The mutation we are looking for is present in all of the body's cells, including the blood. Which DNA are we going to use to check for this
... [Show More] mutation?
genomic DNA
Which part of the blood contains the highest concentration of genomic DNA?
white blood cells
How should we perform a linkage analysis?
genotype the microsatellite markers
Who would be the ideal healthy control in linkage analysis?
a close relative who does not have breast cancer
What is a microsatellite?
repeating sequences of two to six base pairs of DNA
We want to analyze four different microsatellite using four different primer pairs in two samples and prepare a master mix for each set. What should be included in the master mix?
primers, nucleotides, and polymerase
Why is it important to change the pipette tip?
avoid cross-contamination
What is the purpose of PCR?
to create many copies of DNA template
What happens next in the PCR process under this condition?
the DNA separates into two strands
What causes this separation?
the high temperature 95
What is this step in a PCR reaction called?
annealing
The area where the primers bind marks which part of the PCR product?
beginning
The PCR products get a certain length due to which fact?
the placements of the primers
How does the DNA polymerase extend the primers into the primers into a new DNA strand?
adding nucleotides to the 3' end of the primers
Which is NOT the purpose of the loading buffer?
To fasten the electrophoresis experiment
How many copies of DNA are required to see bands on the electrophoresis gel?
millions of copies
Which phrase describes the charge of the DNA?
negatively charged
DNA is negatively charged. To which location in the gel will it migrate?
to the positive pole
The patient has two bands for the D17S855 marker. What does this mean for the patient?
they are heterozygous for this gene
For the microsatellite marker D13S1699 there is only one band corresponding to one PCR product. Which word describes both the patient and healthy control?
homozygous
Both the markers D13S1699 and D13S1701 are located proximal to the BRCA2 gene. Are they linked to the disease?
no
What about the markers D17S855 and D17S1323 that are intragenic to the BRCA1 gene; are they linked to the hereditary breast cancer in this family?
they could be linked, but I need more information
In a pedigree, what is the symbol for a woman that is affected with the disease?
filled circle
What does a diagonal line over the symbol denote?
the subject is deceased
What does an arrow pointing at a symbol denote?
this indicates the proband
Using the pedigree information, can you determine which marker is linked to the hereditary breast cancer in this family?
yes, it is D17S1323
What can we do next to identify the mutation and the consequences?
perform a protein truncation test
What type of mutation can we detect using the protein truncation test?
nonsense mutation
What is the purpose of adding SDS (sodium dodecyl sulfate) when running a polyacrylamide gel electrophoresis?
to denature the protein [Show Less]