Clostridium acetobutylicum capable of co-utilizing glucose and xylose and application of clostridium acetobutylicum
A kind of technology of Clostridium acetobutylicum and glucose, applied in the field of microorganisms
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Embodiment 1
[0034] Mixed sugar (glucose and xylose 1:1) as carbon source Clostridium acetobutylicum Isolation and screening of LY4:
[0035] Weigh 1g of the soil sample collected from Jinghu Lake of Nanjing University of Technology, dilute it with normal saline, pipette 200μL onto a plate with mixed sugar as the only carbon source, place it in an anaerobic incubator at 37°C for 5 days, and streak and purify the grown colony for five generations , screen out the strains that can utilize glucose and xylose mixed sugars, and carry out anaerobic culture on the screened strains, and use liquid phase to detect the daily consumption of glucose and xylose, so as to obtain the co-utilization of mixed sugars (glucose and xylose sugar 1:1) strain.
[0036] The medium formula of the above plate is NaCl 1 g / L, KH 2 PO4 0.75 g / L, K 2 HPO 40.75 g / L yeast powder 3 g / L, CaCl 2 2H 2 O 0.015 g / L, FeCl 2 4H 2 O 1.5 g / L, KCl 0.3 g / L, adjust the pH to 5.0-6.0, carbon source 60 g / L, add agar powder 15-...
Embodiment 2
[0038] Mixed sugar (glucose and xylose 1:1) as carbon source Clostridium acetobutylicum Identification of LY4 and its growth characteristics:
[0039] Identification of LY4:
[0040] For 16S rDNA identification: use primer 27F:5 ， -AGAGTTTGATCCTGGCTCAG-3 , and 1492R: 5 ， -TACCTTGTTACGACTT-3 , The LY4 16S rDNA of the amplified strain was connected to the cloning vector pMD19T by T / A cloning to construct the recombinant cloning vector pMD19T-16S, which was transformed into the cloning host strain Escherich coil DH5α, to obtain recombinant microorganisms Escherich coil DH5α (pMD19T-16S), sequence the exogenous fragment of the obtained recombinant microorganism, compare the 16S rDNA sequence with the NCBI database, and identify strain LY4 to Clostridium acetobutylicum Fungus. Its 16S rDNA nucleotide sequence is shown in SEQ ID NO:1.
[0041] LY4 growth and metabolic characteristics:
[0042] Strain LY4 can grow well at 37°C and grow well at pH 5.0-6.0 (preferably ...
Embodiment 3
[0044] carbon source Clostridium acetobutylicum LY4 utilizes the growth and fermentation characteristics of mixed sugar in different proportions:
[0045] strain Clostridium acetobutylicum LY4 can use different proportions of mixed sugars as carbon source for growth ( figure 1 , image 3 , Figure 5 ), strain Clostridium acetobutylicum LY4 Pick the strain LY4 from the plate and inoculate it into the fermentation medium, culture at 37°C with 120r / min shaking, adjust the pH to 5.0-6.0 every 24h, and measure the concentration of various products with GC after 72h. After 120 hours of fermentation, the concentration of the fermentation product was as follows: figure 2 , Figure 4 , Image 6 .
[0046] The above fermentation medium formula is NaCl 1 g / L, KH 2 PO4 0.75 g / L, K 2 HPO 4 0.75 g / L, yeast powder 3g / L, CaCl 2 2H 2 O 0.015 g / L, FeCl 2 4H 2 O 1.5 g / L, KCl 0.3 g / L, adjust the pH to 5.5, carbon source 60g / L, blow nitrogen for 10-20min, and sterilize at 120°C ...
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