DNA Fingerprinting

1. DNA Fingerprinting http://www.youtube.com/watch?feature=player_embedded&v=fV5kWZWY0P4

2. Where is DNA?

3. Why DNA? • Amino acids are the code that make up proteins. • Nucleotide bases are the code that makes up DNA • With only 4 nucleotide bases, but 20 amino acids, scientists believed that it must to be amino acids that are the genetic material due to their greater chance of variability • In 1944, McCarty, Avery, and MacLeod proved that DNA actually carried the genetic material

4. Nucleotides Bases of DNA • In 1952 Chargaff discovered in his experiments that the amount of adenine (A) in the DNA being used was ALWAYS equal to the amount to thymine (T) and the amount of guanine (G) was ALWAYS equal to the amount of cytosine (C)! • A – T, C – G • Known as Chargaff’s rule • He later met with Watson and Crick to share his discoveries. This knowledge helped them to discover the structure of DNA (1953)

5. DNA

6. Nucleotides Bases of DNA

7. Structure of DNA

8. DNA Replication

9. Polymerase Chain Reaction (PCR) Polymerase Chain Reaction was discovered by Kary Mullis in 1983, winning the Nobel Prize in 1993! (This is a huge accomplishment that the Nobel Prize was awarded only 10 years after the techniques introduction!) It is a technique used to amplify DNA Due to the limited amount of forensic samples, PCR has become an invaluable tool for the forensic scientist Scientists only need 0.1-1 ng of DNA… that’s 1 x 10-9 g! http://www.youtube.com/watch?v=2KoLnIwoZKU&feature=player_embedded#!

10. Polymerase Chain Reaction (PCR)

11. DNA  proteins • DNA is used as a template to made an RNA strand • From the RNA strand, sequences of 3 nucleotide bases (codon) code for specific amino acids • Amino acids strung together make proteins

12. Splicing Introns

13. Structure of DNA • Humans have 46 chromosomes • Chromosome 1 is the longest one, being 220 million base pairs long! • 99.7% of DNA between 2 people is the same…so we care about the other 0.3% that makes us different (~10 million nucleotides) • Only 3% of DNA is known to code for proteins • The rest (97%) is sometimes known as “junk DNA” because it doesn’t have any known function • The noncoding areas (introns) are interspersed between the coding areas (exons) • When the cell is reading the DNA, it cuts out the introns so it can link the exons together

14. VNTR • Within the introns (the noncoding portion) there are many areas that are known to have repeating sequences of several nucleotides (up to 10bp) • The number of repeating sequences is highly variable among people • Therefore, if in one particular intron, the sequence ATTG is repeated over and over, I may have 1000 copies, but you may have 15 copies, and the person next to you may have 345 copies. • These repeating units (variable number tandem repeats – VNTR) were discovered by Alec Jeffreys in 1985.

15. VNTR • By only looking at 1 set of repeating units, it is possible that 2 people may have the same number, but what if we look at 2 repeating areas… What are the chances 2 people would have the same number of repeats in those 2 areas? • And what if we used 10 repeating areas? • 16 markers (areas of known repeating units) are commonly used to make definitive conclusions

16. DNA Fingerprinting • On either side of the repeating areas are known nucleotide base sequences • Restriction endonucleases are special enzymes that can cut up DNA at specific sequences • When a restriction endonuclease is used on a person’s DNA that has many repeating units, that piece of DNA is bigger than a person who has only a few repeating units • This is done to target several of the areas where VNTRs are known to exist

17. Ultimately, the person’s DNA would be cut up into several smaller pieces and the length of each piece is unique to that person • Electrophoresis is a technique used to separate these pieces of different lengths

18. Electrophoresis

19. Electrophoresis • The smaller fragments travel faster, while the large fragments travel slowly • The pattern of the fragments is then unique to an individual

20. DNA Fingerprinting

21. DNA Fingerprinting Suspect 1 Suspect 2 Suspect 3 Suspect 4

22. DNA Fingerprinting

23. DNA Fingerprinting

25. Blood Samples at a Crime Scene Presumptive Tests A criminalist needs to be quite certain that actual blood was found before sending a sample to the laboratory for DNA testing Presumptive tests are done in the field to determine if a sample is probably blood, or maybe just a ketchup stain

26. Blood Samples at a Crime Scene Presumptive Tests A reddish brown droplet can be many things other than blood – so a presumptive test is useful to determine whether to collect the sample or not Kastle-Meyer color test (phenolphthalein mixed with hydrogen peroxide)– turns blood deep pink color (also reacts with potatoes and horseradish) Hemastix (green indicated blood) Luminol (capable of detecting bloodstains diluted up to 300,000 times!)(good for revealing patterns)

27. Blood Samples at a Crime Scene Confirmatory Tests Confirmatory tests confirm the presence of human blood – Precipitin test These are used in the laboratory before expensive DNA testing is done on a sample that doesn’t contain DNA