1. DNA Technology
Packet #50
Chapter #20

2. Introduction
Since the 1970’s, humans have been attempted to manipulate and modify genes in a way that was somewhat predictable.

3. Introduction II
Scientists would select a gene to be inserted into an organism
Cut two DNA molecules into fragments using restriction enzymes
Splice the fragments together into the desired combination
Producing recombinant DNA
Introduce the new DNA into a living cell for replication

4. Introduction III
Recombinant DNA technology isolates and amplifies specific sequences of DNA by incorporating them into vector DNA molecules.

5. The Players Involved in the Making of Recombinant DNA

6. Introduction
Recombinant DNA is made by splicing a DNA fragment of interest into a small quickly dividing replicating molecule (plasmid).

7. Donor & Transgenic Organisms
The organism providing the DNA is called the donor.
After recombination, an organism that contains an artificially inserted, foreign piece of DNA, is called a transgenic organism.

8. The Importance of Transgenic Organisms
Transgenic organisms allow gene targeting and mutagenesis screening that help identify the function of a gene and its protein product.

9. Vectors
The DNA that is removed from the donating organism.

10. Restriction Enzymes
Enzymes that are used to cut DNA into specific fragments.
Each restriction enzyme recognizes and cuts DNA at a highly specific base sequence.

11. The Making of Transgenic Organisms

12. The Making of a Transgenic Organism
The DNA of interest is excised, from the donor, using “scissors” known as a restriction enzyme.
The excised DNA is called a vector.

13. The Making of a Transgenic Organism
The vector is inserted via one of multiple methods.

14. The Making of a Transgenic Organism
DNA ligase is used to join the DNA fragments together.
DNA ligase joins the vector with the genome of the receiving organism.

15. The Making of a Transgenic Organism
Transgenic organism is duplicated.

16. Vectors Currently Under Study

17. Vectors Under Study… Vectors currently under study include
Retroviruses
Adenoviruses
Herpes simplex virus
Rhinovirus
Human Immunodeficiency Virus (HIV)

18. Genomics & Genetic Libraries

19. Genomic Library & cDNA Library
DNA library containing an organism’s complete genome
In the form of thousands of DNA fragments
cDNA Library
DNA library made up of “DNA clones” reconstructed using reverse transcriptase
Must be made from mRNA
Genomics
Sub-discipline in genetics of characterizing the entire genomes of organisms.

20. Homework Assignment
What are some of the advantages, and disadvantages, of having a cDNA library?

21. Genetic Probes

22. Genetic Probes
Genetic probes are radioactively labeled DNA or RNA sequence that enables geneticists to identify complementary nucleic acid sequences.
If used to identify a DNA strand, the DNA molecule must have been separated into two strands via artificial denaturation—heat.

23. Genetic Probes Southern Blot Technique
DNA fragments, produced using restriction enzymes, are separated via gel electrophoresis.
Fragments are then denatured
Fragments are blotted onto a nitrocellulose or nylon membrane.

24. Homework Assignment
Define Northern Blot.
Define Western Blot.

25. Making Copies of DNA in a Lab Setting

26. Introduction
Once a sequence of DNA has been isolated, it is sometimes necessary to make large amounts of that sequence for study.

27. Polymerase Chain Reaction
Allows rapid, efficient amplification of DNA sequences of interest.
In vitro technique
Researchers target a particular DNA sequence, by specific primers, and then clone the DNA sequence by heat resistant DNA polymerase.
Used to help amplify DNA from crime scenes and archaeological remains

28. Gene Therapy

29. Gene Therapy Simple idea—hard to practice
The use of sequencing, cloning and vector insertion techniques to deliver working versions of genes to individuals who are born with deleterious mutant versions of the gene.
Germ Line Therapy
Somatic Gene Therapy

30. Genetic Engineering & Food

31. Genetic Engineering of Agricultural Species
Foreign genes, under study, for insertion into commercial plant species. Helps provide
Selective herbicide resistance
Increased yield
Plant-grown vaccines and pharmaceuticals
Improved nutrient balance
Problems?
Human allergic reactions to foreign proteins
Increased use of herbicides
“jumping” of plasmids from commercial crops to weed species.
Eco-mayhem!

32. Review