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Nano-silver synthetic protein derived from lysinibacillus sphaericus and application of nano-silver synthetic protein

A technology of bacillus and nano-silver, which is applied in the field of protein purification technology and genetic engineering, and can solve problems such as environmental or biological hazards

Active Publication Date: 2020-02-18
HUAIYIN INSTITUTE OF TECHNOLOGY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The reagents used in chemical reduction may have harmful effects on the environment or organisms during use. Therefore, an environmentally friendly and green silver nanoparticle preparation method is urgently needed

Method used

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  • Nano-silver synthetic protein derived from lysinibacillus sphaericus and application of nano-silver synthetic protein
  • Nano-silver synthetic protein derived from lysinibacillus sphaericus and application of nano-silver synthetic protein
  • Nano-silver synthetic protein derived from lysinibacillus sphaericus and application of nano-silver synthetic protein

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] Preparation of sample strain fermentation supernatant. Take out the sample strain (Lysinibacillus sphaericus) and put it into the ice box to slowly thaw, inoculate 2% of the inoculation amount into a 250mL Erlenmeyer flask containing 100mL liquid seed medium, and culture it on a shaker at 37°C and 200rpm for 24h. Then inoculate the above seed liquid into the fermentation medium according to the inoculum amount of 2%. After fermenting on a shaker at 37° C. and 200 rpm for 48 hours, measure the pH of the fermentation broth. Use a desktop high-speed refrigerated centrifuge to centrifuge at room temperature and 10,000 rpm for 10 minutes, and collect the supernatant of the fermentation broth.

Embodiment 2

[0032] Using the ammonium sulfate fractional precipitation method, the supernatant protein of the fermentation broth was preliminarily purified. Make 7 groups (20%, 40%, 50%, 60%, 70%, 80%, 90%) saturation, each group 100mL fermentation supernatant. Slowly add a corresponding amount of ammonium sulfate solid to each reaction system according to the calculation table for adjusting the saturation of the ammonium sulfate solution. Stir overnight on a magnetic stirrer at 4°C to fully precipitate the protein. The resulting solution was centrifuged at 10,000×g for 30 min, and the supernatant and precipitate were kept in groups. Dissolve the precipitated protein obtained by salting out at various saturations in a small amount (about 10ml) of ultrapure water, put the precipitate into a dialysis bag to dialyze the pure water system for 24 to 48 hours, and change the dialysis buffer every 3 to 6 hours Once (use a dropper or pipette gun to pipette a small amount of solution with BaCl ...

Embodiment 3

[0034] The synthesis ability of nano-silver was measured for the dialyzed protein solution at various saturation levels. Through continuous exploration and experimentation, changing the concentration of silver nitrate substrate, temperature, pH and other influencing factors, optimizing the synthesis conditions, the final reaction conditions were determined to be: 5mL dialyzed protein solution (concentration of about 50ug / mL, pH 9.0-12.0)+AgNO 3 (Substrate concentration 15-20mM) Reaction (water-bath shaker 35-45°C, rotational speed 150-200r) for 24h.

[0035] Prepare 7 test tubes, adjust the concentration of 7 groups of dialyzed protein solutions at different saturations to about 50ug / mL in 5ml systems, react according to the above reaction conditions for 24h, and then take the upper liquid. Dilute the appropriate multiple according to the state (20 and 40 saturations do not need to be diluted, 50 and 60 saturations are diluted 10 times, 70 and 80 saturations are diluted 50 tim...

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Abstract

The present invention discloses a nano-silver synthetic protein derived from lysinibacillus sphaericus, also discloses a specific sequence of the nano-silver synthetic protein, discloses a synthesis mechanism for preparing nano-silver from the lysinibacillus sphaericus, and successfully amplifies corresponding DNA. A large number of bacteria with capacity of efficiently synthesizing the biologicalnano-silver can be prepared through recombinant vectors and prokaryotic expression, and the nano-silver synthetic protein has extremely wide application prospects and industrial value.

Description

technical field [0001] The invention belongs to the technical fields of protein purification technology and genetic engineering, and specifically relates to a nano-silver synthetic protein derived from Bacillus sphaeroides and its application. Background technique [0002] Nanomaterials are solid materials composed of nanoparticles. Since the concept of nanomaterials was formed in the 1980s, this material has attracted great attention. Metal nanomaterials are an important branch of nanomaterials. , copper as the representative, of which nano-silver research results are the most. Nano-silver material is an important new type of nano-material, and its preparation methods mainly include physical, chemical and biological methods. With the popularity of green and environmental protection concepts, the biological method of synthesizing nano-silver particles using biological systems has attracted much attention. Under the protection of biological molecules, the stability and disp...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/32C12N15/70C12N1/21C12P3/00B82Y40/00C12R1/19
CPCC07K14/32C12N15/70C12P3/00B82Y40/00
Inventor 李相前汪强刘培田霞
Owner HUAIYIN INSTITUTE OF TECHNOLOGY
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