You are asked to investigate the level of expression of gene X in human kidney a
ID: 175966 • Letter: Y
Question
You are asked to investigate the level of expression of gene X in human kidney and other organs. Which of the following experiments would you first select?
a) Do a qRT-PCR to compare levels of expression of the gene X in kideny and other organs
b) Do an in situ hybridization to analyze which chromosone the gene X is located in
c) Do a DNA fingerprinting to see how different the gene X is in different organisms.
d) Do a Northern blot probed with an antibody probe to anayze levels of X mRNA in different organisms.
e) Do a Southern blot hybridization to analyze the GC content of DNAs in different organs.
Explanation / Answer
The answer is option d)
The amount of mRNA transcript for a single gene ( gene X) is directly proportional to how much transcription of that gene has occurred. Analysing that quantity will therefore indicate how strongly a gene is transcribed, or expressed. The differences in the quantity of mRNA produced by different groups of cells or at different times can be visualized by the technique known as Northern blot. In this method , the mRNA is first isolated from a biological sample by treating the cells with protease. Protease is an enzyme which breaks down cell membranes and releases the genetic material in the cells. Then the mRNA is separated from the DNA, proteins, lipids, and other cellular contents. The different fragments of mRNA are separated from one another by Electrophoresis, and are then transferred to a filter or other solid support using a technique known as Blotting. The mRNA transcripts produced by a particular gene is identified by incubating the sample with a short piece of single-stranded RNA or DNA (also known as a Probe) which is labeled with a radioactive molecule. The probe is designed to be complementary to the mRNA from the gene of interest ( gene X) , and hence it will bind with it.Then the filter is placed against X-ray film, the radioactivity in the probe will expose the film by making marks on it. The resulting marks are knwon as bands and they tell how much mRNA was in the sample, which is a direct indicator of how strongly the gene of interest is expressed .
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