Recombinant microorganism for diol production
A technology for recombinant microorganisms and diols, applied in microorganisms, microorganism-based methods, recombinant DNA technology, etc., can solve the problems of high synthesis costs, increased separation and recovery costs, environmental pollution, etc., and achieve high generation capacity, generation and recovery. Simple, low-generating effects
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experiment example 1
[0053] Preparation of Recombinant Microorganisms
[0054] Klebsiella pneumoniae GSC123△ldhA (Kp△ldhA) strain
[0055] A lactate dehydrogenase (ldhA)-deleted Klebsiella pneumoniae GSC123ΔldhA (KpΔldhA) strain was prepared in the following manner. First, in order to clone lactate dehydrogenase of Klebsiella pneumoniae, ldhA (SEQ ID NO: 1) as a target gene was amplified by polymerase chain reaction (PCR) using primers of SEQ ID NO: 3 and 4 Homology region 1 (SEQ ID NO: 2). Also, homology region 2 (SEQ ID NO: 5) was amplified by polymerase chain reaction using primers of SEQ ID NO: 6 and 7. Then, the homologous region 1 and the homologous region 2 are used as templates and amplified by polymerase chain reaction at the same time, thereby completing the deoxyribonucleic acid (DNA) fragment (SEQ ID NO: 8) (Table 1).
[0056] In order to increase the probability of recombination of the target gene, the completed deoxyribonucleic acid fragment can include antibiotic resistance gen...
experiment example 2
[0077] Formation of diol
[0078] The recombinant strain prepared in Experimental Example 1 above was cultivated to produce diol. In this case, wild-type Klebsiella pneumoniae GSC123 (Kp wt) was used as a comparative example.
[0079] Each recombinant strain was inoculated into 300 ml of complex medium, and after culturing at a temperature of 37° C. for 16 hours, the above-mentioned culture solution was inoculated into 3 L of complex medium for fermentation. At this time, the fermentation conditions were as follows: micro-aerobic condition (aerobic rate: 0.5vvm, agitation rate: 300rpm), 20g / L glycerol at the beginning, and 30g / L glycerol at the time of feeding , the pH was 6.0, and the culture temperature was 37°C. Ammonia (NH4OH) is used to adjust the pH during fermentation. With respect to the above-mentioned recombinant Klebsiella bacteria, a sample in fermentation was used, and the growth rate was detected by detecting the OD600 (optical density) of the sample used, an...
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