This plasmid is digested with the enzymes listed below. Indicate how many fragments will be generated in each case, and give the sizes of the fragments. PstI XhoI Combination of PstI + XhoI + EcoRI (triple digest) ded

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Chapter1: The Human Body: An Orientation
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5.
Consider the following plasmid (size 8000 bp), with restriction sites at the positions indicated:
PstI
XhoI
7000
8 kb
2000
XhoI
Probe
4200
ECORI
This plasmid is digested with the enzymes listed below. Indicate how many fragments will be
a)
generated in each case, and give the sizes of the fragments.
PstI
XhoI
Combination of PstI + XhoI + EcoRI (triple digest)
b)
Draw the banding pattern you would expect to observe if each of these digestions is loaded
into a separate well of an agarose gel, and the fragments separated by electrophoresis. In the
first well you load a DNA marker (M) containing fragments with sizes of 1000 bp, 2000 bp,
4000 bp and 8000 bp.
This gel is transferred to a membrane in a Southern blot experiment, and hybridised to a
c)
radioactively labelled 200 bp probe, which anneals to the plasmid DNA at the position indicated
on the diagram above. Draw the autoradiographic profile you would expect to observe for the
membrane.
Transcribed Image Text:5. Consider the following plasmid (size 8000 bp), with restriction sites at the positions indicated: PstI XhoI 7000 8 kb 2000 XhoI Probe 4200 ECORI This plasmid is digested with the enzymes listed below. Indicate how many fragments will be a) generated in each case, and give the sizes of the fragments. PstI XhoI Combination of PstI + XhoI + EcoRI (triple digest) b) Draw the banding pattern you would expect to observe if each of these digestions is loaded into a separate well of an agarose gel, and the fragments separated by electrophoresis. In the first well you load a DNA marker (M) containing fragments with sizes of 1000 bp, 2000 bp, 4000 bp and 8000 bp. This gel is transferred to a membrane in a Southern blot experiment, and hybridised to a c) radioactively labelled 200 bp probe, which anneals to the plasmid DNA at the position indicated on the diagram above. Draw the autoradiographic profile you would expect to observe for the membrane.
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