DNA Technology - 2

What are plasmids? Small, circular DNA molecules (Found in bacteria) They are separate from the bacterial chromosome

Why are plasmids useful? They are used to manipulate genes in the lab They are small Contain genes useful to the bacteria And, are easily taken up by bacterial cells When they are taken up they are called vectors What happens when the bacterial cell replicates its chromosome? It also replicates the plasmid DNA (including any foreign DNA as well) vector A DNA carrier that move genes from one cell to another

What does cutting and pasting of DNA mean? Enzymes are involved e.g., A T C G For cutting, what are they called? Restriction enzymes They recognize specific sequence of: A T C G Hundreds have been isolated from bacteria For pasting, what are they called? DNA ligases Its the last step to make recombinant DNA They bind cut ends back together (by covalent bonds between adjacent nucleotides)

So, this cutting and pasting of DNA is used to: Cut gene sequence of interest (e.g., from human DNA) Enzyme name? Cut a plasmid (from a bacterium) Enzyme name? Create a recombinant DNA molecule By using the plasmid And ‘pasting’ a human gene sequence into it Enzyme name?

Polymerase Chain Reaction Or PCR PCR is a technique In which any segment of DNA can be copied Quickly and precisely Why is it so important? Use minute amounts of blood or other tissue To generate enough DNA for analysis e.g., DNA from the follicle of ONE stand of hair

How does PCR work? Make a mixture of: the DNA sample, some nucleotides, an enzyme, DNA polymerase Treat the mixture to: cycles of heating Allows separation of DNA strands cycles of cooling Allows DNA strands to re-form duplexes DNA Replication occurs during cooling cycle

In each cycle, the DNA is DOUBLED How many copies of DNA after each cycle? How many copies after a 5th cycle? ___1st cycle Within a few hours: ___2nd cycle PCR can generate billions of copies From a SINGLE DNA molecule ___3rd cycle ___4th cycle Enough to do extensive analyses

The Human Genome Project 1990 - 2003 What was the goal of HGP? To determine the nucleotide sequence all the DNA In any given human cell To identify the location & sequence of every gene What was discovered? Our DNA contains ~ 2.9 billion nucleotide pairs About 25,000 genes There is a LOT of DNA that isn’t made up of genes About 97% is non-coding DNA

Learning check 1. Why is only the slightest trace of DNA at a crime scene often sufficient for forensic analysis? 2. A carrier that moves DNA from one cell to another, such as a plasmid, is called a ________ 3. What features of a DNA fragment causes it to move through a gel during electrophoresis? Its nucleotide sequence The hydrogen bonds between its base pairs Its double helix shape The electrical charges of its phosphate groups

4. A paleontologist has recovered a bit of organic 4. A paleontologist has recovered a bit of organic material from the 400 year old preserved skin of an extinct dodo. She would like to compare DNA from the sample with DNA from living birds. The most useful method for increasing the amount of dodo DNA available for testing is __________ 5. Why is golden rice pale yellow in color? a It is rich in chlorophyll a. b It is nutrient-poor. c It is rich in beta-carotene. d It is rich in chlorophyll b. e It is rich in phycobilins.

What does this figure show? 2 4 7 3 1 8 6 5 What does each ‘band’ consist of?

Some Review

Structure and Function?

8 5 6 4 7 2 1 3

7 3 4 2 6 8 1 5 Name molecule Name of molecule Name the reaction Name of molecule What does the arrow refer to? Where does this take place?

15 10 4 7 11 1 6 9 14 2 3 12 8 16 17 5 13 18