKAGSQIFSEGGLDYLGNPSLVHAQSILAIWACQVVLMGAVEGYRIAGGPLGEITDPIYPGGS...

Embodiment 2

[0051] Example 2 Constructing a Cucumber Cotyledon Model of Transient Overexpression / Gene Silencing

[0052] (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 48-72 hours.

[0053] (2) Pick a single clone and add it to 4 mL of LB medium containing the corresponding antibiotics and rifampicin, shake the bacteria at 180 rpm for 24-36 hours at 28°C.

[0054] (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.

[0055] (4) Collect the bacteria by centrifugation at 3000rpm for 5 minutes, resuspend the bacteria with a suspension (10mM MES, 10mM MgCl2), adjust the OD600 value to about 0.4, and add 200mM acetosyringone.

[0056] (5) Stand at room temperature for 3-5 hours.

[0057] (6) Prick a needle hole on both sides of the...

Embodiment 3

[0070] Example 3 Cucumber cotyledon disease resistance experiment

[0071] For the cucumber cotyledon model with transient overexpression of CsPS1 successfully constructed in Example 2, the cucumber cotyledon model of CsPS1 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 to 4 days. Cucumber cotyledons were isolated and inoculated with Phytophthora cucurbiti, and cultured in a culture dish at 28°C for 24 hours in the dark.

[0072] 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.

[0073] The results showed that transient overexpression of CsPS1 gene in cucumber cotyledon can significantly enhance the resistance of cucumber cotyledon to blight compared with the control, while tran...

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Abstract

The invention discloses an application of a cucumber light-harvesting chlorophyll a / b binding protein CsPS1 to resisting phytophthora melonis for the first time. And particularly, the CsPS1 has a disease-resistant function on phytophthora melonis infecting melons. Overexpression of the CsPS1 gene can obviously enhance the resistance of cucumber cotyledons and pumpkin cotyledons to epidemic diseases compared with a control group, and transient silencing of the CsPS1 gene in cucumber cotyledons can obviously weaken the resistance of the cucumber cotyledons to epidemic diseases compared with thecontrol group. The CsPS1 gene plays a very important role in resisting diseases of cucumbers, pumpkins and other melons, and has a wide application prospect.

Description

technical field [0001] The invention relates to the fields of plant molecular biology and plant genetic engineering, and more specifically relates to the application of cucumber light-harvesting chlorophyll a / b binding protein CsPS1 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 has occurred in areas where melons are grown in my country. [0003] In the north, it occurs more heavily in summer and autumn open-field cultivation and spring protected sweet melons; in the south, it occurs more frequently in spring, summer and autumn, 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-soaked and const...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/82C07K14/415A01H5/00A01H6/34A01N63/40A01N63/50A01N63/60A01N37/46A01N57/16
CPCA01N37/46A01N57/16A01N63/40A01N63/50A01N63/60C07K14/415C12N15/8282
Inventor 吴廷全杜虎王瑞徐晓美金庆敏杨晓珊
Owner INST OF VEGETABLES GUANGDONG PROV ACAD OF AGRI SCI
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