8 thoughts on “C91 – Pierce

    1. Hi Astrid, we don’t know specifically what caused our primers not to anneal correctly, but it is possible that the annealing temperature was too high or too low. Or it could be that the primers were stuck on the side of the micropipette and were not immersed in the solution. For our experiment, there was annealing of cDNA of Hypzif but it was not amplified correctly because it wasn’t the correct size. The primer did not anneal to the gDNA as expected. We can tell that our primers did not anneal correctly because the cDNA lane was the wrong size and the cDNA band was missing.

  1. So just to clarify, is it thought that Hypzif gene down regulation is a way to alter the cell that DNA damage has occurred, or is it meant to actually be involved in the repair of the damaged DNA? You did a nice job, by the way!

    1. Thank you Feriel! Prior to experimentation, we hypothesized that the gene Hypzif would be involved in DNA damage repair pathways. If this hypothesis was valid/true, we would see an increase of Hypzif expression after DNA damage was induced (using hydroxyurea) and this would be seen as an increase in band intensity/brightness. However, we did not see an increase in gene expression following induction of DNA damage, therefore, the gene isn’t upregulated in response to DNA damage. Hence, it is likely not involved in repairing DNA.

    1. Hi Luke, the annealing temperature is around 45–60 °C. This is important because it promotes the primer binding to the template.

  2. How did the difference in band length effect the experiment? How do you induce DNA damage

    1. Hi Morgane, during gel electrophoresis, molecules are separated based on size (number of base pairs). cDNA is smaller than gDNA. DNA damage was induced by using the reagent Hydroxyurea, which impairs the elongation and initiation phases of replication.

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