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Plant expression vector for targeting silence phytophthora capsici cellulose synthase 3 and application thereof

A technology of plant expression vector and cellulose synthase, which is applied in the field of agricultural biology, can solve the problems of undiscovered plant diseases and other problems, and achieve the effects of delaying the emergence of drug resistance, benefiting the environment, and reducing the amount of use

Inactive Publication Date: 2021-07-23
CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

After searching, there is currently no relevant literature on the control of plant diseases caused by Phytophthora capsici based on RNA silencing technology

Method used

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  • Plant expression vector for targeting silence phytophthora capsici cellulose synthase 3 and application thereof
  • Plant expression vector for targeting silence phytophthora capsici cellulose synthase 3 and application thereof
  • Plant expression vector for targeting silence phytophthora capsici cellulose synthase 3 and application thereof

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0038] Example 1 The source of the sequence for inhibiting the expression of Phytophthora capsici oxysterol protein 1 gene of the present invention

[0039] The sequences in the two plant expression vectors of the present invention are derived from different intervals of the cDNA sequence of PcCes3, the sequences are shown in SEQ ID No.1 or SEQ ID No.2, and the sequences derived from the PcCes3 cDNA can cause different degrees of PcCes3 Gene silencing, therefore, can be used as a plant expression vector for enhancing plant resistance to Phytophthora capsici. The cDNA sequence of PcCes3 is derived from the gene numbered JX905357 in the Phytophthora capsici database (GenBank).

[0040] Select the 1801-2602 cDNA sequence of PcCes3 with a total of 802 base sequences (sequence 2 in the sequence listing) as the plant expression vector expression sequence of the long loop hairpin structure, the 3005-3559 total of 555 base sequences (sequence in the sequence listing 1) The expression...

Embodiment 2

[0044] Embodiment 2 Construction of plant expression vector of the present invention

[0045] The primers for amplifying the sequences involved in the present invention were entrusted to Qingke Biosynthesis. The sequences were amplified using Q5 High Fidelity DNA Polymerases (NEB), and the vectors were constructed after being recovered by gel electrophoresis.

[0046] For the short-circle hairpin structure vector, the fragment described in sequence 1 (using Phytophthora capsici as a template, using pENsenseC5F and pENsenseC5R in Table 2 as the fragment amplified by primers obtained by Cla I and EcoR I double enzyme digestion) was normalized first. Insert between the pENTR1a plasmid restriction site Cla I and EcoR I, after PCR, sequencing, enzyme digestion, electrophoresis verification, use the restriction enzyme site SacII and SpeI to convert the fragment described in sequence 1 (with Phytophthora capsici As a template, the fragment amplified with the primers pENantisenseC5F a...

Embodiment 3

[0051] Embodiment 3 utilizes the plant expression vector involved in the present invention to carry out transgenic plant construction

[0052] Test plants and bacterial liquid: aseptically cultured tobacco seedlings, Agrobacterium tumefaciens strain GV3101 containing the plant expression vector involved in the present invention.

[0053] Test medium:

[0054] (1) 1 / 2 MS solid medium: MS (Murashige&Skoog medium) 2.2g, sucrose 30g, stir to dissolve, adjust pH to 5.6-5.8 with NaOH, add 8g agar to dilute to 1L, high temperature and high pressure sterilization.

[0055](2) MS solid (liquid) pre-medium: MS 4.44g, MES 2.1235g, sucrose 30g, stir to dissolve and adjust pH to 5.8 with NaOH, add 2.4g Phytogel, 1mL 6-BA (1mg / mL) after high-temperature and high-pressure sterilization, add 1 mL of acetosyringone (AS, 100 μM), 1 mL of naphthaleneacetic acid (NAA, 1 mg / mL), and do not add plant gel to the liquid medium.

[0056] (3) Screening medium (sprouting): MS 4.44g, sucrose 30g, stir...

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Abstract

The invention discloses a plant expression vector capable of effectively interfering a phytophthora capsici cellulose synthase 3 gene through host-induced gene silencing and application thereof. The plant expression vector disclosed by the invention can be transcribed to form a double-stranded RNA with a reverse complementary structure, and the double-stranded RNA is formed by respectively inserting a nucleotide sequence shown as SEQ ID No.1 or SEQ ID No.2 into the vector twice in the forward direction and the reverse direction. The invention also discloses application of the expression vectors in preventing and treating plant diseases caused by phytophthora capsici. The plant expressing the vector provided by the invention has obviously enhanced disease resistance to phytophthora capsici, the expression of the cellulose synthase 3 gene of the phytophthora capsici infecting the plants is significantly inhibited, and the cellulose synthase 3 gene is necessary for the development and pathopoiesis of the phytophthora capsici, therefore, the plant expression vector disclosed by the invention can effectively inhibit plant infection caused by phytophthora capsici, and an effective way is provided for preventing and treating related diseases caused by phytophthora capsici.

Description

technical field [0001] The invention relates to the field of agricultural biotechnology, in particular to two plant expression vectors with different secondary structure transcripts that inhibit the expression of Phytophthora capsici cellulose synthase 3 gene, and the use of these plant expression vectors in preventing and treating plant diseases caused by Phytophthora capsici on the application. Background technique [0002] Phytophthora capsici is an important plant pathogenic oomycete, which is distributed all over the world and can infect more than 70 kinds of plants of Solanaceae, Leguminosae and most Cucurbitaceae, causing damping, wilting and root, Stem and fruit rot can cause disease from the seedling stage to the fruit stage, causing serious economic losses. Capsicum blight caused by Phytophthora capsici is a devastating disease that can spread through rain, soil and airflow, causing symptoms such as withered leaves, rotten fruits, and even dead plants, which have ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/113C12N15/82A01H5/00A01H6/82
CPCC12N15/1137C12N9/1059C12N15/8218C12N15/8282C12N2310/14
Inventor 刘西莉王治文钟珊高翔李瑜张博瑞张思聪李腾蛟
Owner CHINA AGRI UNIV
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