Method for biological synthesis of nano-selenium by using silver-resistant Bacillus edaphicus and application thereof
A soil bacillus and nano-selenium technology, applied in the direction of microorganism-based methods, biochemical equipment and methods, applications, etc., can solve problems such as human hazards, achieve environmental friendliness, significant selenium-enriching effect, and high yield
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Embodiment 1
[0049] Example 1 Isolation, purification and identification of silver-resistant Agrobacterium R5-41
[0050] 1. Isolation and purification of bacterial strain R5-41
[0051] A strain R5-41 that can tolerate higher concentrations of selenite and selenate was isolated from the soil.
[0052] 2. Identification of strain R5-41
[0053] 1.2.1 PCR amplification of 16S rRNA gene sequence and sequencing:
[0054] Inoculate strain R5-41 on LB solid medium and culture for 48 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.
[0055] 1.2.2 Construct the PCR reaction system:
[0056] 16S rRNA: Using 8F (5'-CGGGATCCAGAGTTTGATCCTGGCTCAGAACGAACGCT-3') and 1506R (5'-CGGGATCCTACGGCTACCTTGTTACGACTTCACCCC-3') as primers, the 16S rRNA gene sequence was amplified by PCR. 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 solu...
Embodiment 2
[0059] Embodiment 2 The tolerance concentration of silver-resistant Agrobacillus R5-41 to selenite
[0060] 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 0 mM, 100 mM, 120 mM, 150 mM, and 200 mM.
[0061] Pick a single colony of the R5-41 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 -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...
Embodiment 3
[0063] Embodiment 3 silver-resistant Agrobacterium R5-41 is to the tolerant concentration of selenate
[0064] 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 selenate mother liquor, filter Sterilize, add selenate solution, make the selenate content in the culture medium 0mM, 100mM, 150mM, 200mM, 250mM, 300mM respectively.
[0065] Pick a single colony of the R5-41 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 -2 、10 -3 、10 -4 、10 -5 、10 -6 、10 -7 , respectively drop bacterial solution on different concentrations of selenium-containing plates, each concentration has 6 replicates, culture at 28°C for 48 hours, and observe the growth of bacterial colonies and color chang...
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