Tom is a molecular biology graduate student who needs to isolate
a gene from the pathogenic bacteria Salmonella typhi, which he is
studying. He wiselydecides to use PCR to amplify the gene before
trying to purify it. The gene has the general structure shown
below:
~1000 Base Pairs ! ! 5’-ATGGCTTACGTG———–CCATAGGGCTAA-3’ !
! 3’-TACCGAATGCAC———–GGTATCCCGATT-5’
The gene is about 1000 nucleotide base pairs long, but only the
sequences at the ends of the double strand are shown. Use the
information from the video, what you learned about base pairing in
class, and the fact that PCR always proceeds along the parent DNA
strand the 3’ to 5’ direction. Design two, 6-nucleotide long, PCR
primers for Tom. When you have these primers designed, proceed to
the “Exercises and videos” link. You will be prompted for important
information in drop-down menus. Answer each question and click
continue to see a first-person performance of a PCR reaction.
Question #1 (These will be multiple choice and are automatically
graded) a) What different components will do you want to add to
your PCR?
b) Which primers will you use?
c) How many PCR cycles should you set the thermocycler for to
obtain 128 TOTAL strands of DNA at the end of the reaction (this
includes the template strands that you started with)?
d) To analyze the PCR you need to make a DNA gel. What do you
want to add to make the gel?
