Objective: Understand How Humans Benefit from Bacterial Transformation
New Words: Insulin, recombinant DNA, plasmid, gene splicing
Irene is 10 years old and in the last few weeks, she suddenly experienced extreme tiredness,
weight loss, and increased thirst. Her parents were concerned, so they took her to the doctor.
Dr. Ross took Irene’s blood to test for her blood sugar. The results of the test indicated that
she was a diabetic. When the doctor shared the results, Irene broke into tears. The
combination of her sudden poor health and the news of her diabetes was too much for her to
handle. “I don’t understand. What is diabetes and how can I get better?” The doctor felt bad
for Irene. She said, “Irene, I can explain how this happened, and how we can fix it.”
What is Irene’s problem?
Irene is diabetic
Objective: Understand How Humans Benefit from Bacterial Transformation
New Words: Insulin, recombinant DNA, plasmid, gene splicing
People with diabetes may not have enough insulin or may
not be able to use it properly.
Insulin is a hormone that controls the level of blood sugar
(also called glucose) in your body.
The sugar then builds up in the blood and overflows into
the urine, passing out of your body unused. This deprives
you of an important source of energy.
What is Diabetes?
A disease in which a person has high blood sugar, because
the body does not produce enough insulin
Objective: Understand How Humans Benefit from Bacterial Transformation
New Words: Insulin, recombinant DNA, plasmid, gene splicing
All people with type 1 diabetes, and some people with type 2,
need to take insulin to help control their blood sugar levels.
Type 1 diabetes means your body doesn’t make any insulin.
Type 2 diabetes means your body either doesn’t make enough
insulin or doesn’t use it properly.
The goal of taking insulin is to keep your blood sugar level in a
normal range as much as possible so you’ll stay healthy.
Over time, high blood sugar levels can cause serious health
problems such as blindness and kidney failure.
How Can Irene Get Better?
Irene needs to take insulin
Objective: Understand How Humans Benefit from Bacterial Transformation
New Words: Insulin, recombinant DNA, plasmid, gene splicing
The first successful insulin preparations came from cows (and later pigs).
In the 1980's technology had advanced to the point where we could make
human insulin. The technology which made this approach possible was
the development of recombinant DNA techniques. In simple terms, the
human gene which codes for the insulin protein was cloned (copied) and
then put inside of bacteria.
A number of tricks were performed on this gene to make the bacteria
want to use it to constantly make insulin. Big vats of bacteria now make
tons of human insulin. From this, pharmaceutical companies can isolate
pure human insulin
How Can We Make Insulin?
The human gene which codes for the insulin protein was put inside of
bacteria
Objective: Understand the Main Steps in Bacterial Transformation
New Words: Insulin, recombinant DNA, plasmid, gene splicing, restriction enzyme
Step #1
• Remove DNA from human cell.
• Use restriction enzymes to cut a segment of DNA that contains a
gene of interest, for example, the gene regulating insulin production.
Objective: Understand the Main Steps in Bacterial Transformation
New Words: Insulin, recombinant DNA, plasmid, gene splicing, restriction enzyme
Step #2
• Remove a plasmid from a bacterium and treated with the same
restriction enzyme.
What is a plasmid?
A small, circular, DNA molecule present in Bacteria
Objective: Understand the Main Steps in Bacterial Transformation
New Words: Insulin, recombinant DNA, plasmid, gene splicing, restriction enzyme
Step #3
• Bind the insulin gene with the opened plasmid to form a recombinant plasmid.
Objective: Understand the Main Steps in Bacterial Transformation
New Words: Insulin, recombinant DNA, plasmid, gene splicing, restriction enzyme
Step #4
• The recombinant plasmid is re-inserted back into the bacterium.
Objective: Understand the Main Steps in Bacterial Transformation
New Words: Insulin, recombinant DNA, plasmid, gene splicing, restriction enzyme
Step #5:
• Bacteria clone into a large number of identical daughter cells
Objective: Understand How Humans Benefit from Bacterial Transformation
New Words: Insulin, recombinant DNA, plasmid, gene splicing
Step #6:
• The recombinant plasmid replicates as part of the bacteria’s DNA.
Objective: Understand How Humans Benefit from Bacterial Transformation
New Words: Insulin, recombinant DNA, plasmid, gene splicing
Step #7
• Insulin is produced by bacteria and extracted for human use.
Objective: Understand How Humans Benefit from Bacterial Transformation
New Words: Insulin, recombinant DNA, plasmid, gene splicing
Figure 13-9 Making Recombinant DNA
Gene for human insulin
Gene for human insulin
Bacterial cell containing gene for human insulin
DNA recombination
Recombinant DNA
Sticky ends
Plasmid
DNA insertion
Recombinant
DNA
Gene for human
insulin
Gene for human
insulin
Human Cell
Sticky
ends
Bacterial Cell
DNA
recombination
DNA
insertion
Bacterial
chromosome
Plasmid
Bacterial cell containing
gene for human insulin
Objective: Understand How Humans Benefit from Bacterial Transformation
New Words: Insulin, recombinant DNA, plasmid, gene splicing
8 •Bacteria clone into a large number of identical daughter cells
5 •Bind the insulin gene with the opened plasmid to form a recombinant plasmid.
1 •Extract the DNA from a human cell.
9 •Insulin is produced by bacteria and extracted for human use.
3 •Remove the plasmid from a bacterium.
6 •The recombinant plasmid is re-inserted back into the bacterium.
7 •The recombinant plasmid replicates as part of the bacteria’s DNA.
4
•Treat the bacterial plasmid with the same restriction enzyme.
2 •Use restriction enzymes to isolate a segment of (DNA) that contains the insulin gene.
Objective: Understand the Main Steps in Bacterial Transformation
New Words: Insulin, recombinant DNA, plasmid, gene splicing, restriction enzyme
•Describe the picture above following the numbered sequence.
•Assure to include these terms: restriction enzymes, DNA, plasmid, gene, recombinant
plasmid, replication, cloning, insulin.
Objective: Understand the Main Steps in Bacterial Transformation
New Words: Insulin, recombinant DNA, plasmid, gene splicing, restriction enzyme
Step #1
• Extract the DNA from a human cell
• Use restriction enzymes to isolate a segment of (DNA) that contains the
insulin gene
Objective: Understand the Main Steps in Bacterial Transformation
New Words: Insulin, recombinant DNA, plasmid, gene splicing, restriction enzyme
Step #2
• Remove the plasmid from a bacterium
• Treat the bacterial plasmid with the same restriction enzyme
What is a plasmid?
A small, circular, DNA molecule present in Bacteria
Objective: Understand the Main Steps in Bacterial Transformation
New Words: Insulin, recombinant DNA, plasmid, gene splicing, restriction enzyme
Step #3
• Bind the insulin gene with the opened plasmid to form a recombinant plasmid.
Objective: Understand the Main Steps in Bacterial Transformation
New Words: Insulin, recombinant DNA, plasmid, gene splicing, restriction enzyme
Step #3
• The recombinant plasmid is re-inserted back into the bacterium
Objective: Understand the Main Steps in Bacterial Transformation
New Words: Insulin, recombinant DNA, plasmid, gene splicing, restriction enzyme
Step #4:
• Bacteria clone into a large number of identical daughter cells .
Objective: Understand How Humans Benefit from Bacterial Transformation
New Words: Insulin, recombinant DNA, plasmid, gene splicing
Step #4:
• The recombinant plasmid replicates as part of the bacteria’s DNA.
Objective: Understand How Humans Benefit from Bacterial Transformation
New Words: Insulin, recombinant DNA, plasmid, gene splicing
Step #4
• Insulin is produced by bacteria and extracted for human use.
Objective: Understand How Scientists Manipulate DNA
New Words: E. coli, Ampicillin, GFP, plasmid, antibiotic
• Do Now: List 3 things you learned in the Harlem DNA Lab
http://www.dnalc.org/harlemdnalab/BacterialTrans.html
Objective: Understand How Scientists Manipulate DNA
New Words: E. coli, Ampicillin, GFP, plasmid, antibiotic.
•
Complete the table below:
LB
+GFP
LB/Amp
LB
•Non transformed bacteria •Non transformed bacteria •Transformed bacteria
What do
•LB (food)
you have in •LB (food)
•LB (food)
•Ampicillin
the dish?
•GFP
What is
your
hypothesis?
Non transformed
bacteria spread across
the entire plate
What are
the results?
Same as expected in
hypothesis
Same as expected in
hypothesis
How do you
explain it?
•Non transformed
bacteria feed on LB
•Nothing stop them from
growing
•Non transformed
bacteria killed by
antibiotic ampicillin
No growth
•All bacteria grow
across the entire
plate
•Less green colonies
+GFP
LB/Amp
•Transformed bacteria
•LB (food)
•Ampicillin
•GFP
Transformed bacteria
grow in the form of
green colonies
Same as expected in
hypothesis
•Hypothesis is true
•Transformed bacteria
grow in the form of
green colonies
•Non transformed and
transform bacteria feed
on LB
•Nothing stop them from
growing
• Only transformed
bacteria can survive and
grow in the presence of
antibiotic ampicillin