The purpose of this project is to examine if the extent of resistance in E.coli MG1655 is depending on which kind of plasmid the resistance gene is placed on, a high copy number plasmid or a low copy number plasmid. We wish to see if the copy number makes a difference, as it so far never has been examined whether the amount of antibiotics needed to prevent the bacteria from growing is determined by the copy number of the plasmids or not. Furthermore, we wish to see if the specific target of the different antibiotics has an effect on the extent of resistance. We have chosen to work with three different antibiotics; tetracycline, ampicillin and chloramphenicol. The reason for selecting these antibiotics is that they each have different targets, and therefore uses different mechanisms to inhibit the bacteria's growth. This means that the bacteria have to use different methods to inactivate each of the antibiotics. Our method is to insert resistance genes towards the three antibiotics to both a high copy plasmid and a low copy plasmid. These plasmids are then transformed into MG1655. Finally we use MIC measurements to compare the extent of resistance. To make an MIC measurement means to measure the lowest amount of antibiotics needed to inhibit the bacteria's growth. We started by assuming that the resistance of tetracycline would depend of the copy number of the used plasmid and the resistance of ampicillin and chloramphenicol would not. We based this theory on our knowledge about their mechanisms, respectively. Unfortunately we only succeeded in getting clear results from the experiment with ampicillin. These results showed that the extent of resistance in the case of ampicillin and its corresponding resistance gene ampR is depending on, whether ampR is placed on a high copy number plasmid or a low copy number plasmid. In the case with chloramphenicol and cat, our results indicate that the resistance is not depending on the copy number of the plasmid. However, one of our plasmids did not have a completely correct insert, so the experiment will have to be repeated before anything can be concluded. We did not get any results from the experiment with tetracycline.
|Uddannelser||Basis - Naturvidenskabelig Bacheloruddannelse, (Bachelor uddannelse) Basis|
|Udgivelsesdato||24 jun. 2010|