How would u generally set this up? Hypothesis: you believe that a gentic disorde

Question

How would u generally set this up?
Hypothesis: you believe that a gentic disorder (Duchene's Muscular Dystrophy) is caused by a deletion mutation in the dystrophin gene.

Step 1: designing the overall study: how is the study designed specifically the independent and dependent variables and how will up the structure of study? Hypothesis too.

Step 2: what procedure we learned in class is used to extract the DNA

Step 3: make copies of the DNA (no questions for this part)

Step 4: Now that u have the copies of the Dystrophin gene. How do u test the hypothesis? Specifically, what technique will you use?

Step 5: what would be your expected results if the hypotheisis is supported?

Step 6: Lets say in the last step, you did not see any results and decided to start parameterizing your study. How might you break down parametrization process? Where would you start and what would you do?

Explanation / Answer

Deletion mutations causes frameshift in the transation and the final polypeptide will be different from the initial one.

1. We could purify DNA from a patient sample, amplify it and express it in some organism like bacteria with a tag, purify this protein and do an assay to see whether it is dystrophin. We can simply run both crude dystrohin and the protein obtained on a gel. Proteins formed from deletion mutations are usually smaller in length.

To be sure, we can sequence this amplified gene by Chip-seq method and compare it with the original gene.

2. We can use Alkaline Lysis method with RNAase treatment for DNA extraction.

3. Using PCR amplify the gene.

4. We can test the hypothesis by the methods on Step 1. We could also do a monoclonal antibody based assay to see if it is dystrophin.

5. When running the gel the band from the suspected protein shall be of lower molecular weight. In Sequence we can pinpoint the deletion if any.

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