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Clostridium beijerinckii strain and applications thereof

A technology of Clostridium beijerinckii and microbial strains, applied in the field of microorganisms, can solve problems such as competition with people for food and food shortages, and achieve the effects of good stability and increased total solvent output

Active Publication Date: 2016-06-29
CHINA PETROLEUM & CHEM CORP +1
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Problems solved by technology

[0005] Due to the large population in my country, the production of butanol by fermentation with traditional raw materials (corn and sugarcane) will inevitably cause competition for food and food shortages. Therefore, it is important to carry out biotransformation of renewable biomass materials as raw materials Research directions in line with national conditions

Method used

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  • Clostridium beijerinckii strain and applications thereof

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Comparison scheme
Effect test

Embodiment 1

[0026] The ultraviolet mutagenesis screening process of embodiment 1 Clostridium beijerinckii CM20 bacterial strain

[0027] Clostridium beijerinckii Clostridium beijerinckii NCIMB8052 (purchased from the British National Center for Industrial, Marine and Food Culture Collection) was subjected to ultraviolet mutagenesis, and was screened on a starch-glucose solid plate to obtain high-activity strains, and then screened by anaerobic fermentation to obtain target strains with high production of butanol, acetone, and ethanol .

[0028] The specific process is as follows: Clostridium beijerinckii NCIMB8052 is the starting strain. First, it is cultivated in P2 (glucose as the substrate) liquid fermentation medium. After growing to the exponential phase, take 600 microliters of bacterial liquid and carry out mutagenesis under anaerobic conditions. The ultraviolet mutagenesis condition is UV : 15W, height: 10cm, time: 2-3min. After the mutagenization, the bacterial solution was ap...

Embodiment 2

[0034] The passage stability of embodiment 2 Clostridium beijerinckii CM20 bacterial strains

[0035] In the P2 fermentation medium with glucose as the carbon source, the passage stability of Clostridium beijerinckii CM20 was detected.

[0036] In the 7 consecutive transfer cultures, CM20 used glucose as the carbon source to produce butanol and the total solvent were relatively stable, and had good passage stability. The specific results are shown in Table 2.

[0037] The passage stability test result of table 2 mutant strain CM20

[0038]

[0039] When glucose was used as the carbon source, the total solvent yield was 15.3-17.6g / l, and the butanol yield was 10.6-12.1g / l, which indicated that the genetic traits of the mutant strain CM20 were stable.

Embodiment 3

[0040]Sequence identification and bacterial strain identification of embodiment 3 mutant strain CM2016sRNA

[0041] Cultivate the mutant strain CM20 in the P2 fermentation medium with glucose as the carbon source, spread the plate during the logarithmic growth phase, and obtain a single clone, pick a single clone into a PCR tube with a sterile toothpick as a PCR template, and perform 16sRNA sequence expansion and primers Use universal primers, 27F: AGAGTTTGATCMTGGCTCAG and 1492r: TACGGYTACCTTGTTACGACTT. The PCR cycle conditions are: 95°C, 10min; 95°C, 20s; 55°C, 20s; 72°C, 1min; 32 cycles; 72°C, 10min. The amplified bands were sent to Huada Gene Company for sequencing, and the sequences were compared with the NCBI website. The sequences and Clostridium beijerinckii The 16sRNA of NCIMB8052 has 100% similarity, proving that the mutant strain is Clostridium beijerinckii .

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Abstract

The invention discloses a clostridium beijerinckii strain CM 20 with high yield of butanol, wherein the classification name of the clostridium beijerinckii strain CM 20 is Clostridium beijerinckii, and the clostridium beijerinckii strain CM 20 is preserved in the China General Microbiological Culture Collection Center (CGMCC) on June 17, 2014, and has the preservation number CGMCC No.9354. The clostridium beijerinckii strain CM 20 provided by the invention can be used for producing butanol with high yield through fermentation by utilizing lignocellulose enzymatic hydrolysate detoxified by calcium hydroxide, the total solvent yield can achieve 19g / l, the sugar utilization ratio is high, the yield of butanol is high, and the problems that the capacity of the strain and the raw materials are insufficient when the traditional biological fermentation method is adopted for producing butanol are solved.

Description

technical field [0001] The invention belongs to the technical field of microbes, and in particular relates to a high-butanol-producing Clostridium beijerinckii and its application in producing butanol by fermenting lignocellulosic raw materials. Background technique [0002] Butanol is an important organic chemical raw material, which is widely used in chemical, pharmaceutical and petroleum industries. And as a potential renewable bioenergy that can replace gasoline, butanol has attracted more and more attention from all over the world. [0003] The production process of butanol mainly includes chemical synthesis method and microbial fermentation method. With the depletion of petroleum resources, it is difficult to produce butanol by propylene oxo synthesis using petroleum as raw material, and due to backward technology and small equipment, the production capacity is insufficient, resulting in a long-term shortage of butanol market in China, which cannot meet the demand of ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20C12P7/16C12R1/145
Inventor 李福利张全刘自勇曹长海田磊刘振
Owner CHINA PETROLEUM & CHEM CORP
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