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Transcript of Agenda – Applying DNA knowledge to diabetes Warm-up: Attractive & professional thank-you notes...
Agenda – Applying DNA knowledge to diabetes
Warm-up: Attractive & professional thank-you notes
Diabetes, Insulin, and rDNA Review
Recombinant DNA Plasmids Recombinant Paper Plasmid
Recombinant DNA project
Notes on what you have learned – due tomorrow
Electronic presentations http://prezi.com/ezf2bged4bia/genetic-
engineering-and-biotechnology/Genetic Engineering and Biotechnology by Dong Min Kim on Prezi
Diabetes and Biotechnology
Biotechnology solutions
Recombinant DNA
Questions- What is diabetes? Why is there growing concern about
diabetes? What is the role of insulin?
What is the role of biotechnology in the treatment of diabetes?
Biotechnology & Diabetes Treatment Read Calorie-Coated Diabetes
Beta Cell in Pancreas
Pancreas, thymus
Cell Biology
Of Insulin
Response
Cell Biology Of Insulin Response
Cell Biology Of Insulin Response
Insulin
A small, simple protein
Gene is on the 11th
Chromosome(short arm)
It contains 153 bases.
Recombinant DNA: A desired gene to the plasmid. Bacteria is transformed to accept the plasmid.
Recombinant DNA with insulin gene inserted
Chromosome & Plasmid
Interesting facts about plasmidsWhat do plasmids do in bacteria?
In nature?
Why are plasmids used in recombinant DNA?
Interesting facts about plasmidsWhat do plasmids do in bacteria?
Extra genes Not essential for living Benefit for survival under
certain conditions
In nature, some plasmids Antibiotic genes Antibiotic resistant genes
Why are plasmids used in recombinant DNA?
Bacteria can be transformed by adding circular DNA but not linear
Exonuclease: Breaks down genes in a
linear process Outside to the inside
Making a Recombinant DNA Product What are the steps needed?
Paper plasmids - Construction of the pAMP and pKAN Plasmids Questions
Genetic Engineering –Recombinant DNA How? Identify a molecule produced by a living
organism Isolate the instructions (DNA sequence =
gene) Put the instructions into another cell or
organism Allow the cell to replicate Harvest the desired product
The Vector (E. coli bacteria).
What is needed to produce a product with recombinant DNA?
Recombinant DNAInsulin
Identify a insulin gene in humans Use restriction enzymes to isolate the gene in a
DNA fragment DNA fragment is added to another DNA source =
vector such as plasmids of bacteria or yeast
Recombinant DNA is placed in a host cell As the host cell divides (replicates), the rDNA also
replicates Harvest, purify, test & market
What we need to learn about:
Bacteria (Vector) Copying cells - Mitosis Proteins How are enough insulin is made? PCR
Recombinant DNA & Cloning
What is DNA cloning?
• When DNA is extracted from an organism, all its genes are obtained
• In gene (DNA) cloning a particular gene is copied (cloned)
Why Clone DNA?
A particular gene can be isolated and its nucleotide sequence determined
Control sequences of DNA can be identified & analyzed
Protein/enzyme/RNA function can be investigated Mutations can be identified
e.g. gene defects related to specific diseases Organisms can be ‘engineered’ for specific purposes
e.g. insulin production, insect resistance and more
How is DNA cloned?, I
• DNA is extracted- here from blood
• Restriction enzymes, e.g. EcoR I, Hind III, etc., cut the DNA into small pieces
• Different DNA pieces cut with the same enzyme can join, or recombine.
Blood sample
DNA
Restriction enzymes
DNA Cloning, II
• Bacterial plasmids (small circular DNA additional to a bacteria’s regular DNA) are cut with the same restriction enzyme
• A chunk of DNA can thus be inserted into the plasmid DNA to form a “recombinant” molecule
DNA cloning, III
• The recombinant plasmids are then mixed with bacteria which have been treated to make them “competent”, or capable of taking in the plasmids
• This insertion is called transformation
DNA Cloning, IV
• The plasmids have naturally occurring (or inserted) genes for antibiotic resistance
• Bacteria containing plasmids with these genes will grow on a medium containing the antibiotic- the others die, so only transformed bacteria survive
DNA Cloning, V
• The transformed bacterial cells form colonies on the medium
• Each cell in a given colony has the same plasmid (& the same DNA)
• Cells in different colonies have different plasmids (& different DNA fragments)
Screening, IScreening can involve:1. Phenotypic screening-
the protein encoded by the gene changes the color of the colony
2. Using antibodies that recognize the protein produced by a particular gene
Screening, II 3. Detecting the DNA sequence of a cloned gene
with a probe (DNA hybridization)
rDNA project Choice of format:
PowerPoint Presentation with notes Globster (electronic poster) Museum boxes
Presentation: Process to make rDNA Process before rDNA Benefits Disadvantages or bioethical issues Regulation
Equivalent to Test Grade
Additional Resources http://www.accessexcellence.org/AE/AEC/CC/
restriction.php
Discovery and Applications of REs - http://videos.howstuffworks.com/hsw/22635-discoveries-with-bill-nye-restriction-enzymes-video.htm
http://tools.neb.com/NEBcutter2/
http://www.accessexcellence.org/AE/AEC/CC/activity1.php