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Method for biosynthesis of nano-selenium by utilization of Agrobacterium tumefaciens and application thereof

A technology based on Agrobacterium tumefaciens and nano-selenium, applied in the direction of microorganism-based methods, biochemical equipment and methods, applications, etc., can solve problems such as human hazards, and achieve environmental friendliness, high yield, and remarkable effects

Active Publication Date: 2017-04-19
CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Excessive selenium can also cause harm to the human body

Method used

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  • Method for biosynthesis of nano-selenium by utilization of Agrobacterium tumefaciens and application thereof
  • Method for biosynthesis of nano-selenium by utilization of Agrobacterium tumefaciens and application thereof
  • Method for biosynthesis of nano-selenium by utilization of Agrobacterium tumefaciens and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0048] Example 1 Isolation, purification and identification of Agrobacterium tumefaciens S2-323

[0049] 1. Isolation and purification of strain S2-323

[0050] A strain S2-323 that can tolerate higher concentrations of selenite and selenate was isolated from the soil.

[0051] 2. Identification of strain S2-323

[0052] (1) PCR amplification of 16S rRNA gene sequence and sequencing:

[0053] Inoculate strain S2-323 on LB solid medium and culture for 24 hours, take 0.2mL sterilized PCR tube, add 10μL ddH 2 O, a sterile toothpick picked a single colony into the PCR tube and stirred to mix.

[0054] (2) Construct the PCR reaction system:

[0055] 16S rRNA: Using 8F (5'-CGGGATCCAGAGTTTGATCCTGGCTCAGAACGAACGCT-3') and 1506R (5'-CGGGATCCTACGGCTACCTTGTTACGACTTCACCCC-3') as primers, the 16S rRNA gene sequence was obtained by PCR amplification. The PCR reaction system is: ddH 2 O, 18.5 μL; 10×Buffer, 2.5 μL, dNTP Mix, 2 μL; primer 8F, 0.5 μL; primer 1506R, 0.5 μL; bacterial solut...

Embodiment 2

[0058] Embodiment 2 Agrobacterium tumefaciens S2-323 is to the tolerance concentration of selenite

[0059] Prepare solid LB selenium-containing medium with different concentrations (per liter medium contains 10g NaCl, 10g tryptone, 5g yeast extract, 15g agar, 1L deionized water), autoclave at 121°C for 20min; prepare 1M selenite The mother liquor is sterilized by filtration, and a selenite solution is added to make the selenite contents in the medium respectively 0mM, 50mM, 75mM, and 100mM.

[0060] Pick a single colony of the S2-323 strain and inoculate it in LB liquid medium for 8 hours (150rpm, 28°C), and 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 -1 、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, each concentration has 6 replicates, culture at 28°C for 48 hours, and observe the ...

Embodiment 3

[0062] Embodiment 3 Agrobacterium tumefaciens S2-323 is to the tolerance concentration of selenate

[0063] 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, and 1L of deionized water), and autoclave at 121°C for 20min; prepare selenate mother liquor, filter Sterilize, add selenate solution, make the selenate content in the culture medium 0mM, 200mM, 300mM, 400mM, 500mM respectively.

[0064] Pick a single colony of the S2-323 strain and inoculate it in LB liquid medium for 8 hours (150rpm, 28°C), and take the above bacterial solution and dilute it to OD 600 =0.8 mother liquor, then dilute the mother liquor to 10 -1 , respectively drop 2.5 μL of bacterial solution on different concentrations of selenium-containing plates, each concentration was replicated three times, cultivated at 28°C for 48 hours, and observed the growth of colonies and color changes.

[...

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Abstract

The invention provides a method for biosynthesis of nano-selenium by the utilization of Agrobacterium tumefaciens and application thereof. According to the method, Agrobacterium tumefaciens S2-323 capable of tolerating high-concentration selenite and selenate is separated from soil; the bacterial strain S2-323 is utilized to synthesize bio-nano-selenium and the bio-nano-selenium is purified by separation; and the bio-nano-selenium is prepared in quantity and applied in fields of fertilizer, feed, selenium-enriched functional food processing, health care products and pharmaceutical products. The biological fermentation technology for preparation of nano-selenium is environment-friendly, has high yield, and is safe and efficient. The bio-nano-selenium obtained by production is used in selenium-enriched fertilizers and selenium-enriched feeds. After fertilization or feeding, crops, melons and fruits, vegetables and meat, milk and eggs have remarkably selenium-enrichment effect.

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 using non-pathogenic Agrobacterium tumefaciens and its application. Background technique [0002] Selenium (Selenium, Se) element is one of the essential trace elements in many organisms, and it is an essential component of various selenium-containing enzyme proteins such as thioreductase, deiodinase, and glutathione peroxidase in organisms, and participates in Various metabolic pathways in the human body. Studies have found that selenium deficiency in the human body can lead to various diseases and increase the risk of cancer. my country is rich in selenium resources, but the distribution of selenium in nature is extremely uneven, resulting in selenium deficiency in more than two-thirds of my country. The Chinese Nutrition Society and the FAO / WHO / IAEA Joint Expert Committee determined that...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20C12P3/00A23K20/20A23L33/16C05D9/02A61K33/04C12R1/01
CPCA23V2002/00A61K33/04C05D9/02C12N1/20C12P3/00C12N1/205C12R2001/01A23V2250/1626
Inventor 郭岩彬李奎陈志蓥赵桂慎吴文良李柯谢斌王浩阳方勇胡秋辉
Owner CHINA AGRI UNIV
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