Observed colony growth in LB+ampicillin+kanamycin plate that the bacterial cells
ID: 224509 • Letter: O
Question
Observed colony growth in LB+ampicillin+kanamycin plate that the bacterial cells are resistant to both ampicillin and kanamycin. Is there another way that the cell could have been transformed and have resistance to both ampicillin and kanamycin and yet not contain your recombinant plasmid? Observed colony growth in LB+ampicillin+kanamycin plate that the bacterial cells are resistant to both ampicillin and kanamycin. Is there another way that the cell could have been transformed and have resistance to both ampicillin and kanamycin and yet not contain your recombinant plasmid?Explanation / Answer
Answer:
Yes, the cell could have been transformed and have resistance to both ampicillin and kanamycin and yet not contain your recombinant plasmid.
If your plasmid vector contains the kanamycin and ampicillin resistance genes and do not have your insert DNA (gene of interest), colonies will still appear on the LB+ampicillin+kanamycin plate.
In that case, plasmids need to be isolated from the corresponding colonies and confirm for the presence of the insert through restriction digestion of the plasmid with restriction enzymes used for cloning the gene of interest.
This might happen if the plasmid vector gets religated after restriction digestion during the cloning step or the vector did not get digested at all. Use of antarctic phosphatase may solve the problem associated with the religation of the vector during the cloning step.
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