Application of Bacillus mojavensis and method for degrading naphthalene (NAP) by using Bacillus mojavensis
A technology of Bacillus virulina and Bacillus, which is applied in the field of environmental microbial applications, to achieve significant degradation effects, simple operation, and short degradation cycle
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Embodiment 1
[0031] Identification of Example 1 bacterial strain B1811
[0032] Strain B1811 was isolated from soil, and its morphology was as follows: figure 1 As shown, it is a rod-shaped Gram-negative bacteria that can form spores. Genomic DNA was extracted, and the genomic DNA was subjected to polymerase chain reaction (Polymerase Chain Reaction, PCR) using 16s rDNA and BCR1 primers to propagate specific DNA sequences, and the National Center for Biotechnology Information (National Center for Biotechnology Information, NCBI) database for strain comparison.
[0033] (1) 16S rDNA sequence analysis:
[0034] 16s rDNA forward primer 27F: 5'-AGA GTT TGA TCC TGG CTC AG-3', as shown in SEQ ID NO.1;
[0035] 16s rDNA reverse primer 1541R: 5′-AAG GAG GTG ATC CAG CC-3′, as shown in SEQ ID NO.2;
[0036] The amplified 16S rDNA sequence is shown as SEQ ID NO: 3, and the full length of the sequence is 1461bp. The amplified 16S rDNA sequence was compared with the gene sequence of the related b...
Embodiment 2
[0042] Example 2 Preparation of Bacillus mohewei B1811 liquid shake flask culture solution
[0043] (1) Slant culture: Bacillus mohaiwei B1811 was inoculated on the slant medium and cultured at 40°C for 48 hours to obtain the slant strain. The slant medium used contained the following ingredients per 1L: 2.0g glucose, 1.0g peptone, 0.5g yeast extract, 1.8g agar, the rest of distilled water, neutral pH, and sterilized at 115°C for 20 minutes.
[0044](2) Take the slant strain and inoculate it into the sterilized seed culture medium, and culture it under the conditions of 175 rpm and 30°C for 24 hours with constant temperature and shaking to obtain the seed culture solution; the ingredients contained in each 1 L of the seed culture medium are: Ammonium sulfate 0.5g, sodium chloride 4.0g, potassium phosphate trihydrate 0.5g, magnesium sulfate heptahydrate 0.4g, agar powder 15g, yeast extract 5g, the rest of distilled water, pH 7.0, sterilized at 121°C for 25min.
[0045] (3) Add...
Embodiment 3
[0046] Example 3 NAP standard curve formulation and content determination
[0047] (1) Prepare 400mg / L naphthalene mother liquor with n-hexane as solvent, dilute 20 times, 25 times, 30 times, 35 times, 40 times, 80 times, 100 times, 200 times respectively for later use, take 2.5mL of different dilution factors The naphthalene solution is placed in a 3mL quartz cuvette, and the absorbance value under the 275nm wavelength is measured respectively with a UV spectrophotometer, and the ordinate is used as the absorbance value, and the concentration of naphthalene is used as the abscissa to draw a standard curve; the standard curve is as follows figure 2 Shown; and then obtain the absorbance value-concentration equation: y=0.0424x-0.0033, x is the concentration of naphthalene, and y is the absorbance value.
[0048] (2) Take 2.5mL of the fermentation broth obtained in Example 1 and put it in a 3mL quartz cuvette, measure the absorbance at 275nm wavelength with a UV spectrophotomete...
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