Transcript Engineering Recombinant DNA Vaccines

Nekeisha Nogal, Gabriel Pineda,
Keith Neeves, Judy Schoonmaker,
Delphi Chatterjee, Diane Ordway,
Erika Tinoco
Goals and Outcomes
• Unit: Engineering recombinant DNA vaccines
– Goal: To understand the molecular biology sequence of
antigen  vaccine
• Goals for Today: Making a recombinant DNA vector
– Students will understand how restriction enzymes work
– Students will understand how DNA ligase works
• Outcomes
1. Students will be able to organize the main steps in DNA
vaccine development
2. Students will be able to recognize the specificity of
ligation sites following cleavage by restriction enzymes
3. Students will be able to create a script that applies the
concepts of gene insertion
Course Context
Biology I or II
– Freshmen
– ~100 students
Prior knowledge
– DNA structure
– Central dogma
– Restriction enzymes
– Vaccines and immune response
Have you gotten YOUR HPV Vaccine?
Recombinant Vector
HPV Vaccine
http://hplusmagazine.com/2009/08/31/take-shot/
Accessed 8/3/12
Steps in DNA vaccine development
Cut vector and DNA insert with restriction enzymes
Purify recombinant vector
Ligate DNA insert into vector
Deliver recombinant HPV vector vaccine
Determine coding DNA for insert
Identify HPV immunogenic protein
Choose a vector
Culture transformed bacterial host cells
Steps in DNA vaccine development
Identify HPV immunogenic protein
Determine coding DNA for insert
Choose a vector
Cut vector and DNA insert with restriction enzymes
Ligate DNA insert into vector
Culture transformed bacterial host cells
Purify recombinant vector
Deliver recombinant HPV vector vaccine
Cut vector and
DNA insert with
restriction
enzymes
Campbell Essential Biology with
Physiology, Simon et al., 3rd Ed.
The restriction enzymes RE1 and RE2 cut along the red lines:
If one sample of DNA was cut with RE1 and another with RE2, and
these two samples were mixed and treated with DNA ligase, what
would happen?
A. DNA cut with RE1 and RE2 would be ligated (joined together)
with no preference.
B. The RE1-cut DNA would be ligated to itself.
C. The RE2-cut DNA would be ligated to itself.
D. Both B and C.
E. No DNAs would be ligated.
Ligate HPV specific DNA insert into vector
1. Write a script for this movie using these
terms:
A.
B.
C.
D.
E.
F.
Restriction enzyme
Plasmid
DNA ligase
DNA insert
Sticky ends
Restriction site
2. Peer review script
3. Turn in revised script to teacher.
http://www.youtube.com/watch?v=aA5fyWJh5S0
Have you gotten YOUR HPV Vaccine?
Recombinant Vector
HPV Vaccine
http://hplusmagazine.com/2009/08/31/take-shot/
Accessed 8/3/12
Homework Assignment
(summative assessment)
• Make a concept map that shows how a scientist would
proceed from a gene sequence for a protein to a vaccine
using recombinant technology.
• You should use the terms below as a skeleton framework.
Add as many additional terms as you feel appropriate.
•
Restriction enzyme
1.
2.
3.
4.
5.
Vector
DNA ligase
DNA insert
Sticky ends
Vaccine
Exam Question
(summative assessment)
If you had an HPV-specific DNA insert that has sticky
ends created by digestion with BamHI, which of the
three restriction enzymes below could be used to cut a
vector to create a recombinant vector vaccine for HPV?
A.
B.
C.
D.
E.
BamHI only
BamHI and BslI
BslI and EcoRI
EcoRI only
EcoRI and BamHI
BamHI – 5’-G|GATCC
BslI – 5’-C|GATCG
EcoRI – 5’-G|AATTC
Alignment
Learning Objective
Assessment
Active
learning
Low Order/
High Order
Students will be able to
organize the main steps in
DNA vaccine development
Formative: Selfcheck based on key
Strip
sequence
Low Order
Bloom 3
Students will be able to
recognize the specificity of
ligation sites following
cleavage by restriction
enzymes
Formative: Clicker
question-two polls
Clicker
question +
peer
instruction
High Order
Bloom 3-4
Students will be able to
create a script that
synthesizes the key
concepts of gene insertion
Formative: Peer
evaluation
Writing script
of movie
High Order
Bloom 5
Summative: Grade
the script
Diversity
Different learning styles:
Kinesthetic – strip sequence
Visual – movie
Writing – script
Auditory – movie sounds
Different personalities:
Activities that appeal to introverts and extroverts