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Broad-spectrum and highly effective new Bacillus thuringiensis strain, its bacterial agent and application

A technology of Bacillus thuringiensis and Bacillus aureus, applied in the fields of application, bacteria, biocide, etc., can solve the problems of isolation and application of Bacillus thuringiensis, few reports, and few Bt strains

Inactive Publication Date: 2013-06-19
HEBEI AGRICULTURAL UNIV.
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the isolation and application of broad-spectrum and high-efficiency Bacillus thuringiensis are still rarely reported, especially the Bt strains against Lepidoptera pests on fruit trees and vegetables

Method used

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  • Broad-spectrum and highly effective new Bacillus thuringiensis strain, its bacterial agent and application
  • Broad-spectrum and highly effective new Bacillus thuringiensis strain, its bacterial agent and application
  • Broad-spectrum and highly effective new Bacillus thuringiensis strain, its bacterial agent and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0056] Example 1. Isolation and identification of Bacillus thuringiensis YX-1

[0057] The inventors isolated the Bt strain YX-1 from soil by themselves, and the soil comes from Yi County, Baoding City, Hebei Province.

[0058] Weigh 10g of soil sample into 50mL sodium acetate liquid medium (peptone 0.5g, yeast powder 0.25g, NaCl 0.5g, sodium acetate 1.8g, water 50mL, adjust the pH to 7.2-7.4, 121℃, autoclave Add 400μg / mL penicillin sodium salt to a 30min) shake flask, and incubate on a shaker (200rpm / min, 30℃) for 4h. After the incubation, take 10 mL of the soil suspension and add it to a sterile centrifuge tube at 308 g and centrifuge for 15 min. Take 1.5 mL of the upper turbid liquid and bath in a 65 ℃ water bath for 15 min. Take 0.2 mL of the heat-treated turbid liquid and coat it with LB solid medium (peptone 2g, Yeast powder 1g, NaCl 2g, agar powder 3.2g, water 200mL, adjust pH to 7.2-7.4, 121°C, autoclave for 30min) plate, put the plate in a 30°C incubator. 3-4d Pick a sme...

Embodiment 2

[0061] Example 2. Observation of the growth characteristics of Bacillus thuringiensis YX-1

[0062] Inoculate Bt YX-1 in LB liquid medium for activation overnight, inoculate the bacteria in LB liquid medium according to 1% of the inoculum, culture with shaking (28℃, 230rpm / min), take samples every 2h to determine OD 600 , A total of 24h. At the same time, the pH value of the culture broth was measured, and the growth and development of the strains during the fermentation process were observed.

[0063] Bacillus thuringiensis YX-1 grows well in LB liquid medium and can normally produce spores and parasporal crystals within 24 hours of the growth cycle. Sampling every 2h to determine the OD of the culture medium 600 Value and pH value to draw growth curve and pH change curve (see figure 2 ), and observe the development of the strain during the fermentation process. Among them, 0~4h is the incubation period, and the pH value does not change much; 4~14h is the logarithmic growth peri...

Embodiment 3

[0064] Example 3. Genotype identification of Bacillus thuringiensis YX-1

[0065] Bt YX-1 total DNA extraction refer to Narva (Novel Bacillus thuringiensis microbes active against nematodes,and genes encoding novel nematodes-active toocin from Bacillus thuringiensis isolates. European Patent Office. 1991, EP0462721) method.

[0066] Using the Bt cry gene identification system (Identification of novel cry-type genes from Bacillus thuringiensis strains on the basis of restriction fragment length polymorphism of the PCR-amplified DNA. Appli- ed and Environmental Microbiology.1996,62:1369-1377; Establishment of PCR-RFLP identification system for cry gene of Bacillus thuringiensis. Chinese Agricultural Sciences, 1998, 31(3):13-18), using the total DNA of Bt YX-1 as a template, using 31 pairs of universal primers for PCR amplification, the results show that Bt YX-1 contains cry1 , cry2 , cry1I , vip3A , cry34-35 Gene (see image 3 ). Four pairs of primers K5un2 / K3un2, K5un3 / K3un3...

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Abstract

Belonging to the field of biological control of pests, the invention discloses a Bacillus thuringiensis strain that has a broad spectrum and is highly effective on lepidoptera pests, preparation of its wettable powder and application. The Bacillus thuringiensis YX-1 strain involved in the invention is preserved in China General Microbiological Culture Collection Center, and the preservation number is CGMCC No. 5256. Containing a plurality of insecticidal genes, the strain has a highly effective lethal effect on Cydia molesta, cotton bollworm, fall webworms, Adoxophyes orana Fisher von Roslerstamm, Phalera flavescens Bremer et Grey, carpocapsa pomonella, prodenia litura, plutella xylostella, beet armyworm and other lepidoptera pests. The invention also discloses a method for preparation of wettable powder through the YX-1 strain. The Bacillus thuringiensis YX-1 strain and its wettable powder provided in the invention are mainly used for prevention and treatment of lepidoptera pests on various fruit trees and vegetables, also can be used for prevention and treatment of lepidoptera pests on field crops and garden plants, and have the characteristics of broad spectrum, high efficiency, safety to people, livestock as well as animals and plants, and no environmental pollution, etc.

Description

technical field [0001] The invention relates to a strain of bacillus thuringiensis, and also relates to a microbial insecticide prepared by using the bacillus thuringiensis, a preparation method and an application, and belongs to the field of pest biological control. Background technique [0002] Lepidoptera belongs to the subclass Pteroptera and holometabolism. About 200,000 species are known in the world, and more than 8,000 species are known in China. This order is the second largest order after Coleoptera in Insecta, with a wide range of distribution and the most abundant species in the tropics. This order includes two types of insects, moths and butterflies, which harm food, cotton, oil plants, vegetables, tobacco, flowers, pastures, lawns, fruit trees and other plants. Almost every crop is affected by one or more of them Lepidoptera pests harm. In order to control the harm of these pests, a large number of highly toxic and high-residue chemical pesticides are used un...

Claims

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Application Information

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IPC IPC(8): C12N1/20A01N63/02A01N25/14A01P7/04C12R1/07
Inventor 王勤英苏俊平南宫自艳宋萍曹克强
Owner HEBEI AGRICULTURAL UNIV.
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