Method for simultaneously removing bacillus anthracis virulence megaplasmids pXO1 and pXO2
A Bacillus anthracis and plasmid technology, applied in the field of genetic engineering, can solve the problems of unclear replication and separation of Bacillus anthracis large plasmids, slow research progress, etc.
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[0045] Example 1. Acquisition and Identification of Recombinant Bacteria Eliminating pXO1
[0046] 1. Repel plasmid construction
[0047] 1.1 Temperature-sensitive plasmid linearization
[0048] The temperature-sensitive shuttle plasmid pKSV7 (6.9Kb) was linearized with restriction enzymes HindIII and EcoR I (HindIII and EcoR I are the multiple cloning sites of pKSV7), and then separated by 0.6% agarose gel electrophoresis, The target band was excised from the gel, and the linearized vector pKSV7 fragment was recovered with a DNA gel extraction kit, and the operation steps were performed according to the instructions.
[0049] Backbone plasmid: pKSV7 plasmid (neither transposon nor insert sequence on it): Smith K, Youngman P.Use of a new integrational vector to investigate compartment-specific expression of the Bacillus subtilis spoIIM gene[J].Biochimie, 1992 , 74:705-711., publicly available from the Institute of Bioengineering, Academy of Military Medical Sciences, Chinese...
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