Jackofknives
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Spinobulbar muscular atrophy (SMA) is an X-linked recessive motor neuron disease, arising from a mutation in the androgen receptor gene. Shown in the diagram below are results of DNA sequencing studies of the receptor gene from healthy individuals and SMA patients. The normal allele of the gene has 10 adjacent glutamine codons (CAG) in exon 1, whereas the SMA allele has 40 of these CAG repeats in the same region (a mutation referred to as triplet repeat expansion).
Normal gene … 5’UTR …CAG(CAG)8CAG…
SMA allele … 5’URT…CAG(CAG)38CAG…
The most appropriate DNA-based test to detect this mutation would utilize PCR primers:
A) Complementary to the CAG repeat followed by gel electrophoresis of the PCR products
B) Flanking the CAG repeat followed by a dot-blot using a CAG repeat-specific DNA probe.
C) Flanking the CAG repeat followed by gel electrophoresis of the PCR product
D) Complementary to the CAG repeat followed by a dot-blot using a CAG repeat-specific DNA probe.
Please explain your answer
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| new_n_lost
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B) Flanking the CAG repeat followed by a dot-blot using a CAG repeat-specific DNA probe.
PCR and Southern Blot thnking outside the box not sure with the answer and cant explain will have to work over it to explain.
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| Jackofknives
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The answer is A
Flanking is not involved in PCR only denaturation. Correct me if im wrong but flanking means breakage of the DNA into pieces, that is not done here. In PCR with 2 complementary probes you will get the desired piece of DNA as soon as at the 2nd copy.
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| doc179
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no way ,the answer can not be A. the answer is definitely C. Flanking does not mean breaking the DNA. Flanking means adding the complementary sequences to both sides of the CAG repeat for a PCR.
and this qn is in kaplan notes.
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| Jackofknives
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oops my bad
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| sfk
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doc179 u r right, ans is c. the q is in KLN, but could u plz throw some more light on the explanation..
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| yoga
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plz set the record straight
what is the answer
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| drgho
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 C.
The main diagnostic is to detect the different sizes of CAG repeats sequence between two groups.
so
A : can replicate the sequence anywhere in CAG sequence and not sure to see the size differences.
B: dot-blot can't detect different sizes and it is maily for nucleotide polymorphism/mutation.
C: ensure the whole CAG repeat sequences are replicated and then the size detect. so in the diseased state the sequence will migrate less.
D: dot blot cannot detect the size differences and the PCR products from that primers not guaranteed to get different sized products in two states.
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| asha_usmle
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Flanking means addition of short primers complementary to the 3' regions bordering the sequence to be amplified..
PCR primers flanking the mutation, followed by Southern blotting, would be needed to detect the extent of the triplet repeat expansion
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