Difference between revisions of "Part:BBa K3809012"

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Characterization of BBa_K3809012<o:p></o:p>

For the characterization of the protein BBa_K380912 we transformed the plasmid in E. coli BL21. We then examined the growth of different colonies and performed a Colony PCR to determine if they had the insert or not. The result can be seen in Figure 1. And 2. and illustrates that only 6 out of 30 colonies we chose had the insert. <o:p></o:p>

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Figure 1. Gel electrophoresis of the colony PCR we performed for the transformed bacteria BL21 from 1 to 19. We selected only those colonies (marked in blue, M8, M13 and M18) that showed the insert at the size we expected (1948 bp).<o:p></o:p>

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Figure 2. Gel electrophoresis of the colony PCR we performed for the transformed bacteria BL21 from 20 to 30. We selected only those colonies (marked in blue, M21, M23 and M26) that showed the insert at the size we expected (1948 bp).<o:p></o:p>

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After that, we propagated the selected colonies on liquid LB+CAM medium and took samples of all the cultures at 1.5, 3, 4.5 and 48 hours after their inoculation. We ran an SDS-PAGE electrophoresis expecting to see a protein of around 69.5 kDa (corresponding to the size of the protein receptor ESR1) but discovered that only the colonies M18 and M23 had produced a visible protein at the expected site. The best result corresponded to M23, which we describe below. <o:p></o:p>

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Figure 3. SDS-PAGE electrophoresis showing the apparition of a protein of around 70 kDa which corresponds to ESR1.<o:p></o:p>

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