1. DNA Technology and Genomics
Chapter 20
DNA Technology and Genomics

2. DNA Cloning

3. Restriction enzymes Where are restriction enzymes found?
In bacterial cells, to help fight viruses
What is the difference between blunt and sticky ends?
What is a restriction site?
The area that the restriction enzyme recognizes
Why are restriction sites often palindromes?
A MAN A PLAN A CANAL PANAMA

5. Cloning The original plasmid is called the
cloning vector
After insertion of the gene of interest into the plasmid the plasmid is called
Recombinant DNA
How would the recombinant plasmid that we engineered in class be used in the lab?

7. Identifying the gene of interest
Use different probes
How would you identify different genes?

8. DNA Libraries
Shotgun approach makes thousands of different recombinant plasmids
This set of plasmid clones is called a
Besides using plasmids scientists use as cloning vectors
genomic library
bacteriophages

9. cDNA Library

10. Different libraries have different uses
Which library would you use in the following scenarios?
You want to clone a gene, without knowing where and how it is expressed.
You want to know regulatory sequences.
You want to know the coding sequences of a gene.
genomic library
genomic library
cDNA library

11. Expressing Eukaryotic Genes
How can scientists get bacterial cells to express eukaryotic genes?
Insert the desired gene with a prokaryotic promoter attached.
How do they remove introns?
Insert cDNA in the first place.
Use YACs (Yeast artificial chromosomes)…eukaryotic hosts

12. DNA Amplification
PCR = polymerase chain reaction
PCR Animation

13. Gel electrophoresis Uses to separate DNA DNA has a charge.
DNA moves towards the electrode.
DNA molecules of sizes move the furthest through the gel.
electrical current
negative
positive
smaller

15. RFLP Restriction enzymes DNA. Fragments that result are of sizes.
These differently sized fragments are called .
RFLP’s are found on noncoding segments of DNA and are due to .
cut
different
RFLP’s
VNTR’s

17. Southern Blotting Technique

18. Uses recombinants
Mapping the Genome
Uses restriction fragments

19. Sanger Sequencing

20. Shotgun Approach

21. Identifying Protein Coding Genes
What would you look for if you wanted to find a protein coding gene?
Start and stop codons
These sequences are called EST’s (expressed sequence tags)

22. Genome Sizes

23. Genomes
If there is no pattern between complexity and number of genes what makes organisms more complex?
More involved regulation
Alternative splicing

24. Determining Gene Function
How can we determine the function of different genes?
RNAi; silence a gene and see what it did
Insert synthetic double stranded RNA’s that will activate dicer.
This was used to identify the function of C. elegans genes

25. Genome wide expression
Resulting colors shows which genes were being expressed in each tissue
Use two tissue samples with differently colored fluorescent nucleotides

26. Comparing Genomes
Has revealed many similar regions even between yeast and us.
Can be used to derive evolutionary relatedness.
Can use differences between closely related species to explain phenotypic differences. (chimps and humans with regard to language acquisition).
Has led to proteomics.

27. Practical Applications
Medical
Disease diagnosis
Gene Therapy
Medicines
Forensics
Environmental Cleanup
Aggriculture
Pharm animals
GMO foods/plants

28. Safety and Ethics What are the fears? Is this ethical? Is it too fast?
Is it really that good in the long run?