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Gene Technology Vocab Crunch! Target gene DNA we want to work with Target gene DNA we want to work with Reverse transcriptase enzyme that makes DNA from mRNA Reverse transcriptase enzyme that makes DNA from mRNA cDNA complementary DNA strands made from mRNA cDNA complementary DNA strands made from mRNA DNA Polymerase copies DNA DNA Polymerase copies DNA Vector object DNA will be placed into Vector object DNA will be placed into Plasmid small ring of DNA that can be added to a bacterial genome Plasmid small ring of DNA that can be added to a bacterial genome DNA Ligase reconnects sections of DNA backbone (C-P-C) DNA Ligase reconnects sections of DNA backbone (C-P-C) Restriction Enzymes cut DNA and plasmid in specific places so to make “sticky ends” (matching ends) Restriction Enzymes cut DNA and plasmid in specific places so to make “sticky ends” (matching ends)
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Gene Technology
What Is Gene Technology?• Gene Tech the alteration of genes inside a living organism to produce a specific results • Examples?• GM Foods:• Disease resistant potatoes• Pesticide producing tomatoes• Golden Rice• GM Pets:• Cloned pets• Glow in the dark pets
Gene Technology Vocab Crunch!• Target gene DNA we want to work with• Reverse transcriptase enzyme that makes DNA from mRNA• cDNA complementary DNA strands
made from mRNA•DNA Polymerase copies DNA• Vector object DNA will be placed into• Plasmid small ring of DNA that can be
added to a bacterial genome• DNA Ligase reconnects sections of DNA backbone (C-P-C) • Restriction Enzymes cut DNA and plasmid in specific places so to make “sticky ends” (matching ends)
I totally didn’t steal this image
Steps of Producing Insulin
1) mRNA is extracted from pancreas cells
2) Reverse transcription using reverse transcriptase
3) Single stranded DNA is made (cDNA)
4) DNA Polymerase is used to make dsDNA
5) Restriction enzyme is used to cut dsDNA to make
sticky ends6) Same restriction enzyme is used
to cut plasmid vector
Steps of Producing Insulin (II)
7) Open plasmid vector, ds DNA, and DNA ligase all mixed together
8) Plasmid vector produced with insulin gene inside (WOOT!)
9) Plasmid vector added to bacteria by transformation (electric shock)
10) Grow bacteria and test for insulin gene
11) Grow bacteria that successfully make insulin
12) Isolate, clean, and sell insulin protein
*le badly drawn mRNA
Insulin mRNA isolatedRT
*le incredibly well drawn cDNA
DNA polymerase
*le ok-drawn dsDNA
Restriction Enzyme
dsDNA with sticky ends
AATTTTAA
dsDNA with sticky ends
SameRestriction Enzyme
AATTTTAA
Plasmid
Cut plasmid
AATTTTAA
AATTTTAA
DNA Ligase
Insulin gene BacteriaTransformation
Antibiotic selection
PCR testing for gene
How Do We Know It Worked?1) Antibody selection• Add in penicillin resistant gene
to plasmid; only bacteria with plasmid should grow
2) PCR sample and run on a gel• Make copies of bacteria plasmid
and digest with same restriction enzymes and separate on a gel
• Same band = same gene3) Green Fluorescent protein• Add in gene to make green
fluorescent protein• Only green bacteria have
plasmid
Verbal cards
Debate:
“Should there be more regulation on the production and distribution of GM foods?”
