Method for inducing phytophthora capsici to generate toxic secretory proteins

A technology of Phytophthora capsici and protein secretion, applied in the direction of microorganism-based methods, biochemical equipment and methods, microorganisms, etc., can solve problems such as limiting in-depth research on pathogenic mechanisms

Active Publication Date: 2015-08-12
INST OF PLANT PROTECTION FAAS
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, the current research shows that the lack of a method to induce Phytophthora capsici to secrete a large amount of toxic proteins limits our in-depth study of its pathogenic mechanism

Method used

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  • Method for inducing phytophthora capsici to generate toxic secretory proteins

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Embodiment 1

[0014] Scrape off the surface of the base of the stem of the diseased plant stem of pepper blight with a scalpel, and then rinse it with tap water. After natural air drying, take a 0.5 cm tissue block from the junction of the disease and health and place it on the V8 medium. Cultured in the dark at 25°C for 4 days, the 15% (final concentration) solid V8 medium mainly consisted of the following parts: V8 vegetable juice (purchased from Campbell Soup Company, USA) 150 mL, CaCO 3 0.2 g, 20 g agar, 1000 mL distilled water; Inoculate the isolated weak pathogenic strain FZ-1 of Phytophthora capsici (self-isolated) on the V8 medium, and culture it in the dark at 25°C for 4 days until the formation of Larger colonies; transfer 20 plates with a diameter of 5 mm to 150 mL toxic protein liquid induction medium, culture at 25°C, 150 rpm shaking for 18 days, toxic secretory protein liquid induction medium Main component: yeast extract 0.6 g, KH 2 PO 4 0.6 g, MgSO4.7H 2 O 0.25 g, VB1 0...

Embodiment 2

[0016] Scrape off the surface of the base of the stem of the diseased plant stem of pepper blight with a scalpel, and then rinse it with tap water. After natural air drying, take a 0.5 cm tissue block from the junction of the disease and health and place it on the V8 medium. Cultivate in the dark at 25°C for 3 days, inoculate the isolated strong pathogenic strains ND-2 and XM-3 of Phytophthora capsici into 15% solid V8 medium (V8 vegetable juice 150 mL, CaCO 3 0.2 g, 20 g agar and 1000 mL distilled water), cultured at 25°C in the dark for 3 days until larger colonies were formed; transfer 20 plates of bacteria with a diameter of 5 mm to 150 mL toxic protein liquid to induce culture medium, at 25°C, 150 rpm shaking culture for 20 days, toxic secreted protein liquid induction medium main components: yeast extract 0.8 g, KH 2 PO 4 0.7 g, MgSO4.7H 2 O0.3 g, VB10.002 g, glucose 30 g, asparagine 1 g, ?-sitosterol 0.02 g, set the volume to 1000 ml; filter with Whatman filter ...

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Abstract

The invention relates to a method for inducing phytophthora capsici to generate toxic secretory proteins. The method comprises the following steps: bringing pepper phytophthora blight tissues back to a laboratory, scraping off the basal part surfaces of stems of diseased plants by a surgical knife blade at first, then washing the diseased plants by running water, taking 0.5cm of tissue blocks from disease-health junctions and placing the tissue blocks in a V8 culture medium, culturing for 3-4 days under a 25 DEG C dark condition until a large bacterial colony is formed, transferring a bacterial disk into a toxic protein liquid induction culture medium, carrying out shaking culture for 15-20 days, filtering mycelia by filter paper, centrifuging the filtrate for 30min at a speed of 8000rp, adding saturated ammonium sulphate in the supernatant, centrifuging the filtrate for 30min at a speed of 12000rp, carrying out a dialysis treatment on a precipitate, freeze-drying to obtain the toxic proteins secreted by the phytophthora capsici, and storing the toxic proteins at -70 DEG C. The method is capable of inducing the phytophthora capsici to generate lots of the toxic secretory proteins, thus filling in the gap that lots of toxic secretory proteins cannot be obtained in the domestic and overseas phytophthora capsici pathogenesis researches, and having great significance.

Description

technical field [0001] The invention relates to the technical field of plant protection, in particular to a method for inducing Phytophthora capsici to produce toxic secretion protein. Background technique [0002] Pepper blight is a devastating fungal disease that occurs worldwide. It was first discovered in New Mexico, USA in 1918. Phytophthora capsici caused by pepper blight Phytophthora capsici Leonian was named by Leon Leonian in 1922, and it belongs to plant pathogenic oomycetes. Phytophthora capsici overwinters in diseased soil as oospores or chlamydospores, and can survive for several months or even longer. It has a wide range of hosts and mainly infects many important vegetable crops such as peppers, tomatoes, eggplants, cucumbers, and pumpkins. Capsicum blight can occur in both open fields and protected areas. The prevalence of the disease has caused serious losses of peppers. The general rate of diseased plants is 15-30%, and it can reach more than 80% in sever...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12P21/00C12R1/645
Inventor 刘裴清陈庆河李本金丁雪玲翁启勇
Owner INST OF PLANT PROTECTION FAAS
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