Science Buddies: "Ask an Expert"

I have been doing research on trying to get bacteria to produce more of a certian type of enzyme.

My question is, if you know the DNA marker (ex. a resistance to a certian antibodic) for the gene that produces the RNA for an enzyme, could you not just use that resistance to remove that DNA segement from another bacteria culture, and then add the segement to another bacteria (as all the other DNA strains would be "dead")? Would you need to add more DNA that codes for the repressor for this enzyme, if you didn't want continous enzyme secreation? Would this extra RNA "overpower" other RNA strains to the point where the cell would die becuase no other RNA was getting to the ribosomes? OR would the extra DNA not make much of a difference at all?

I learned all of this from the interent, so I could be completly wrong about how this works. If so, I would love some 'for dummies' microbiology resources.

Thank you in advance for your help!

Hi Danni,

Great questions. I am going to try to answer, although I am not entirely sure I understand your question.

A little background: When we get bacteria to produce large amounts of a specific, we often will introduce a new plasmid DNA (encoding a resistance gene and the enzyme expressing gene). When we transform the bacteria, we attempt to get MANY copies of the plasmid into the cell so they will make high amounts of the enzyme.

So to answer your questions:

could you not just use that resistance to remove that DNA segement from another bacteria culture, and then add the segement to another bacteria (as all the other DNA strains would be "dead")? Yes, in molecular cloning we do move the gene segments around to builf what we want.

Would you need to add more DNA that codes for the repressor for this enzyme, if you didn't want continous enzyme secreation? If you don't want continuous production you would want to use an inducuble system (can turn it on/off). A common one is IPTG. Others are tet or lac. These systems are based on either repressors or inducers.

Would this extra RNA "overpower" other RNA strains to the point where the cell would die becuase no other RNA was getting to the ribosomes? Possibly. We know that often the cells stop multiplying during times of high expression because of the resource constraints (not just cell machinary, but building blocks and nutrients, too).

OR would the extra DNA not make much of a difference at all? It's hard to tell - you would have to have lots of copies in there to actually kill the cell. However, expressing a toxic protein could kill the cell pretty easily.

I hope that helps. Let me know if further questions.
Tonya