Klebsiella for strengthening expression of citT gene and application of klebsiella in production of 1,3-propylene glycol
A Klebsiella, propylene glycol technology, applied in the direction of microorganism-based methods, bacteria, microorganisms, etc., can solve the problem of no research report on the citT gene of Klebsiella
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Embodiment 1
[0052] The present embodiment provides a construction method for enhanced expression of citT genetically engineered Klebsiella pneumoniae, which comprises the following steps:
[0053] Step 1, using the DNA of the genome sequence of Klebsiella ATCC BAA-830 (the preservation number of the NCBI database is NZ_CP010523.2) as a template, design primers, and the primers are designed as follows:
[0054] Kv-citT-F: GGCCGAATTCATGAAAGAGAAAAAGACAAC (as shown in SEQ ID NO: 1)
[0055] Kv-citT-R: GAGCGGATCCTCAGAACATCATGGACATCC (as shown in SEQ ID NO: 2),
[0056] The citT gene was completely cloned by PCR to obtain the citT gene.
[0057] Step 2: Use EcoRI and BamHI to digest the citT gene and the expression vector pDK6, connect the citT gene to the EcoRI and BamHI restriction sites of the expression vector pDK6, and construct the citT enhanced expression plasmid pDK6-citT.
[0058] Step 3: Prepare electroporation-competent Klebsiella pneumoniae ATCC 25955, transform the citT-enhanced ...
Embodiment 2
[0066] The present embodiment provides a construction method for enhanced expression of citT genetically engineered Klebsiella pneumoniae, which comprises the following steps:
[0067] Step 1, using the DNA of the genome sequence of Klebsiella pneumoniae ATCC 25955 (the preservation number of the NCBI database is AQQH00000000) as a template, design primers, and the primers are designed as follows:
[0068] Kp-citT-F: GAGCGGATCCATGAAAGAGAAAAAGACAAC (as shown in SEQ ID NO: 3)
[0069] Kp-citT-R: GGCCAAGCTTTCAGAACATCATGGACATCC (as shown in SEQ ID NO: 4),
[0070] The citT gene was completely cloned by PCR to obtain the citT gene. Through detection, the nucleotide sequence of the citT gene of Klebsiella pneumoniae ATCC25955 and the citT gene of Klebsiella mutans ATCC BAA-830 have 93% identity, and the amino acid sequence has 97% identity.
[0071] Step 2: Digest the citT gene and the expression vector pDK7 with BamHI and HindIII, connect the citT gene between the BamHI and HindI...
Embodiment 3
[0080] The present embodiment provides a construction method for enhanced expression of citT genetically engineered Klebsiella oxytoca, which comprises the following steps:
[0081] Step 1, using the DNA of the genome sequence of Klebsiella pneumoniae ATCC 25955 (the preservation number of the NCBI database is AQQH00000000) as a template, design primers, and the primers are designed as follows:
[0082] Kp-citT-F: GAGCGGATCCATGAAAGAGAAAAAGACAAC (as shown in SEQ ID NO: 3)
[0083] Kp-citT-R: GGCCAAGCTTTCAGAACATCATGGACATCC (as shown in SEQ ID NO: 4),
[0084] The citT gene was completely cloned by PCR to obtain the citT gene. Through detection, the nucleotide sequence of the citT gene of Klebsiella pneumoniae ATCC25955 and the citT gene of Klebsiella mutans ATCC BAA-830 have 93% identity, and the amino acid sequence has 97% identity.
[0085] Step 2: Digest the citT gene and the expression vector pDK7 with BamHI and HindIII, connect the citT gene between the BamHI and HindIII ...
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