Geotrichum candidum MF5 with effect of removing heavy metal ions, microbial inoculum and application of microbial inoculum
A technology of heavy metal ions, Geotrichum candidum, applied in the treatment of polluted groundwater/leachate, fungi, microorganism-based methods, etc., can solve problems such as limited
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Embodiment 1
[0040] Enrichment culture of fungi: Take 50 mL of acidic mine wastewater in eastern Anhui (pH is 3.67, sulfate concentration is 3870 mg / L, Fe 3+ The concentration of 398.7mg / L, Al 3+ The concentration of 912.3mg / L, Mn 2+ The concentration of 776.2mg / L, Cu 2+ The concentration of 43.9mg / L, Zn 2+ The concentration of the -1 Centrifuge at low speed to remove impurities. Pipette 1 mL of the supernatant after centrifugation and inoculate it into 50 mL of Char type liquid medium (sodium nitrate 3.00 g, potassium dihydrogen phosphate 1.00 g, magnesium sulfate 0.50 g, ferrous sulfate 0.01 g, sucrose 30.00 g, chloramphenicol 0.10 g) , distilled water 1000mL), modified Martin-style liquid medium (peptone 5.00g, yeast extract 2.00g, glucose 20.00g, potassium dihydrogen phosphate 1.00g, magnesium sulfate 0.50g, chloramphenicol 0.10g, distilled water 1000mL) and sand type Liquid culture medium (10.00 g of peptone, 40.00 g of glucose, 0.10 g of chloramphenicol, 1000 mL of distilled wat...
Embodiment 2
[0044] Morphological identification: The bacterial strain I obtained in Example 1 was observed with a high-power fluorescence microscope on structures such as colonies, hyphae, and spores, and the fungal color, size, and shape were recorded. Chinese Mycology" (Zhang Zhongyi, 2014), for identification.
[0045] Morphological identification results such as figure 1 Shown: the width of the hyphae is 3-7μm, and the hyphae are broken into single or chained, long-cylindrical, blunt-ended arthrospores after maturation. The colony spreads flat, grows fast, flat, milky white, fluffy or almost powdery, with concentric circles that can be radiated, and some are central protrusions.
Embodiment 3
[0047] Molecular biological identification: Extract the screened fungal DNA according to the Ezup column fungal genomic DNA extraction kit (SK8259) provided by Shanghai Bioengineering Co., Ltd., and perform PCR amplification on the extracted DNA. The primer is 18SrDNA universal primer NS1:5 '-GTAGTCATATGCTTGTCTC-3' (primer F, SEQ ID NO.1), NS6: 5'-GCATCACAGACCTGTTATTGCCTC-3' (primer R, SEQ ID NO.2), the sequence of PCR amplification is shown in Table 1; The increase program is shown in Table 2.
[0048] Table 1 PCR amplification sequence
[0049] reactive components Volume (μl) 10×PCR Buffer dNTP(each 10mM) Taq Plus DNA Polymerase (5U / μl) 50mM MgSO 4
12.5 in total Primer F (10mM) 1 Primer R (10mM) 1 Template(DNA) 1 ddH 2 O
9.5 Total 25
[0050] Table 2 PCR amplification program
[0051] temperature(℃) time cycle 95 5min 94 30s 57 30s 30cyc 72 90s 72 ...
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