Quick Lab 15.2
Inserting Genetic Markers
Problem
How are restriction enzymes used to recombine DNA?
Introduction
Scientists know how to combine DNA from two or more sources. When the process is done right, a cell
will copy a gene that was inserted into its DNA when it copies the full strand of DNA. This technology has
been used to turn bacteria into “factories” for human proteins, such as insulin.
Scientists will often work with small circular pieces of DNA called plasmids, which are found in bacteria.
The human genes are inserted into the plasmids. Genetic markers are also inserted. The markers allow
scientists to track whether the recombined DNA was successfully introduced into the bacteria.
Scientists use restriction enzymes to control the location of inserted genes and markers. Bacteria
produce restriction enzymes. The enzymes as a defense against viruses that invade bacteria
Procedure
For each restriction enzyme, there is a recognition sequence that determines the location where the
enzyme can cut a DNA strand. For example, the enzyme Eco RI can cut a DNA strand between guanine
(g) and adenine (a) when the sequence GAATTC appears in the strand. In the table below, an asterisk (*)
shows the location of the cut in the recognition sequence for six restriction enzymes. Restriction
enzymes can read the sequence in both directions.
Enzyme
Bacterial Source
Recognition Sequence
Eco RI
Escherichia coli
G*AATTC
Hae III
Haemophilus aegyptius
GG*CC
Hha I
Haemophilus haemolyticus
GCG*C
Sma I
Serratia Marcescens
CCC*GGG
Taq I
Thermophilus aquaticus
T*CGA
1. Examine the base sequence below. Use a caret (^) to mark the location in the sequence where the Eco
RI enzyme could cut the DNA. Use a vertical line to mark a location where the Taq I enzyme could cut
the DNA.
CTT CGA TAT CAG ATT TAA ATC GCG TGT GTA CTT ACT GGG GAT CGC ATC ACG GTG AAT TCC CCC CTT
CTC TAC CTC CTT TGA ATT CGA ACT ACA GCG CTT CCC GGG TTT
2. Use a caret to mark the locations where the Hha I enzyme could cut the DNA. Read the sequence in
both directions
CTT CGA TAT CAG ATT TAA ATC GCG TGT GTA CTT ACT GGG GAT CGC ATC ACG GTG AAT TCC CCC CTT
CTC TAC CTC CTT TGA ATT CGA ACT ACA GCG CTT CCC GGG TTT
3. Use a caret to mark the location where both the Sma I enzyme and the Hae III enzyme could cut the
DNA
CTT CGA TAT CAG ATT TAA ATC GCG TGT GTA CTT ACT GGG GAT CGC ATC ACG GTG AAT TCC CCC CTT
CTC TAC CTC CTT TGA ATT CGA ACT ACA GCG CTT CCC GGG TTT
Analyze and Conclude
1. Predict Suppose the recognition sequence for a restriction enzyme appears three times along a DNA
strand. What could happen when the enzyme is used to insert a gene into a plasmid?
2. Infer Which restriction enzyme might give scientist more control over the location of an inserted
gene- Eco RI or Hae III? Why?
3. Infer How could restriction enzymes help bacteria defend against viruses? (Hint- viruses contain
nucleic acids.)