8. Do you think the transformation was successful? Compare the results in Figure

Question

8. Do you think the transformation was successful? Compare the results in Figures 3-6 with the results in Figures 7-10. 9. Another researcher performed the same exx 100 and obtained the results show in the table bel pGRN Plate Light number of added? colonies + NA visible +NA UV + AMP visible + AMP UV 10,000 10,000 50 50 10,000 10,000 50 50 NA visible NA UV -AMP visible AMP UV Explain these results, focusing on the shaded rows. Do you think your transformation was successful? Explain your answer 10. As a general principle, in order to validate results in scientific experiments, scientists use controls. A control is a group that is designed to demonstrate that the results/changes observed in an experiment were because of the procedure you performed. It sets a baseline value with which the scientist can compare with experimental sample and interpret results, as was demonstrated in the previous question. In this question, we are asking you to describe the control experiments, using the same transformation protocol, with pGRN plasmid. You want to test whether the E. coli cells were viable (alive). How would you test this? Consider the following (a) do you add the plasmid to the cells? (b) what plate do you use NA or NA-AMP and (c) do you use visible light or UV light?

Explanation / Answer

9. The result in the two shaded rows can be explained as follows:

a) pGRN+AMP+UV/Visible ---> 50 (no. of colonies)

Explaination: the green color is observed hence the bacteria is observed only when the pGRN is integrated in to the bacterial chromosome and is expressed. The plasmid is not responsible for the fluoroscence but the product of GFP is responsible for the color. In this experiment we can conclude that the plasmid was taken up successfully (maybe) but not integrated and expressed and that is why the number of colonies observed is less.

b) AMP+UV/Visible ---> 50 (no. of colonies)

In this experiment there was not plasmid and plates were added with the ampicillin antibiotic. Since there was not plasmid added, there was no ampr gene and no resistance developed in the colonies. herefore the bacterial growth was inhibited by ampicillin reducing the number of colonies

10. To check whether the E.coli cells were viable, you do not use plasmid, you use just NA, withou selection i.e. without AMP and you use visible light.

E.coli+NA+Visible light

UV damages the bacteria.

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