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Amgen Biotech Labs
OverviewIn this set of labs, students will:
• Lab 1.1: Learn how to use a pipette to dispense small volumes of solutions
• Lab 1.2: Learn how to use the technique of gel electrophoresis to separate molecules based on their size
• Lab 2: Learn how to use restriction enzymes to cut DNA at specific locations
• Lab 3: Learn how to use DNA Ligase to join together DNA fragments
• Lab 4: Learn how to use gel electrophoresis to check to verify successful creation of a plasmid with the desired genes
• Lab 5: Learn how to transform competent cells with a plasmid containing a desired gene (that is, insert a desired gene into bacteria so you can grow it to produce the desired protein)
• Lab 6: Isolate and purify the desired protein
Animations on DNA Sequencing, DNA Extraction, Cloning, Gel Electrophoresis, loading a gel, etc are all available on the Amgen Biotech site:http://www.amgenbiotechexperience.com/curriculum/curriculum-resources
Biotechnology Overview with a lot of good info: http://biotechnology.amgen.com/index.html
Amgen Biotech Labs
What is Genetic Engineering?
…the branch of biology that uses special procedures and techniques to change an organism’s DNA.
One example is removing DNA from one organism and inserting it into another organism.
In this image, the gene for insulin has been remove from human DNA and inserted into a bacterial plasmid. The plasmid with the human insulin gene is then put into a bacterium which reproduces to make many copies. Each bacterial cell produces the human insulin protein which can be purified from the solution.
Amgen Biotech Labs
Insulin is the hormone responsible for regulating glucose levels in the blood
In this image, the gene for insulin has been remove from human DNA and inserted into a bacterial plasmid. The plasmid with the human insulin gene is then put into a bacterium which reproduces to make many copies. Each bacterial cell produces the human insulin protein which can be purified from the solution.
Diabetes Movie: http://www.amgenbiotechexperience.com/sites/abe.edc.org/files/abe_sg_s2.pdf (8:40 min)
Amgen Biotech Labs
OverviewIn this set of labs, students will:
• Lab 1.1: Learn how to use a pipette to dispense small volumes of solutions
Amgen Biotech Labs
OverviewIn this set of labs, students will:
• Lab 1.1: Learn how to use a pipette to dispense small volumes of solutions• Lab 1.2: Learn how to use the technique of gel electrophoresis to separate
molecules based on their size
Negatively charged molecules (like DNA) are attracted to the positive electrode. Smaller fragments move through the porous gel faster than larger fragments.
Amgen Biotech Labs
OverviewIn this set of labs, students will:
• Lab 1.1: Learn how to use a pipette to dispense small volumes of solutions• Lab 1.2: Learn how to use the technique of gel electrophoresis to separate
molecules based on their size
Amgen Biotech Labs
OverviewIn this set of labs, students will:
• Lab 2: Learn how to use restriction enzymes to cut DNA at specific locations.
Amgen Biotech Labs
OverviewIn this set of labs,
students will:• Lab 2: Learn how
to use restriction enzymes to cut DNA at specific locations.
Amgen Biotech Labs
OverviewIn this set of labs, students will:
• Lab 2: Learn how to use restriction enzymes to cut DNA at specific locations
Amgen Biotech Labs
OverviewIn this set of labs, students will:
• Lab 2: Learn how to use restriction enzymes to cut DNA at specific locations.
Cut two genetically engineered plasmids into DNA fragments that contain specific genes of interest:
From the Plasmid pKAN-R red fluorescent protein (rfp) – the protein we want to grow and isolate promotor (pBAD) – site at which RNA polymerase binds to DNA to initiate
transcription (an RNA copy of the genes are made which then moves to the ribosomes which use them to build the proteins)
From the plasmid pARA ampicillin resistance gene (ampR) – enables us to selectively grow only bacteria
that have the plasmids with the desired genes in them arabinose activator gene (araC) – codes for a protein that blocks the promoter
unless the sugar Arabinose is present to remove it (thus the genes are only able to be activated and their proteins made when arabinose is mixed in)
Restriction Enzymes being used to make the cut: BamH I and Hind III
Amgen Biotech Labs
OverviewIn this set of labs, students will:
• Lab 3: Learn how to use DNA Ligase to join together DNA fragments
Join together the DNA fragments produced in Lab 2, resulting in the production of different plasmids with various combinations of genes:
Amgen Biotech Labs
OverviewIn this set of labs, students will:
• Lab 3: Learn how to use DNA Ligase to join together DNA fragments
Join together the DNA fragments produced in Lab 2, resulting in the production of different plasmids with various combinations of genes:
Amgen Biotech Labs
OverviewIn this set of labs, students will:
• Lab 4: Learn how to use gel electrophoresis to check to verify successful creation of a plasmid with the desired genes
Amgen Biotech Labs
OverviewIn this set of labs, students will:
• Lab 4: Learn how to use gel electrophoresis to check to verify successful creation of a plasmid with the desired genes
Amgen Biotech Labs
OverviewIn this set of labs, students will:
• Lab 4: Learn how to use gel electrophoresis to check to verify successful creation of a plasmid with the desired genes
Amgen Biotech Labs
OverviewIn this set of labs, students will:
• Lab 5: Learn how to transform competent cells with a plasmid containing a desired gene
1) Insert plasmid with desired gene into E coli bacteria 2) Verify that it worked by growing the bacteria in the presence of ampicillin (which acts as an indicator for transformation) and arabinose (which is required to activate the rfp gene)
Amgen Biotech Labs
OverviewIn this set of labs, students will:
• Lab 5: Learn how to transform competent cells with a plasmid containing a desired gene
1) Insert plasmid with desired gene into E coli bacteria
Amgen Biotech Labs
OverviewIn this set of labs, students will:
• Lab 5: Learn how to transform competent cells with a plasmid containing a desired gene
2) Verify that it worked by growing the bacteria in the presence of ampicillin (which acts as an indicator for transformation) and arabinose (which is required to activate the rfp gene)
Amgen Biotech Labs
OverviewIn this set of labs, students will:
• Lab 5: Learn how to transform competent cells with a plasmid containing a desired gene
2) Verify that it worked by growing the bacteria in the presence of ampicillin (which acts as an indicator for transformation) and arabinose (which is required to activate the rfp gene)
Amgen Biotech Labs
OverviewIn this set of labs, students will:
• Lab 5: Learn how to transform competent cells with a plasmid containing a desired gene
2) Verify that it worked by growing the bacteria in the presence of ampicillin (which acts as an indicator for transformation) and arabinose (which is required to activate the rfp gene)
