Recombinant DNA Lab

Process: To produce recombinant DNA, you first need a plasmid that has some antibiotic resistance. Our plasmid was resistant to ampicillin. This plasmid is cut open by a restriction enzyme which also cuts the insulin gene out of another piece of DNA. For us finding an enzyme that did all of this was quite tedious, and unfortunately, the one that worked was the last one to try.This insulin gene that was chopped out and some of its surrounding code are inserted into the plasmid and stuck together by using the enzyme ligase. This plasmid with the insulin gene and antibiotic resistance is then inserted into a bacteria. This bacteria then replicates and passes the antibiotic resistance on to all of its offspring. Ampicillin can then be added to the petri dish to kill off all the bacteria that don't have antibiotic resistance, thus leaving only the bacteria that produce insulin.

1. In my petri dish I would only use antibiotics that I know the plasmid carried resistance for. Otherwise, I might kill off the bacteria that have the insulin gene too. I also won't use antibiotics that all the bacteria in the species are resistant too, because that wouldn't refine the types of bacteria in the dish.

2. Restriction enzymes are enzymes that cut open DNA when they read a certain sequence. They work by cutting in patterns that create "sticky ends". I used HIND III because it cut extremely close to the insulin gene on both sides, as well as cut open my plasmid in one place.

3. If my enzyme cut the plasmid in 2 locations, then once the insulin gene is inserted in, I would be left with a string of DNA rather than a loop. However, I think that if ligase is introduced, the plasmid will reconnect to itself and everything would proceed as normal.

4. This process is important in our daily lives, because bacteria are extremely useful factories for protein. Although right now the only well-known use for bacteria-produced protein is insulin, I am sure in the future, we will be using bacteria as factories for all types of materials.

5. This process could eventually actually be used for bacteria to convert things like plastic and feces into usable fuel or other goods. For example, we would first have to engineer an enzyme for the digestion of plastics. We may then have to convert this into a genetic code. The code could be produced by various DNA "printer" technologies that are being innovated today. This gene will then be spliced by a restriction enzyme that also splices open a plasmid. The process will then be repeated except with an enzyme for the construction of propane or another fuel. Once all the necessary plasmids are inserted into bacteria, we could essentially feed the colony our plastic waste, and fill our cars or propane tanks out of the other side.

The plasmid with insulin gene and ampicillin resistance