1. Studying and Manipulating Genomes
Chapter 15

2. How are Genetic changes produced?
Natural processes
Mutation, variation
Humans have been changing the genetics of many species for thousands of years
Artificial selection of plants and animals
Humans changing genetics by combining genes between species
Genetic Engineering

3. Purpose of Genetic Engineering
Cure hereditary diseases
Engineer livestock to
Provide more nutrients
Make vaccines, vitamins
Engineer crops to
Produce pesticides
Increase productivity
Better nutrients
Basic research
Investigate basic genetic processes
Reconstruct life’s evolutionary history
Devise counterattacks against rapidly mutating pathogens

4. First Recombinant DNA
Paul Berg and associates were first to make recombinant DNA in 1972
Fused fragments of DNA from one species into the genetic material from another
Allowed them to isolate and replicate subsets of DNA from any organism

5. Tools for Constructing Recombinant DNA
Restriction Enzymes
Hamilton Smith was studying Haemophilus influenzae discovered bacteria have an enzyme that chops up viral DNA.
Will cut at a specific site on a specific sequence of DNA. The section has to be a palindrome!
Ex. GAATTC is cut by EcoRI
The same enzyme is used for both the DNA fragments so the overhanging ends are identical and complementary
Ligase used to glue them together

6. Making Recombinant DNA

7. Tools for Constructing Recombinant DNA
Plasmids as Vectors
Small circular DNA pieces,
can be used to transfer foreign DNA to bacteria
Why: So that the gene can be expressed and studied
Can also be inserted into eukaryotic cells directly Ex. Humans, plants etc.

8. Making Recombinant DNA
Recombine DNA to be studied with a Plasmid DNA. Transform bacteria with this plasmid. Can be studied and manipulated.

9. Tools for Constructing Recombinant DNA
PCR – Polymerase Chain reaction
Many rounds of DNA replication in a test tube to generate thousands of copies of a DNA segment.
Used for
forensics
Insertion and cloning
sequencing of DNA

10. Polymerase Chain Reaction

11. Polymerase Chain Reaction

12. Polymerase Chain Reaction

13. Polymerase Chain Reaction DNA replication in a tube!
Sequence to be copied is heated to break H bonds
Primers are added and bind to ends of single strands
Taq DNA polymerase uses free nucleotides to create complementary strands
Repeat Steps 1-3
Doubles number of copies of DNA at each round

14. Tools for Constructing Recombinant DNA
Gel Electrophoresis
Technique by which molecules of different sizes are separated and analyzed
Separated by electric charge - DNA is negatively charged and will migrate towards positive electric field
Rate of migration depends on size of molecule +

15. Tools for Constructing Recombinant DNA
T C C A T G G A C C
T C C A T G G A C
Tools for Constructing Recombinant DNA
T C C A T G G A
T C C A T G G
T C C A T G
Sequencing DNA
T C C A T
T C C A
Fluorescent labeled and fragmented DNA placed on gel
Fragments move off gel in size order; pass through laser beam
Color each fragment fluoresces is recorded on printout
electrophoresis
gel
T C C
T C
one of the many fragments of
DNA migrating
through the gel T
one of the DNA fragments
passing through a laser beam
after moving through the gel
T C C A T G G A C C A

16. Uses of genetic engineering: The Human Genome Project
Goal – Use these tools to map the entire human genome
Initially thought by many to be a waste of resources
Process accelerated when Craig Venter used bits of cDNAs as hooks to find genes
Sequencing was completed ahead of schedule in early 2001

17. Human Genome

18. Uses of genetic engineeringDNA Fingerprinting
Unique array of DNA fragments
Inherited from parents
Even full siblings can be distinguished from one another by this technique

19. Analyzing DNA Fingerprints
DNA is stained or made visible by use of a radioactive probe
Pattern of bands is used to:
Identify or rule out criminal suspects
Determine paternity