q5 - raji_321

[ArchivalUser]

A patient presents with retinoblastoma. He has a single tumor in one eye. What test could be used to determine whether this is a heritable or sporadic tumor?
A. DNA footprinting
B. Fluorescence-activated cell sorting (FACS)
C. Northern blotting
D. PCR
E. Western blotting

[ArchivalUser]

no idea...we need indirect genetic analysis i think??

[ArchivalUser]

i think its b.

Rb, tumor supressor gene.
checks the G1 point in cell cycle.
two hit hypothesis.

[ArchivalUser]

dd

[ArchivalUser]

The correct answer is D. Forty percent of retinoblastomas are heritable; the child inherits one mutant allele through the germline and a somatic mutation in the other allele occurs, resulting in the loss of function of that gene. This leads to tumor development, often with multiple tumors in both eyes. Penetrance, however, is not complete, since the second mutation is a chance event. Sixty percent of retinoblastomas are sporadic; both alleles are inactivated by somatic mutation. Because this is a rare event, tumors usually are in only one eye and present at a later age.

To detect a mutation in the Rb gene and differentiate between heritable and sporadic cases of retinoblastoma, PCR is the most appropriate technique of the choices given. Using the appropriate primers, a deletion can be detected as a change in size of the DNA band amplified. In heritable cases, one chromosome will show the altered length from the deletion or the translocation in any cell in the body. In sporadic retinoblastoma, samples isolated from anywhere other than the tumor should have two normal alleles.

DNA footprinting (choice A) is a protocol used to detect specific protein binding to DNA molecules by comparing the fragmentation patterns of DNA bound with the protein of interest vs. unbound DNA following digestion with DNase I. If the bound protein protects against DNase I digestion, the position of the protein on the DNA can be deduced by the loss of the characteristic fragment.

FACS (choice B) uses antibodies coupled to fluorescent markers to determine cell surface molecules on whole cells. It is very useful for determining the stage of development or activation of cells, but does not give you any insight into the DNA.

Northern blotting (choice C), the RNA counterpart to Southern blotting, will determine the size and abundance of mRNA of a specific gene in a given sample of RNA. RNA is not able to be cleaved by restriction enzymes the way the DNA can; however, RNA molecules for different proteins are of different lengths. The sample is separated by agarose gel electrophoresis, transferred to a nitrocellulose membrane, and probed with a specific, labeled probe. This will be exposed to film and the bands will appear, revealing the size and the abundance of specific mRNAs.

Western blotting (choice E) is the protein counterpart of Southern and Northern blotting. Proteins are isolated from the cell type of interest and separated by size on a polyacrylamide gel, then transferred to a nitrocellulose membrane. The filter is then incubated with antibodies that specifically react with the protein of interest, then it is incubated with a labeled antibody (e.g., radioactive) that reacts with the first antibody. After this has been washed off, the membrane is exposed to film and bands characteristic of the protein being studied will appear. This is useful to determine whether a protein is present or absent in a patient or if it is the wrong size, indicating an error in the DNA for this protein.

[ArchivalUser]

the logic answer is the correct one. i dont have to forget that... i keep forgeting that...

the logic answer is the correct one... :O)

ok ok.