Thermophilic bacillus pumilus strain Tamy12 and high-temperature amylase produced by same

A technology of Bacillus pumilus and high-temperature amylase, applied in the field of applied microorganisms, can solve the problems of no Bacillus pumilus, no high-temperature amylase, etc.

Inactive Publication Date: 2009-12-23
KUNMING UNIV OF SCI & TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] So far, there are no reports of the Bacillus pumilus sp. Tamy 12 strain in the prior art, nor of the high-temperature amylase produced therefrom

Method used

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  • Thermophilic bacillus pumilus strain Tamy12 and high-temperature amylase produced by same
  • Thermophilic bacillus pumilus strain Tamy12 and high-temperature amylase produced by same
  • Thermophilic bacillus pumilus strain Tamy12 and high-temperature amylase produced by same

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0027] Isolation, screening and identification of Bacillus pumilus sp. Tamy12 strain:

[0028] Tamy12 strain was isolated from Rehai, Tengchong, Yunnan Province. The soil in Rehai, Tengchong was cultured in a strain screening liquid medium at 55°C for a short time in a shaker flask, spread on a strain screening solid medium, and cultured at 55°C for 1 to 2 days, and the strains were picked out and planted on the strain screening solid medium Above, after 3-4 days of culture at 55°C, the iodine solution was colored, and the size of the transparent circle was observed, and the amylase-producing strain Tamy12 was screened out. The strain selection liquid medium is (g / L): (NH 4 ) 2 SO 4 , 1.3; peptone, 1; KH 2 PO 4 , 0.276; MgSO 4 ·7H 2 O, 0.25; yeast extract, 1; Na 2 MoO 4 2H 2 O, 0.000025; CuSO 4 , 0.000016; MnSO 4 ·H 2 O, 0.0022; H 3 BO 3 , 0.0005; ZnSO 4 ·7H 2 O, 0.0005; CoCl 2 ·6H 2 O, 0.000046; CaSO 4 ·H 2 O, 0.06; FeSO 4 , 0.099; pH7.5. Strain screeni...

Embodiment 2

[0031] The Bacillus pumilus sp. Tamy12 strain of Example 1 was inoculated into 50 ml strain screening liquid medium, and cultured on a shaker at 55° C. for 24 hours. Inoculate into 500ml strain screening liquid medium at a ratio of 1:10, and culture on a shaker at 55°C for 4 days. The fermentation broth was centrifuged at 6500 rpm for 10 min at 4°C, the fermentation supernatant was collected, and filtered twice with filter paper. Use a 10,000MWCO membrane to perform ultrafiltration and concentrate to 50ml, and store the concentrate at 4°C.

Embodiment 3

[0033] The Native-PAGE of embodiment 2 concentrates:

[0034] ①Native-PAGE: In discontinuous electrophoresis, the concentration of the separating gel is 12% (starch solution with a final concentration of 0.1% is added to the separating gel), and the concentration of the stacking gel is 5%. Electrophoresis conditions: After adding 15 μl of sample, the voltage was adjusted to 130V, and the electrophoresis was performed at 4°C for 3 hours.

[0035] ②After electrophoresis, half of the gel was soaked in 30% Tris-Tcl buffer, incubated at 55°C for 6 hours, and stained with iodine solution. According to the nature of iodine-starch color development, there will be obvious hydrolysis circles around the amylase hydrolyzed.

[0036] ③ Gel staining and decolorization: Stain and decolorize the other half of the gel. The steps are:

[0037] a. Put the gel in a plastic box, add Coomassie Brilliant Blue R250 staining solution (submerged gel), and stain on a decolorizing shaker for 30 minute...

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Abstract

The invention discloses a bacillus pumilus novel strain Tamy12, high-temperature amylase obtained by utilizing the same through fermentation and a preparation method thereof. The colony of the Tamy12 strain is white, the surface is dry, and the edge is round; cells are in a short bar shape and are Gram positive bacteria, and the temperature range is 38-70 DEG C. Native-page analysis indicates that coarse enzyme liquid obtained by fermenting the Tamy12 contains amylase, and soluble starch can be resolved into glucose, maltose and oligosaccharide with 3-6 glucosyls. The related high-temperature amylase of the invention has higher operative temperature and wider pH adaptation range and has a certain application potential in application to the fields of the food brewing industry, the environmental protection industry, the energy utilization industry, the medicine industry and the like.

Description

technical field [0001] The invention belongs to the field of applied microorganisms. In particular, it relates to a strain of Bacillus pumilus Tamy 12, and the high temperature amylase produced therefrom. Background technique [0002] The term "Extremophiles" was proposed by MacElroy in 1974, which refers to the ability to survive normally in extreme environments (high temperature, cold, strong acid, strong alkali, high salt, high pressure, high radiation, etc.) A collective term for living microbial populations. Mainly include acidophilic microorganisms, alkaliphilic microorganisms, halophilic microorganisms, marine microorganisms, barophilic microorganisms, psychrophilic microorganisms, thermophilic microorganisms, etc. [0003] Thermophilic microorganisms are a type of microorganisms that live in high-temperature environments, such as craters and their surrounding areas, hot springs, high-temperature wastewater discharge areas of factories, etc. Extremophilic enzymes d...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20C12N9/28C12R1/07
Inventor 林连兵李秋鹏魏云林季秀玲井申荣洪伟陈波陆月情熊学权
Owner KUNMING UNIV OF SCI & TECH
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