Life Science 15: Concepts and IssuesLecture 10: Modern genetics and biotechnology: DNA fingerprinting2/9/12I. DNA fingerprinting overviewII. Individual differences in DNA- VNTRsIII. RFLP analysis and DNA fingerprintingIV. Difficulties and controversiesWhy are we interested in DNA fingerprinting?- real fingerprinting not left behind in many crime scenes- they are essentially uniqueDNA fingerprinting:- Uses:o Solve unsolvable crimeso Resolve paternity issueo Can prove innocence of people unjustly convictedo ImmigrationDNA is a long sequence of ‘letters’ that carry the information of how to build molecules for the body.A A G T C C A G G G G C T ________________________T T C A G G T C C C C G AFour ‘letters’: A,T,C,GHow much spelling difference is there between individuals?- almost none- 99.9% identical- different enough, though: 3 million different base pairs in that .1%On average, individuals differ in their DNA sequence at 0.1% of their bases: 3 million differences out of 3 billion base pairs.The fundamental question: We have DNA from several samples. How can we tell if the DNA came from the same person?- what sections of DNA do we want to look at?o High variable regionso Variable Number of Tandem Repeats (15-100 nucleotides long)VNTR1:- allele 1:o ___I___I___ < chromosomeo ^’ATCGATC’- allele 2:o ___I___I___I___ - allele 3:o ___I___I___I___I___ - allele 4:o ___I___I___I___I___I___ - allele 5:o ___I___I___I___I ___I___I___ - allele 6:o ___I___I___I___I___I___I___I___ What do individuals look like?VNTR1:- individual A:o allele 5: ___I___I___I___I ___I___I___ o allele 6: ___I___I___I___I___I___I___I___ Individuals have numerous VNTR loci:- Ind A: 10,13- Ind B: 7,8- Ind C: 13,11- For any VNTR locus- many (2-3 dozen or more) o Alleles exist within the population, but each individual can only have 2 of them.Comparing VNTR loci:- choose 6 features for which individuals vary a lot- compare DNA sample for those 6 featureso if they’re the same at all of them… the samples come from the same individualProducing a DNA fingerprint:1. Break-up DNA with restriction enzymes- Ind A:5: ___ I___I___I___I ___I___I ___ 6: ___ I___I___I___I___I___I___I ___ - Ind B:5: ___ I___I___I___I ___I___I ___4: ___ I___I___I___I___I ___ - Ind C:2: ___ I___I___I ___ 1: ___ I___I ___ (Pink denotes Restriction Fragment Lengths)At VNTR loci, a restriction enzyme cleaves the DNA into fragments of different lengths, depending on the number of repeated sequences present. The differences are RFLPs: Restriction Fragment Length Polymorphisms. 2. Run GelDNA Samples lined up at top A B CNegative ___ ___ ___ChargeFragments migrate down gel:Small: fastPositive Long: slowCharge- Gel from VNTR: A B CNegative Charge ______same allele > ___ _________PositiveChargeHuman Paternity Testing:- Analyzing one locus:Parents Children 1 2 3 4 5 6 A B C___ ___ ___ ___ ___ ___ _________ ______ ___ ___ ______ ___ ___ ___Which pair of people are the parents of each?- A: 5 and 2- B: 3 and 1- C: 4 and 6Difficulties and Controversies with DNA fingerprinting:1. Overlap of VNTRs2. How many VNTRs should we compare?3. How likely is a match?a. P (one band) x p (another)…b. Can juries understand this?4. are there sub-population differences?a. P= 1/10 * 1/10 * 1/10 * 1/10 = 1/10,000b. P= 1/10 * 1/10 * 1/1 * 1/1 = 1/100c. 2 loci, 10 alleles each5. Do humans make mistakes? YES.6. How sure do we want to be?a. What is a ‘reasonable’ level of certainty to warrant imprisoning someone?7. Should we keep DNA databases? a. Who would have access to them?b. People could be framed; unlawful search/seizureDespite universally accepted methods, DNA fingerprinting is not foolproof. We should never blindly draw