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Team:Elan Vital South Korea/p results and conclusion
From 2014hs.igem.org
Results and Conclusion
We first grew the MRSA in liquid LB broth, and then placed the MRSA in solid LB plate with a variety of antibiotics, and observed its growth. This is the resulting growth of the MRSA.
Then we wanted to transform the E. coli with the DNA from the MRSA and test its resistance, but first, we had to test the original E. coli in the drugs, as without that, we wouldn’t know if the drug resistance was from the MRSA DNA, or from the E. coli. We placed the E. coli on a LB plate with a variety of antibiotics and observed its growth. This will be the control of this section of the experiment. This is the resulting growth of the E. coli. The results show that the E. coli has no resistance to gentamycin, kanamycin, ampicillin, and tetracycline.
Growth of E. coli 24 hours it was placed in different antibiotics (top left: kanamycin, top right: gentamycin, bottom left: ampicillin, bottom right: tetracycline.) with 1/1000 concentrations.
Then to test the drug resistance of the transformed E. coli, we placed the E. coli in a LB plate and observed its growth. This is the resulting growth of the transformed E. coli.
7 A, G, K, T: DNA of MRSA from patient 7 grown in ampicillin, gentamycin, kanamycin, and tetracycline, respectively.
8 A, G, K: DNA of MRSA from patient 8 grown in ampicillin, gentamycin, and kanamycin, respectively.
7 GA, KA, TA: DNA of E. coli grown in ampicillin, which were transformed with the DNA of MRSA from patient 7 grown in gentamycin, kanamycin, and tetracycline, respectively.
8 GK, KK: DNA of E. coli grown in kanamycin, which were transformed with the DNA of MRSA from patient 8 grown in gentamycin and kanamycin, respectively.
8 GG: DNA of E. coli grown in gentamycin, which were transformed with the DNA of MRSA from patient 8 grown in gentamycin.
The results show that wells 2-8 has about 300 base pairs, which means that most likely the type V primers were active in PCR, and the section of DNA between V-F and V-R with about 300 base pairs were multiplied in PCR of 7A, 7G, 7K, 7T, 8A, 8G, and 8K. This most likely means that 7A, 7G, 7K, 7T, 8A, 8G, and 8K all included the section of 300 base pairs that is between V-F and V-R. So, this 300 base pair long section of DNA has a possibility of coding for multidrug resistance. The results also show that wells 9-11 has about 1000 and 1600 base pairs, which means that most likely the primers ccrAB-α3 and ccrAB-α4 were active in PCR, and the section of DNA between ccrAB-α3 and ccrAB-α4 with about 1000 base pairs and the section of DNA between ccrAB-α3 and ccrAB-α4 with about 1600 base pairs were multiplied in PCR of 7K, 8G, and 8K. This most likely means that 7K, 8G, and 8K all included the sections of 1000 and 1600 base pairs that is between ccrAB-α3 and ccrAB-α4. So, this 1000 base pair long section of DNA and the 1600 base pair long section of DNA has a possibility of coding for multidrug resistance.
The results show that wells 12-17 has about 150 base pairs, which means that most likely the primers MecA147 or the primers mecI were active in PCR, and the section of DNA between MecA147-F and MecA147-R or the section of DNA between mecI-F and mecI-R with about 150 base pairs were multiplied in PCR of 7GA, 7KA, 7TA, 8GK, 8GG, and 8KK. This most likely means that 7GA, 7KA, 7TA, 8GK, 8GG, and 8KK all included the section of 150 base pairs that is between MecA147-F and MecA147-R or the section of 150 base pairs between mecI-F and mecI-R. So, this 150 base pair long section of DNA has a possibility of coding for multidrug resistance.
The investigation showed that MRSA showed resistance to some of the antibiotics, and some of the transformed E. coli showed antibiotic resistance. That most likely means that the DNA that codes for antibiotic resistance was successfully transported into the E. coli that survived the antibiotics. By running PCR, we were able to amplify some of the DNA in the drug resistant MRSA and transformed E. coli. Since the section of DNA that codes for the multidrug resistance is most likely shared in all the bacterium that survived the antibiotics, by analyzing the DNA through gel electrophoresis, we were able to get an idea about which section of DNA was shared. If possible, the next step forward would have been to analyze the sequence of the section of DNA, but we were not able to procede that far.
