1. Genetic Engineering Ch 15
“Real World Biology”

2. Selective Breeding Selective Breeding
People select organisms with desired characteristics to produce next generation
Takes advantage of naturally occurring variation
Selective breeding of teosinte grass by native Americans 6000 years ago led to corn as we now know it

3. Selective Breeding Hybridization
Cross dissimilar organisms to bring together best of both organisms
Ex: disease resistance + increased yield
Benefits include hardier plants
American botanist Luther Burbank developed more than 800 varieties of plants using selective breeding methods.

4. Selective Breeding Inbreeding
Breeding a line of organisms with similar characteristics
Ex: dog breeds
Risks- decreased genetic variation and increased susceptibility for certain diseases/disorders
Ex: hip dysplasia

5. Increasing Variation
Process used to increase the variation normally present in nature
But why?
Biotechnology is the application of a technological process, invention, or method to living organisms.

6. Increasing variation Can be accomplished through mutations
Mutations are usually random, but can be induced via radiation and chemical exposure
Potential to yield few beneficial mutants with desirable characteristics not found in original population

7. Increasing Variation
Bacteria- can treat millions at a time increasing chances of producing useful mutants
Ex: oil-digesting bacteria

8. Increasing Variation
Plants-arresting chromosome separation during meiosis to produce polyploids
Known to be more vigorous than diploid relatives

9. 13-2 Manipulating DNA Mutations are random
Having a way to alter DNA in a very specific way to achieve a particular result has huge advantages
Scientists can now use the knowledge of DNA structure and its chemical properties to study and change DNA molecules

10. Tools of Molecular Biologists
Genetic engineering allows biologists to rewrite the DNA code of an organism
Modern techniques employed can
Extracting DNA from cells
Cutting it into smaller pieces
Identifying sequences of bases in DNA (genes)
Making unlimited copies

11. Finding Genes Started with Douglas Prasher (1987)
Prasher wanted to find a specific gene in a jellyfish that codes for a molecule called green fluorescent protein, or GFP
GFP is a natural protein that absorbs energy from light and makes parts of the jellyfish glow
Prasher thought that GFP from the jellyfish could be linked to a protein when it was being made in a cell
bit like attaching a light bulb to that molecule

12. Finding Genes (GFP specifically)
Prasher compared part of the amino acid sequence of the GFP protein to a genetic code table
was able to predict a probable mRNA base sequence that would code for this sequence of amino acids
Then used a complementary base sequence to “attract” an mRNA that matched his prediction and would bind to that sequence by base pairing.
After screening a genetic “library” with thousands of different mRNA sequences from the jellyfish, he found one that bound perfectly

13. Finding Genes
To find the actual gene that produced GFP, Prasher took a gel in which restriction fragments from the jellyfish genome had been separated and found that one of the fragments bound tightly to the mRNA
That fragment contained the actual gene for GFP
This method is called Southern blotting, after its inventor, Edwin Southern.

14. Finding Genes- Southern Blot Analysis

15. Finding Genes
Today it is often quicker and less expensive for scientists to search for genes in computer databases where the complete genomes of many organisms are available.

16. Copying DNA (specific genes)
First step is a polymerase chain reaction (PCR)
Heat a piece of DNA
separates its two strands
DNA cools and added primers bind to the single strands
DNA polymerase starts copying the region between the primers
These copies can serve as templates to make still more copies.

17. Polymerase Chain Reaction
Once biologists find a gene, a technique known as polymerase chain reaction (PCR) allows them to make many copies of it.
1. A piece of DNA is heated, which separates its two strands.

18. Polymerase Chain Reaction
2. At each end of the original piece of DNA, a biologist adds a short piece of DNA that complements a portion of the sequence.
These short pieces are known as primers because they prepare, or prime, a place for DNA polymerase to start working.

19. Polymerase Chain Reaction
3. DNA polymerase copies the region between the primers. These copies then serve as templates to make more copies.
4. In this way, just a few dozen cycles of replication can produce billions of copies of the DNA between the primers.

20. Copying DNA
It is relatively easy to extract DNA from cells and tissues.
The extracted DNA can be cut into fragments of manageable size using restriction enzymes.
These restriction fragments can then be separated according to size, using gel electrophoresis or another similar technique

21. Gel Electrophoresis

22. Recombinant DNA Technology
It is a form of genetic engineering that cleaves DNA into small fragments and inserts those fragments into a host organism
Host may be the same or a different species

23. Transgenic Organisms
Organisms who have incorporated foreign DNA in their chromosomes and use this new DNA as their own

24. How to Produce a Transgenic Organism
Step 1: Isolate the gene in the foreign DNA that you want to insert
Ex: isolate the gene for beta carotene in a daffodil so you can then add it to rice

25. Step 2: Cut it out of the chromosome (in daffodil) using restriction enzymes.
Restrictions enzymes are bacterial proteins that have the ability to cut both strands of the DNA molecule at a specific nucleotide sequence
Resulting fragments can have blunt ends or sticky ends

26. Some Commonly used REs EcoRI (eco r one) HindIII (hindi three)
BamHI (bam h one)
TaqI (tack one)

27. Step 3: Cut host’s DNA with the same RE so cut ends will match up
When DNA from two different organisms joins up- recombinant DNA is formed

28. Vectors Getting DNA from one organism into another requires a vector
The vector introduces the new DNA into the host cell
Bacterial DNA is often used as a vector

29. Bacterial DNA
Bacteria contains plasmids- small rings of DNA separate from the bacterium’s larger circular chromosome
The foreign DNA is inserted into the plasmid by cleaving both using the same restriction enzyme
Sticky ends match up and foreign DNA becomes part of plasmid

30. Gene Cloning
Plasmid with foreign DNA (Now considered recombined DNA) is inserted into a bacterial cell
Plasmids can replicate within the cell and can produce up to 500 copies in the cell

31. Soon Tons of Copies! Bacteria clones the recombinant DNA How?
Clones-genetically identical copies
How?
Bacterial cells themselves will reproduce quickly, each with hundreds of copies of the recombinant DNA inside (plasmid + foreign DNA)

32. Introduction into Host Cell
Plasmid is then inserted into a host’s chromosome where it will be replicated each time the cell replicates along with the organism’s other chromosomes
The host cell can transcribe/translate that recombinant DNA into protein just like all other proteins coded in its DNA