Mr. Poruban
Lab Activity 4.4: Restriction Enzyme
Genetics/Biotechnology
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There is no specific pattern associated with the upper sequence of bases
A pairs with T and C pairs with G
CTTAAG
Restriction enzyme cuts between the G and A in the palindromic sequence: GAATTC
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4
10
No, it is larger
Fragment 1 is a 4-base-pare fragment
Fragment 2 is a 10-base-pare fragment
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5
Random sized fragments will be produced
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The DNA is colorless
NO, the DNA in solution is not visible
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The DNA in the uncut lambda DNA tube, which gets no enzyme, should remain intact as a single
band
No restriction enzymes were added to the tube, thus no digestion will occur, and no fragments
will be produced. Tube L is the control tube
P has PstI; there should be an enzymatic digest in this tube which will produce fragments. The L
tube has DNA and no enzyme..so no restriction fragments.
With no restriction enzyme present…no digestion (cutting of DNA) will occur.
Some restriction enzyme was added accidentally? Maybe not changing pipet tips?
No. the DNA still appears colorless.
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3
Fragment 3 is the largest
Fragment 2 is the smallest
6
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Answers will vary
Answers will vary
1 |Chapter 4: Lab Activity 4.4: Restriction Digest
Mr. Poruban
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The enzyme would cut only at site B, producing 2 DNA fragments
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The large fragments would be towards the top of the gel because it is more difficult for the larger
pieces to be strained through the gel
There would still be only 4 bands present
Fragment D would be the heaviest because it is the largest piece of DNA and would thus have the
greatest mass.
The larger the DNA fragment, the slower it migrates through an agarose gel
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8
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The right lane is the molecular weight standards because it contains the known DNA fragments
Upper band- 5,000
Lower band- 4,000
Compared to both unknown bands to the migration of the known bands in the reference lane
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4000 bp
This standard curve is fairly linear, this estimation is fairly accurate, probably within 10% of true
value
2 |Chapter 4: Lab Activity 4.4: Restriction Digest
Mr. Poruban