What do these terms mean to you?

1. DNA sequencing • DNA profiling/fingerprinting • Gene cloning • Transformation • Making artificial genes What do these terms mean to you? You have 5 min to discuss possible meanings and examples with your group!

2. DNA Amplification • Using the technique called polymerase chain reaction (PCR), researchers are able to create vast quantities of DNA identical to trace samples. This process is also known as DNA amplification. • Many procedures in DNA technology require substantial amounts of DNA to work with, for example; • DNA sequencing • DNA profiling/fingerprinting • Gene cloning • Transformation • Making artificial genes • Samples from some sources,including those shown here,may be difficult to obtain inany quantity. A crime scene (body tissue samples) A single viral particle (from an infection) Fragments of DNA from a long extinctanimal

4. PCR Equipment • Amplification of DNA can be carried out with simple-to-use PCR machines called thermal cyclers (shown below). • Thermal cyclers are in common use in the biology departments of universities as well as other kinds of research and analytical laboratories.

5. A DNA sample called the target DNA is obtained DNA is denatured (DNA strands are separated) by heating the sample for 5 minutes at 98C The temperature is lowered to 50C Primers (short strands of mRNA) are annealed (bonded) to the DNA Primer annealed The Process of PCR 1

6. The sample is heated to 60°C. A thermally stable Taqpolymeraseenzyme binds to the primers on each side of the exposed DNA strand. This enzyme synthesizes a complementary strand of DNA using free nucleotides. After one cycle, there are now two copies of the original sample. The Process of PCR 2 Nucleotides Nucleotides

7. Polymerase Chain Reaction Original DNASample Although only three cycles of replication are shown here, following cycles replicate DNA at an exponentialrate and can make literally billions of copies in only a few hours. Cycle 1 Cycle 2 Cycle 3

8. Can you match thedefinitionsbelowtothekeywordsonyourworksheet? • This enzyme can add complementary nucleotides to a DNA strand during DNA synthesis. It is similar to the human DNA polymerase responsible for copying your genome every time one of your body cells divides. • These are short pieces of single-stranded DNA that match up to DNA sequences either side of the region of genomic DNA that you would like to copy. One is a forward primer and one is a reverse primer. When they have bound to the complementary sequences on the genomic DNA template strand, they show the Taq polymerase where to start DNA synthesis. The primers are responsible for making sure that only the region of interest is copied. • This is double-stranded genomic DNA isolated from the cells of the organism being studied. It can be human DNA, plant DNA, mouse DNA, bacterial DNA, whatever DNA you would like to have copied! • Free floating single nucleotides must be present in the reaction because they are what the Taq puts in place during DNA synthesis.

9. Lets act it out!

10. Man it´s hot in here! 98°C

11. Phew…I´m glad it´s cooler! 50°C

12. Seems to be warming up again! 60°C

13. Man it´s hot in here! 98°C

14. Phew…I´m glad it´s cooler! 50°C

15. Seems to be warming up again! 60°C

16. Now complete your PCR notes!

17. FILL IN THE BLANKS!!