Antibacterial composition for combating carbapenem-resistant gram-negative bacteria comprising ADK protein as active ingredient
一种抗菌组合物、抗碳青霉烯的技术,应用在含有效成分的医用配制品、抗细菌药、食品成分的功能等方向,能够解决用途限制等问题,达到优异抗菌活性的效果
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[0062] Example 1. Cloning of recombinant ADK (Rv0733)
[0063] [44] used the genomic DNA of Mycobacterium tuberculosis H37Rv (ATCC 27294) as a template (primer: 5'- CATATG AGAGTTTTGTTGCTGGGACCG-3' (SEQ ID NO: 3) and 5'- AAGCT TCTTTCCCAGAGCCCGCAACGC-3' (SEQ ID NO: 4)) amplifies the ADK (Rv0733) region by PCR. The isolated PCR product was digested with NdeI and HindIII enzymes and inserted into the expression vector pET22b. Escherichia coli BL21 transformed with the ADK gene-inserted vector pET22b was cultured in LB medium (containing 100 µg / ml ampicillin) at 37°C for 12 hours. Subsequently, 1 mM isopropyl-D-thiogalactopyranoside (IPTG) was added thereto, followed by culturing for 6 hours, and the cells were lysed with a lysis buffer (containing 1 M DTT, lysozyme, and PMSF). Recombinant proteins were purified using nickel-nitrilotriacetic acid agarose (Ni-NTA, Invitrogen, Carlsbad, CA, USA) according to the manufacturer's protocol. Finally, the purified recombinant ADK pro...
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