Transcript Genetic Engineering via Bacterial Transformation

• Summer 2008 Workshop
• in Biology and Multimedia
• for High School Teachers
Life Sciences-HHMI Outreach. Copyright 2008 President and Fellows of Harvard College.
AP Biology Lab 6:
Genetic Engineering
via
Bacterial Transformation
Making E. coli glow like jellyfish
Amy Dickson, Prospect Hill Academy Charter School
All images by Christine Rodriguez and Amy Dickson
Life Sciences-HHMI Outreach. Copyright 2008 President and Fellows of Harvard College.
WHY SHOULD WE DO THIS?
Genetic Engineering is now widely used:
•
Bacteria that produce human insulin
•
Corn that produces insecticide
•
Rice that produces extra vitamin A
•
Goats that produce spider silk
Life Sciences-HHMI Outreach. Copyright 2008 President and Fellows of Harvard College.
WHY SHOULD WE DO THIS?
To SEE the Central Dogma in action:
DN
A
GFP Gene
RN
A
• found in jellyfish
Protein
Green
Fluorescent
Protein
Trait
GLOWING
CELLS
• engineered into
bacteria
Life Sciences-HHMI Outreach. Copyright 2008 President and Fellows of Harvard College.
QUICK REVIEW
Promoter Plasmid -
an “on/off” switch for a gene
a small, circular piece of bacterial DNA
that is not part of the chromosome
Transformation - a process in which bacteria take up
DNA from their environment
- can be triggered by electric shock or
heat shock
Life Sciences-HHMI Outreach. Copyright 2008 President and Fellows of Harvard College.
STARTING MATERIALS
E. coli cells
• sensitive to antibiotics
• can’t glow
Bacterial
chromosome
• competent - able to be
transformed
Life Sciences-HHMI Outreach. Copyright 2008 President and Fellows of Harvard College.
STARTING MATERIALS
Plasmid containing:
• Ampicillin resistance
gene (always expressed)
• Ara promoter - turned on
in the presence of
arabinose
Life Sciences-HHMI Outreach. Copyright 2008 President and Fellows of Harvard College.
STARTING MATERIALS
Jellyfish DNA
GFP
gene
GFP = Green Fluorescent Protein
glows under UV light
Life Sciences-HHMI Outreach. Copyright 2008 President and Fellows of Harvard College.
STARTING MATERIALS
E. coli cells
Plasmid
GFP
Jellyfish DNA
Life Sciences-HHMI Outreach. Copyright 2008 President and Fellows of Harvard College.
END RESULT
GROW ON
AN AGAR
PLATE
Recombinant Bacteria…
… that can GLOW!
Life Sciences-HHMI Outreach. Copyright 2008 President and Fellows of Harvard College.
HOWEVER…
things are actually a bit more complex.
makes all transformed bacteria
resistant to ampicillin
pGLO
plasmid
controls GFP gene expression
only turned on in the presence of
arabinose
Life Sciences-HHMI Outreach. Copyright 2008 President and Fellows of Harvard College.
YOUR TASK:
Design an experimental procedure for
genetically engineering glowing bacteria.
Goals to consider:
#1 - Make recombinant bacteria
#2 - Select for only the recombinant bacteria
#3 - Make the recombinant bacteria glow
#4 - Establish a control for your experiment to
demonstrate that it’s the plasmid that causes
ampicillin resistance and the ability to glow.
Life Sciences-HHMI Outreach. Copyright 2008 President and Fellows of Harvard College.