Question

A biologist is interested in purifying a large amount of a protein she is studying. The protein is a fascinating new type of enzyme from a rare bacterium that grows only in deep-sea hydrothermal vents. She clones the gene and expresses it in E. coli, hoping to use the E. coli as a factory for that protein. The plasmid she uses is similar to the pGLO plasmid—it includes the bla selectable marker (encoding ampicillin resistance) and it uses the arabinose-dependent inducer of GFP expression (araC) to regulate the expression of the gene. Her initial transformation efficiency is high, and she continues growing transformants on LB/amp/ara plates. After several weeks, however, she notices that the transformants are less viable—they are growing more slowly, and then not at all. Through a quick test, she sees the bacteria still appear to have the plasmid. What might be going wrong? Can you suggest a solution that might help?

          A biologist is interested in purifying a large amount of a protein she is studying. The protein is a fascinating new type of enzyme from a rare bacterium that grows only in deep-sea hydrothermal vents. She clones the gene and expresses it in E. coli, hoping to use the E. coli as a factory for that protein. The plasmid she uses is similar to the pGLO plasmid—it includes the bla selectable marker (encoding ampicillin resistance) and it uses the arabinose-dependent inducer of GFP expression (araC) to regulate the expression of the gene.

Her initial transformation efficiency is high, and she continues growing transformants on LB/amp/ara plates. After several weeks, however, she notices that the transformants are less viable—they are growing more slowly, and then not at all. Through a quick test, she sees the bacteria still appear to have the plasmid.

What might be going wrong? Can you suggest a solution that might help?

Added by Rocio B.

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