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Queens UniversityBCHM 218PBA3

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BCHM 218 – Fall 2019 – Problem-Based Assignment #3 1. You are a scientist studying the members of a family with a rare lung disease. The disease has been linked to a genetic mutation involving t ... he insertion of a 180 bp segment within exon 3 of the BCHM2 gene. The mutation is illustrated in the schematic diagram below. M u t a n t B C H M 2 N o r m a l o r “ W i ld t y p e ” B C H M 2 F w d 1 8 0 b p P r i m e r 6 0 0 b p , G + C = 3 5 % G + C = 5 6 % R e v P r i m e r R e v P r i m e r F w d P r i m e r You use a set of forward and reverse primers (shown above) to perform PCR reactions on separate DNA samples from three members of the affected family. Family member 1 does not carry the mutation. Family member 2 has one normal copy of the BCHM2 gene and one mutated copy. Family member 3 has two copies of the gene bearing the insertion mutation. You take your three reaction products and run half of the volume in three separate wells of an agarose electrophoresis gel. a) Please draw a picture of the gel that you would expect to see (make sure to specify the sample in each lane and the size of all products in bp) (3 marks). Family Member 1 Family Member 2 Family Member 3 600 bp 420 bp More DNA in a band gives more intense staining of that band (black = more DNA). b) Upon completing your electrophoresis, you remember that you forgot to add ethidium bromide to your gel. What does ethidium bromide do in this protocol (2 marks)? What would be the consequence of this error (1 mark)? Ethidium bromide is a planar molecule that can act as an intercalating agent by inserting itself in between nucleic acid strands in a nonspecific manner. It works to stain the samples in the gel since it fluoresces when exposed to UV light. Without ethidium bromide the fragments in the gel will not be seen [Show More]

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