Question

You are interested in studying the expression of Neurogenesis-2 (NG-2), a protein that may have a role in neurogenesis in the brain. You know that neurogenesis slows/stops as mice age, so you hypothesize that it will be highly expressed in young mice compared to old mice. (NG-2 is a fake protein used for this test question). Please note that you are being asked to analyze NG-2 at the PROTEIN level. Select appropriate techniques and technologies. (20 pts total).

a) Describe a method that could be used to quantify NG-2 protein expression that will test this hypothesis and what experimental groups and controls would be included (6 pts).

b) Having determined that your hypothesis is correct, you seek to identify where it is expressed in the brain. You suspect that NG-2 will be expressed in the hippocampus, where neurogenesis occurs. Describe one of the methods that could be used to detect NG-2 in order to identify where it is expressed in brain tissues. What are your expected results if your hypothesis is correct and what experimental groups and controls would be included (6 pts).

c) The results of the previous experiment have confirmed that NG-2 is indeed expressed in the hippocampus. A recently published study used X-ray crystallography suggests that NG-2 contains a putative DNA-binding site. Based on this new information and the results of your previous experiments you propose that it may be a transcription factor that binds to the promoter regions of genes involved in neurogenesis. Describe one approach that could be used to determine if a protein-DNA interaction occurs, what are your expected results (if your hypothesis is correct)? (4 pts)

d) The results of the previous experiment have shown that NG-2 binds to DNA. You hypothesize that one of the sequences it binds to is a promoter region of Sox2, the master regulator of neurogenesis in the brain. What are the next steps to determine if the NG-2 bound DNA sequences are actually from the promoter of Sox2? (4 pts)

Answer 1

First , western blot can be done or Elisa can be performed to determine the protein expression.

And if I am successful and got that young brain has more protein than aged and willing to defect where this protein is synthesized , we can perform immunohistochemistry the groups will be control young aged .

Then to determine that whether it is bound to DNA or not. We can perform chromatin immune precipitation which detect if any protein is bound to a DNA regions.

If can perform many experiment like we can see the expression of mRNA of Sox 2 or protein level of Sox 2 by Western blot in presence of ng2 and in absence of Ng 2.