Download Activity 1.4.2: Vaccine Development Introduction

Activity 1.4.2: Vaccine Development
Introduction
The discovery of vaccination was one of the farthest reaching medical discoveries of
all time. Vaccines have dramatically decreased the impact of infectious diseases
which were once considered deadly. Although the overall goal of creating a
weakened version of the antigen to kick-start the immune system is the same,
various laboratory techniques are used to generate vaccines. Scientists must be
concerned with safety of the vaccine, ease of the delivery system, and its overall
effectiveness over time. Technology for producing vaccines continues to evolve. In
this activity, you will explore the methods used to develop vaccines as well as
simulate the use of recombinant DNA techniques to produce these powerful
interventions.
Recombinant DNA technology is the process of cutting and recombining DNA
fragments. Since DNA is comprised of the same nucleotides in all species,
segments of DNA can be combined from two different people, or even two different
species. In addition to one large chromosome, bacteria carry additional genes in
rings of DNA called plasmids. Plasmids can be employed as an important tool in
genetic engineering and can serve as vectors, vehicles for the movement of genetic
information. Genes, segments of DNA that code for the production of a protein, can
be inserted into plasmids and the plasmids can then be inserted into bacterial cells.
Cells containing a new plasmid are cloned, the plasmid replicates and proteins are
transcribed and translated in the host cell. Scientists can use tiny bacterial cells as
factories for the production of proteins. These proteins can be used for various
medications, including vaccines.
In this activity, you will research the various methods used to produce vaccines. You
will then try your hand at genetic engineering and simulate the formation of a
recombinant plasmid used to produce a vaccine for the Hepatitis B virus.
Equipment
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Computer with Internet access
Activity 1.4.2 Student Resource Sheet
Paper plasmid DNA
Scissors
Scotch tape
Colored pencils
Laboratory journal
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MI Activity 1.4.2 Vaccine Development – Page 1
Procedure
Part I: Vaccine Basics
1. Take notes in your laboratory journal as the teacher presents the Making
Vaccines presentation.
2. Visit the NOVA Making Vaccines site at
http://www.pbs.org/wgbh/nova/meningitis/vaccines.html. Complete the
Making Vaccines activity. Take notes on the six different methods for producing
vaccines. For each type of vaccine make sure to describe the steps involved in
the development of the vaccine, summarized in no more than four steps, as well
as include an example of a vaccine produced by each method.
3. Answer Conclusion question 1 and 2.
Part II: Recombinant DNA Technology
Now that you have researched the types of vaccine preparations, you will simulate the
production of a vaccine through recombinant DNA technology.
4. Open your laboratory journal to the drawing of a bacterial cell you created in
Lesson 1.2 and review the way in which bacteria store genetic information.
5. Answer Conclusion question 3-4.
6. Visit the Dolan DNA Learning Center DNA Interactive site at
http://www.dnai.org/b/index.html.
7. Choose the Revolution tab at the bottom of the page to learn more about how
scientists are able to manipulate and work with particular genes.
8. Choose the Problem tab at the top of the page. Read the information presented
in this section using the small arrow at the bottom of the screen to advance the
slides.
9. In your laboratory journal, describe the two tools of molecular biology that are
often described as the scissors and glue for manipulating DNA. Describe how
these tools of molecular biology combined with plasmids can be used to make
copies of an important gene.
10. At the end of the Problem section, click on the Techniques tab at the bottom of
the page.
11. Click on Cutting and Pasting.
12. Click on Cutting and Pasting DNA to view the 2D animation. Use the arrow at the
bottom right-hand corner of the screen to advance the animation.
13. Click on the Recombinant DNA tab at the left side of the page to view the 3D
animation. Watch the video and read the text below.
14. View the Access Excellence diagram Inserting a DNA Sample into a Plasmid at
http://www.accessexcellence.org/RC/VL/GG/inserting.php and read the
description found below the diagram. Connect concepts you learned from the
DNA Interactive site to the process shown on the diagram.
15. Answer Conclusion question 5.
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MI Activity 1.4.2 Vaccine Development – Page 2
Bacterial plasmids can be used to generate protein products that are part of a disease
antigen. These products can be used to trigger an immune response. In this activity,
you will engineer a plasmid to produce a protein used as a vaccine against the virus,
Hepatitis B. You will transfer a gene for a viral envelope protein for Hepatitis B into a
bacterial plasmid. This protein resides on the surface of the virus and assists with
identity and infection. Once copies of this protein are made in bacterial cells, the protein
can be purified for use as a vaccine.
16. Recall that each restriction enzyme recognizes a specific DNA sequence called a
restriction sequence or restriction site. When the enzyme encounters this
sequence, it makes a distinct cut in the DNA. Refer to the chart below for the
restriction sites of five common enzymes.
Restriction Enzyme
BamHI
EcoRI
HaeIII
HindIII
PstI
Restriction Site
Cut
5’ GGATCC
5’ ---G
GATCC--- 3’
3’ CCTAGG
3’ ---CCTAG
5’ GAATTC
5’ ---G
3’ CTTAAG
3’ ---CTTAA
5’ GGCC
5’ ---GG
CC--- 3’
3’ CCGG
3’ ---CC
GG--- 5’
5’ AAGCTT
5’ ---A
3’ TTCGAA
3’ ---TTCGA
A--- 5’
5’ CTGCAG
5’ ---CTGCA
G--- 3’
3’ GACGTC
3’ ---G
G--- 5’
AATTC--- 3’
G--- 5’
AGCTT--- 3’
ACGTC--- 5’
17. Answer Conclusion question 6.
18. Obtain a Student Resource Sheet, a paper plasmid, scissors, tape and colored
pencils from your teacher.
19. Refer to the table of restriction enzymes and their restriction sequences shown
above or on the Student Resource Sheet. Scan the plasmid DNA and locate the
restriction sites for each enzyme. These sites have been shaded in or colored for
you. Identify the enzyme that would target each designated site and create a
color key in your laboratory journal.
20. Scan the viral DNA and locate restriction sites for each enzyme. Using the same
color key you designated in Step 20, shade each restriction site with the
appropriate color using colored pencils.
21. Scan the plasmid DNA as well as the gene from the Hepatitis B virus to find an
appropriate enzyme to engineer the plasmid. Your goal is to open the plasmid
ring and insert a piece of the viral gene. Follow the guidelines listed below to
determine the most efficient enzyme to complete your task.
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MI Activity 1.4.2 Vaccine Development – Page 3
o Use an appropriate restriction enzyme to open the plasmid ring as
well as cut out a piece of the viral gene. You do not want to use an
enzyme that digests the plasmid into many tiny pieces.
o Do not cleave more than twenty base pairs off either side of the viral
DNA. The remaining base pairs are required for production of the
protein of interest. Important promoters at the beginning of the
sequence are vital to transcribing the genetic code.
o Choose an enzyme that does not interrupt the antibiotic resistance
gene in the plasmid or the origin of replication (ori), the sequence
that initiates replication of the plasmid. The antibiotic resistance
gene provides scientists with a way of determining if an engineered
plasmid successfully made it into a target cell. If bacterial cells that
were not antibiotic resistant now grow on medium with antibiotics
present, they must contain the new plasmid.
o Make sure you create sticky ends on both the plasmid and the viral
DNA.
22. When you think you have located an appropriate enzyme, present your plan to
the teacher. Be prepared to defend your recommendation with evidence on your
DNA.
23. Use scissors and tape to create your recombinant plasmid. Be sure to make your
cuts according to the pattern shown on the restriction enzyme table.
24. Examine your completed plasmid. Compare your final product to that of a
classmate’s. Discuss any differences.
25. Note that an engineered plasmid must be put back into a bacterial cell to produce
the desired protein. Refer to your discussion of bacterial gene transfer in Lesson
1.2. In your laboratory journal, describe which gene transfer method can most
likely be used to insert the new plasmid into a bacterial cell.
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MI Activity 1.4.2 Vaccine Development – Page 4
Name: ____________________________
Date: ________________
Conclusion
1. Which types of vaccines described on the website are examples of live vaccines?
Explain your reasoning. (2)
2. How does vaccination contribute to the development of herd immunity in a
community? (2)
3. What are the similarities and differences between the organization and structure
of DNA in bacterial cells and in human cells? (3)
4. Why do you think plasmids and bacterial cells are such important tools in genetic
engineering? (1)
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MI Activity 1.4.2 Vaccine Development – Page 5
5. Explain why a plasmid and a segment of DNA to be inserted into a plasmid must
be cut with the same restriction enzyme. Use the term sticky ends in your
response. (2)
6. Explain why HaeIII is unlikely to be used to create recombinant DNA. How does
this enzyme differ from the others described in the table? (2)
7. What molecular tools do scissors and tape represent in this activity? Explain. (1)
8. Edible vaccines, a more controversial approach to vaccine development, have
been investigated by scientists. Plants can be genetically modified to produce
viral proteins and when ingested, these proteins ignite the immune system of the
consumer. What are the advantages to edible vaccines? What are some of the
possible disadvantages? (2)
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MI Activity 1.4.2 Vaccine Development – Page 6
9. How could you compare the original plasmid and the engineered plasmid in the
lab using restriction digestion and gel electrophoresis. Look at your paper
plasmid and think about the number of restriction sites for the enzyme you used
in the activity before and after you altered the plasmid. Describe your experiment
below. (3)
10. Jimmy has always been told that if you have chicken pox once, you will not get
the disease again. He had chicken pox when he was eight, but he only had five
to ten small pox on his skin. As an adult, Jimmy is again showing symptoms of
chicken pox. Using your knowledge of the human immune system, explain how
this is possible. (2)
11. A new flu vaccine is formulated and released every year. Explain why one shot
against influenza does not necessarily protect a person from year to year. Think
about the genetic material of the virus. (1)
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MI Activity 1.4.2 Vaccine Development – Page 7