drk1980
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A phenotypically normal man is tested for a specific mutation associated with a genetic disease in his family. A polymerase chain reaction (PCR) is run on a small sample of his DNA, using primers to amplify the region around the suspected mutation. The PCR products are tested (Test) by both gel electrophoresis and by using allele-specific oligonucleotide probes on a dot blot. The results of these two tests are shown below. In each case, a sample of DNA from a homozygous normal individual (Control) is shown for comparison.
Dot Blot shows Test has two filled circles and Control only one filled circle.
Gel Electrophoresis shows: Test sample is one line further down(longer) from the one line representing Control.
Which of the following genetic diseases is most likely affecting his family?
A. Cystic fibrosis
B. Duchenne muscular dystrophy
C. Huntington disease
D. Marfan syndrome
E. Sickle cell anemia
Kindly give reasoning for choices.
Edited by prep4usmle on 12/17/05 - 05:12 PM
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| reet
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Couple expecting first child, consulted genetic counselor cuz woman had a brother died of sickle cell anemia. There is also history of ds in MANS family. FETAL CELLS are obtained by amniocentensis. WHICH test would be . (below given). tells fetus having a disease??
A. WESTERN BLOT
B, Hb ELECTROPHORESIS
C.NORTHERN BLOT.
D. PCR with allele specific probes on a DOT-BLOT
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| sturge_weber
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cystic fibrosis
first lets rule out the autosmal dominant, dot blot has filled only one circle in the control showing that it has both the same genes ... but the man who is not manifesting the disease has one mutant and one normal alleele, showing that he is a carrier, this proves that they re talking about an autosomal recessive disease.. so huntingtons , marfans ruled out....
now when we look at the electrophoresis, we see that the test sample is one line further down than the control..
now the rule here is to follow is that the shorter the dna the further down it will go, because in gel electrophoresis we have a negative charge at the top and positive at the botton, and when we give a negative charge , the dna moves to the opposite side , that is the positive which is down, and shorter the dna the more fasrther it will go, hence this implies that the tested sample has a deletion,
out of the remaining choices cystic fibrosis is an autosomal recessive dfisease where the phenylalanine is knocked off leading to an abnormal cl- transporter.
now sickle cell anaemia is also an ar disease but the there is replacement of a nucleotide, with no change in the frame hence it is ruled out.
the dmd also does not have a deletion
hence i think the ans is cystic fibriosis
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| sturge_weber
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and ya i forgot to say about dmd, the point is that it is x linked recessive and hence the males only need to have it, and they only one chr, and hence only one gene, and hence they exhibit, but this guy has two different genes.. as seen in dot blot, hence its also ruled out
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| sturge_weber
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to reet
i will go for pcr with dot blot
western blot, cant be as the fetus has embryonic hb, and not hba which undergoes sickling
do post the answers guys
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| reet
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sturge great job,, but i m still confused ....in this question
Edited by reet on 12/01/05 - 08:21 AM
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| drk1980
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thanks sturge_weber for the detailed explanation!! esp on the gel eletrophoresis...
you narrowed it down pretty well(AR disorder). iam afraid, though, the answer given is slightly different. would you like to try again?
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| sturge_weber
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well whats the answer
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| drk1980
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"sickle cell anemia" !!
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| reet
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huh?? now can u explain difference in both in this ques.
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| sturge_weber
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re u sure on the gel electrophoreis, the diseased was down than the control, or at the same level, if its down it means a smaller gene, which means a deletion
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| drk1980
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i cudnt put up the graphics in the Q...
anyhow...rechecked, yes on gel electrophoresis the Control line shows higher up than the Test line, which has traveled down further.
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| sturge_weber
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well i think there must be a catch in the diagram then, because this guy is normal has two allelles on dot blot, which confirms he is a carrier, and its a recessive disease. now if it were sex linked recessive , the patient a male would have manifested the symptoms, but as he is normal, it gotta be autosomal recessive....
now only two diseases left, cystic fibrosis, and sickle disease, does ur diagram give 63 kb and 60 kb or 1.35kb and 1.15 kb
then we can reach the ans
may i ask where is it from
reet i hope u re not asking me the q u posted, it suggests that the child is fetus, and fetus has embroynic hb and hardly any HbA , and the fetal Hb has a very strong affinity for o2. hence if we do the western blot wont help.
but if we a pcr of the gene which codes for Hba and then do a dot blot , we will reach the answrr... please do post the ans cause thats what i assume
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| reet
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sturge,,
answer fr my ques is right, .. ,
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| drk1980
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iam sorry, i ws under the impression i had already mentioned in the beginning.... this is a Kaplan Simulated Online Test question.
the test comes with explanations of the answers, but i dont hv that with me currently.
Are 'allele-specific oligonucleotide probes on dot blot' used for any disorder other than sickle cell?
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| sturge_weber
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ya they can be used for all the disorders hwere there is a small mutation, eg cystic fibrosis....
see kaplan genetics section, its very well explained there, and buddy do post the ans when u get the explanations, as to where i was wrong, as i still dont get what i am wrong in
thanks
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| drk1980
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picture
Attached Files:
Doc1.doc (24 KB, 32 downloads)
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| drk1980
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hope its easier to understand now...
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| sturge_weber
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well i still stick with cystic fibrosis, becoz the cystic fibrosis, u lose a gene, and hence the the test should be lower than the control, but ib sickle cell anaemia, there is a larger fragment, ie, 1.35kb instead of normal, 1.15 kb in a normal person, so the test should be higher than the the control, as larger the fragment, the less it will move on gel electrophoresis, so i still see the ans, as cystic fibrosis,
i hope someone else clarifies the doubt
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| drk1980
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i agree with ur answer too.
maybe this one Q ws just an error on the part of kaplan 
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| sturge_weber
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| Doc2378
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CF it is...for sure...
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| Geroo
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hi guys,
if we say that it's CF,and the man is a carrier,isn't he supposed to have 2 bands coz he is a carrier??
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| drk1980
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Geroo, can u expand on tht?
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| Geroo
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if the patient is a carrier of CF.the gel electrophoresis supposed to show 2 bands one normal long band and one abnormal short band.right?
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