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Application of Cucumber Phloem Lectin cspl1 in Resistance to Cucurbit Blight

A phloem and lectin technology, applied in the fields of plant molecular biology and plant genetic engineering, can solve the problems of undisclosed anti-epidemic diseases, and achieve the effect of improving resistance and wide application prospects

Active Publication Date: 2021-06-15
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

but did not disclose its anti-epidemic effect

Method used

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  • Application of Cucumber Phloem Lectin cspl1 in Resistance to Cucurbit Blight
  • Application of Cucumber Phloem Lectin cspl1 in Resistance to Cucurbit Blight
  • Application of Cucumber Phloem Lectin cspl1 in Resistance to Cucurbit Blight

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] Example 1 Construction of cucumber CsPS1 gene overexpression vector and silencing vector

[0042] The protein encoded by the cucumber CsPL1 gene is Phloem lectin, which can specifically bind monosaccharides or polysaccharides. The full-length construction of the cucumber CsPL1 gene CDS into pGWB5 (the vector map is as follows figure 1 As shown), the overexpression vector was obtained; the cucumber gene CDS non-conserved segment about 200bp length fragment and its complementary fragment were constructed into pK7GWIW (vector map as shown in figure 2 shown), the gene silencing vector was obtained.

[0043] The amino acid sequence of cucumber phloem lectin CsPL1 is (154aa):

[0044] MAGQSTHYLAFPRASTITWGDDTRYWSWATVDFCSYAIEEARLLQVSWFDCRWSMDASDFKQDIWYNASVEVMLTSNASGWNVPLHLEIELPDGSKQESQIVLAGRQPNVWLKIPIGKFILRGSLTSGTIRFGLYNHEGNWKRGLNIRALAIQA (SEQ ID NO. 1).

[0045] Cucumber phloem lectin CsPL1 gene CDS (462bp) sequence is:

[0046]ATGGCAGGCCAAAGCACACATTATTTGGCATTTCCAAGAGCTTC...

Embodiment 2

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

[0052] (1) The overexpression vector and the silencing vector constructed 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

[0054] 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.

[0055] (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.

[0056] (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.

[0057] (5) Stand at room temperature for 3-5 hours.

[0058] (6) Prick a needle ho...

Embodiment 3

[0071] Example 3 Cucumber cotyledon disease resistance experiment

[0072] Taking the cucumber cotyledon model with transient overexpression of CsPL1 successfully constructed in Example 2, the cucumber cotyledon model of CsPL1 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.

[0073] 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.

[0074] The results showed that transient overexpression of CsPL1 gene in cucumber cotyledon can significantly enhance the resistance of cucumber cotyledon to blight compared with the control, while ...

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Abstract

The invention firstly discloses the application of cucumber phloem lectin CsPL1 in resisting melon blight. In particular, CsPS1 has a disease resistance function against Phytophthora melonis which infects melons. Overexpression of CsPS1 gene can significantly enhance the resistance of cucumber and pumpkin cotyledon to blight compared with control, while transient silencing of CsPS1 gene in cucumber cotyledon can significantly weaken the resistance of cucumber cotyledon to blight compared with control. The CsPL1 gene plays a very important role in the resistance of melons such as cucumbers and pumpkins to diseases, and has broad application prospects.

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 phloem lectin CsPL1 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 planted in my country, and it occurs more frequently in summer and autumn open field cultivation and spring protected melons in the north; in the south, it occurs more frequently in spring and summer and autumn high temperature and humidity seasons. 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 constricted, wilting and dying. In t...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/82A01H5/00A01H6/34A01N63/50A01N37/46A01N57/16A01P3/00
CPCA01N37/46A01N57/16A01N63/50C07K14/42C12N15/8282
Inventor 杜虎吴廷全王瑞徐晓美金庆敏杨晓珊
Owner INST OF VEGETABLES GUANGDONG PROV ACAD OF AGRI SCI