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Method for biologically synthesizing nano-selenium through bacillus subtilis and application of method

A technology of Bacillus subtilis and nano-selenium, applied in the direction of microorganism-based methods, biochemical equipment and methods, applications, etc., can solve the problems of non-factory production and application, and achieve high yield, significant effect, and environmental friendliness Effect

Active Publication Date: 2016-12-07
北京华美源生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, the existing reports have not carried out the research on the factory production and application in the later stage.

Method used

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  • Method for biologically synthesizing nano-selenium through bacillus subtilis and application of method
  • Method for biologically synthesizing nano-selenium through bacillus subtilis and application of method
  • Method for biologically synthesizing nano-selenium through bacillus subtilis and application of method

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Experimental program
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Effect test

Embodiment 1

[0045] Example 1 Isolation, purification and identification of Bacillus subtilis S12

[0046] 1.1 Isolation and purification of strain S12

[0047] A strain S12, which can tolerate higher concentration of sodium selenite, was isolated from the soil.

[0048] 1.2 Identification of strain S12

[0049] 1.2.1 PCR amplification of 16S rDNA and gyrA gene sequence and sequencing:

[0050] Inoculate strain S12 on LB solid medium and culture for 24h, take 0.2mL sterilized PCR tube, add 10μL ddH 2 O,

[0051] Pick up a single colony with a sterile toothpick and put it into a PCR tube and mix well.

[0052] 1.2.2 Construct the PCR reaction system:

[0053] 16S rDNA: Using 8F (5'-CGGGATCCAGAGTTTGATCCTGGCTCAGAACGAACGCT-3') and 1506R (5'-CGGGATCCTACGGCTACCTTGTTACGACTTCACCCC-3') as primers, the 16S rDNA gene sequence was amplified by PCR. The PCR reaction system is: ddH 2 O, 18.5 μL; 10×Buffer, 2.5 μL, dNTPMix, 2 μL; 8F, 0.5 μL; 1506R, 0.5 μL; bacterial solution, 0.5 μL; rTaq DNA poly...

Embodiment 2

[0058] Embodiment 2 Bacillus subtilis S12 is to the tolerance concentration of sodium selenite

[0059] 2.1 Prepare solid LB with different concentrations of selenium-containing medium (each liter medium contains 10g of NaCl, 10g of tryptone, 5g of yeast extract, 15g of agar, 1L of deionized water), autoclave at 121°C for 20min; prepare 1M selenous acid The sodium mother liquor was sterilized by filtration, and sodium selenite solution was added to make the sodium selenite content in the culture medium 0mM, 10mM, 25mM, 35mM, 55mM, 70mM, 80mM respectively.

[0060] 2.2 Pick a single colony of the S12 strain and inoculate it in 5ml LB liquid medium for 8 hours (150rpm, 28°C) and shake it for 8 hours to take the above bacterial solution and dilute it to OD 600 =0.8 of the mother liquor for later use; the mother liquor was diluted to 10 -2 、10 -3 、10 -4 、10 -5 、10 -6 , respectively drop 2.5 μL of bacterial solutions of different concentrations on the selenium-containing plate...

Embodiment 3

[0062] Embodiment 3 Bacillus subtilis S12 is to the conversion efficiency of sodium selenite

[0063] 3.1 Prepare different concentrations of selenium-containing liquid LB medium, take 5mL and divide it into test tubes, and sterilize under high pressure at 121°C for 20 minutes; prepare 1M sodium selenite mother solution, filter and sterilize, add sodium selenite solution, and make the medium The contents of sodium selenite were 1mM, 3mM, 5mM, 7mM, 10mM, 15mM, 20mM, respectively, and each concentration gradient was repeated three times.

[0064] 3.2 Pick a single colony of the S12 strain and inoculate it in 5ml LB liquid medium for shaking culture for 8 hours (150rpm, 28°C) to take the above bacterial solution and dilute to OD 600 =0.8; the diluted bacterial solution was inoculated in LB medium (containing Na 2 SeO 3 ), shake and cultivate for 48 hours.

[0065] 3.3 Prepare 1M Na with distilled water 2 S solution (prepared and used now); take 1ml of bacteria and nano-seleni...

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Abstract

The invention provides a method for biologically synthesizing nano-selenium through bacillus subtilis and application of the method. Bacillus subtilis S12 capable of tolerating high-concentration sodium selenite is separated from soil, biological nano-selenium is synthesized through the strain S12 and then separated and purified, and a large amount of biological nano-selenium is prepared and applied to fertilizer, feed, selenium-rich functional food processing, healthcare products and medicine products. Nano-selenium is prepared through a biological fermentation process, the method has the advantages of being friendly to the environment, high in yield, safe, efficient and the like, and the produced biological nano-selenium is used for selenium-rich fertilizer and selenium-rich feed and has a remarkable selenium enriching effect on crops, fruits, vegetables, meat, milk and egg after applying and feeding.

Description

technical field [0001] The invention relates to the technical field of microbiology and biological nano-selenium preparation, in particular to a method for biosynthesizing nano-selenium by using Bacillus subtilis and its application. Background technique [0002] Selenium (Selenium) element is an essential trace element for animals and humans, and is related to various metabolic pathways in the human body. Selenium deficiency in the human body can cause various diseases, such as muscle atrophy, cardiovascular, bone or immune system diseases, and increase the risk of cancer, while excessive selenium can also cause serious poisoning to the human body. The human body has a relatively narrow range of adaptation to selenium. The Chinese Nutrition Society and the FAO / WHO / IAEA Joint Expert Committee determined that the appropriate range of human intake is 60-250 μg / d, the safe dose is 400 μg / d, and the toxic dose is 800 μg / d. Since there is a natural selenium-deficient belt in my ...

Claims

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Application Information

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IPC IPC(8): C12N1/20C12P3/00A23L33/16A23L7/17A23L29/30A23L33/15A23P10/30A23K20/20A23K10/30A23K20/147A23K10/37A23K10/22A23K10/18A23K20/26A61K33/04C05D9/02C12R1/125
CPCA23V2002/00A61K33/04C05D9/02C12P3/00C12N1/205C12R2001/125A23V2200/30A23V2250/5118A23V2250/712Y02P60/87
Inventor 郭岩彬李奎吴文良赵桂慎李柯谢斌卢鹏王昊
Owner 北京华美源生物科技有限公司
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