1. Polymerase Chain Reaction (PCR)

2. PCRPCR PCR produces billions of copies of a specific piece of DNA from trace amounts of starting material. (i.e. blood, skin cells, bone) Allows scientists to isolate pure quantities of specific DNA sequences 2 30 = over 1 billion copies of a specific DNA fragment; large enough quantity to be analyzed

3. 46 Chromosomes code for 30,000 to 50,000 genes; only 5% of your DNA –Exons = DNA that is coded or expressed into proteins Noncoding DNA has more diversity; since this DNA rarely leaves the DNA to head to ribosomes –Introns = DNA that is rarely expressed –Increased number of mutations

4. Target DNA – the DNA that needs to be copied Primers – short pieces of DNA that are designed to attach to each end of the DNA fragment that will be replicated Taq polymerase – enzyme that reads the DNA –Comes from the bacteria Thermus aquaticus –Lives in the hot springs in Yellowstone; doesn’t fall apart in high temperatures dNTPs – 4 nucleotides with the 4 different bases that are needed to replicate DNA Buffer – gives the best environment for the enzymes to work

6. PCR machine is known as thermal cylcer Machine changes to three different temperatures during one cycle Average number of cycles per run is 30 to 40

7. Temperature at 94°C –The target DNA falls apart –The H bonds holding the nitrogen bases together break –2 individual strands of DNA are now present instead of a double helix.

8. Annealing Temperature Temp between 56-65 –Primers attach to the ends of the Target DNA that needs to be copied –Annealing = attachment of the primers –Attach to complimentary bases of target DNA

9. Extension Temperature Temperature at 72°C –Provides best temp for Taq polymerase to begin reading the DNA –Taq polymerase will synthesize a second strand of complimentary DNA –Taq polymerase always read target DNA from 3’ to 5’ end

10. Repeat 30 times The three temperature changes represents one cycle –Denature –Anneal –Extend Repeat 30 times 2 30 = over 1 billion copies of the Target DNA Once DNA is amplified (copied), it is visible on a gel

11. PCR Animation

12. By the 4th Cycle = 32 Copies

13. D stands for the chromosome, and the S stands for map location of the chromosome, and the 80 is the locus point Locus is on chromosome 1 Intron – noncoding region of Chromosome 1 Each person has two copies of D1S80, one from each parent VNTR – Variable Number of Tandem Repeats Consists of a repeating 16 base pattern (10 repeats to >40 repeats) Depending on how many repeating patterns present, determines the size of your D1S80 locus

14. Homozygous or Heterozygous

15. Determining your genotype!

16. Locus with variability D1S80

17. Huntington’s Chorea Found on Chromosome 4 Noncoding region that actually causes genetic disease People with Huntington’s have a section on chromosome 4 that has 35 or more of three base repeating pattern CAG (trinucleotide repeat) CAG normally codes for glutamine Huntington’s patients will have a long line of glutamine produced

18. Effects of Huntingtons

19. Dominant Autosomal Disease