Restriction Enzyme Activity - Answers
Where the DNA will be cut is indicated by a /
EcoR1:
ATCGG/AATTCCTCGAGCTTCGGATCCTTCTCGGATCCATCGAAGCTTCGAGTCGC
TAGCCTTAAG/GAGCTCGAAGCCTAGGAAGAGCCTAGGTAGCTTCGAAGCTCAGCG

AGTGGATCCAGTCGTCAAGCTTCGAGCTATCTCGAAGCTTCG/AATTCAAGCTTCG
TCACCTAGGTCAGCAGTTCGAAGCTCGATAGAGCTTCGAAGCTTAA/GTTCGAAGC

A
T
BamH1:
ATCGGAATTCCTCGAGCTTCG/GATCCTTCTCG/GATCCATCGAAGCTTCGAGTCGC
TAGCCTTAAGGAGCTCGAAGCCTAG/GAAGAGCCTAG/GTAGCTTCGAAGCTCAGC

AGTG/GATCCAGTCGTCAAGCTTCGAGCTATCTCGAAGCTTCGAATTCAAGCTTC
GTCACCTAG/GTCAGCAGTTCGAAGCTCGATAGAGCTTCGAAGCTTAAGTTCGAAG

GA
CT
HindIII:
ATCGGAATTCCTCGAGCTTCGGATCCTTCTCGGATCCATCGA/AGCTTCGAGTCGC
TAGCCTTAAGGAGCTCGAAGCCTAGGAAGAGCCTAGGTAGCTTCGA/AGCTCAGCG

AGTGGATCCAGTCGTCA/AGCTTCGAGCTATCTCGA/AGCTTCGAATTCA/AGCTTC
TCACCTAGGTCAGCAGTTCGA/AGCTCGATAGAGCTTCGA/AGCTTAAGTTCGA/AG

GA
CT
Alu1:
ATCGGAATTCCTCGAG/CTTCGGATCCTTCTCGGATCCATCGAAG/CTTCGAGTCG
TAGCCTTAAGGAGCTC/GAAGCCTAGGAAGAGCCTAGGTAGCTTC/GAAGCTCAGC

CAGTGGATCCAGTCGTCAAG/CTTCGAG/CTATCTCGAAG/CTTCGAATTCAAG/CTT
GTCACCTAGGTCAGCAGTTC/GAAGCTC/GATAGAGCTTC/GAAGCTTAAGTTC/GAA

CGA
GCT
Taq1:
ATCGGAATTCCT/CGAGCTTCGGATCCTTCTCGGATCCAT/CGAAGCTT/CGAGTCG
TAGCCTTAAGGAGC/TCGAAGCCTAGGAAGAGCCTAGGTAGC/TTCGAAGC/TCAGC

CAGTGGATCCAGTCGTCAAGCTTC/GAGCTATCT/CGAAGCTT/CGAATTCAAGCT
GTCACCTAGGTCAGCAGTTCGAAGC/TCGATAGAGC/TTCGAAGC/TTAAGTTCGA

T/CGA
AGC/T
Questions:

Does each restriction enzyme digest the DNA into the same number of pieces? Why or why not?
Because each restriction enzyme recognizes a different sequence, the number of pieces of DNA will vary.
Which restriction enzyme has the most pieces? Why? Which restriction enzyme has the least pieces? Why?
Taq1 breaks the DNA into 8 pieces. The smaller the recognized sequence, the higher probability of that sequence occurring. Similarly, the longer sequences are less likely to occur often in the DNA, which is why EcoR1 breaks the DNA into only 3 pieces.
DNA has four bases, but there are over 100 different restriction enzymes. Why are there so many different restriction enzymes?
Since each restriction enzyme recognizes more than one base at a time, there is the possibility of many different restriction enzymes. No two enzymes will digest a DNA molecule the same way.
Look at the cuts made by AluI and HindIII. What is the relationship between the two sequences those restriction enzymes recognize?
The sequence recognized by Alu1 ( AGCT ) occurs in the middle of the recognition sequence of HindIII: AAGCTT.
You’re a geneticist that wants to cut out a gene with the sequence GGTCCTCGACGGATCCA out of the follow strand of DNA:
CTGAAGAATTCGGTCCTCGACGGATCCAGCTGAATTCGCTAGGATCCGAT
You don’t want to cut in the middle of the gene, and you don’t want too many excess base pairs on either side of the gene. What is the best restriction enzyme to use? Why?
The sequence recognized by EcoR1-GAATTC-occurs in front of the target sequence, as well as shortly afterwards, and also does not occur in the middle of the target sequence.
What would happen if you put all the DNA fragments into a gel and performed a gel electrophoresis? Mimic a gel electrophoresis with your group’s fragments. Put each different restriction enzyme into a different lane. The biggest pieces will be at the top of the gel, the smallest pieces will be at the bottom of the gel. Why would a scientists use different restriction enzymes?
Different restriction enzymes digest DNA in different ways, so scientists can get an idea of the frequency of a particular sequence when they use different restriction enzymes.