Dna And A Psbc Lacking Species Through Double Homologous Recombination

2204 Words May 6th, 2016 9 Pages
The intent of this experiment was to successfully transform synechocystis to a psbC lacking species through double homologous recombination. Subsequently we intended to retransform the mutated colonies back to the psbC carrying species after kanamycin selection. The initial knockout of the psbC gene ensured that photosystem II wouldn’t work and the colonies would rely on glucose as a source of sustainable nutrition. The sequential addition of kanamycin killed off the species that lacked the KanR gene that was replacing the psbC gene necessary for photosynthesis. Once we were content with the selection we were able to retransform the species back to the wild type carrying the psbC gene and reinstall the ability to use light as a source of energy. We employed a variety of genetic recombination and analytics to successfully achieve our goals.
In Figure 1, the Synechocystis was advantageous in this experiment since it can naturally transform and instantly take up foreign DNA. Synechocystis does this by double homologous recombination and in this case, the process was manipulated by making the plasmid pKCP43 and disrupting the psbC gene. Since the two ends of the plasmid and the gene are extremely similar, the Synechocystis bacteria can easily uptake the plasmid leaving behind the psbC gene during crossover in homologous recombination. As the bacteria replicate the kanamycin resistant gene present in the pkCP43 plasmid, continue to reproduce while the bacteria that did not…

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