What is DNA Fingerprinting?
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1 What is DNA Fingerprinting? A collection of techniques used to determine the genetic identity of an organism based on its unique DNA patterns Can use trace amounts of hair, skin, body fluids Based on the fact that every individual s DNA sequence is unique (except identical twins) Based on known DNA polymorphisms, usually in the non-protein coding regions of the genome
2 SNPs Single Nucleotide Polymorphisms SNPs are produced when individuals in a population have different nucleotides (A,T,C or G) at specific locations. Each sequence change represents a potential allele SNPs are differences in specific sequence locations among individuals in a population.
3 Example of SNP use Recent genome wide study of SNPs to identify genes under positive selection in human populations living at high altitudes Compared SNPs in Tibetan genomes (live at high altitudes) with Han Chinese and Japanese genomes (both do not live at high altitudes) Study identified 1 gene known to affect regulation of RBC production 2 other genes linked to adult and fetal Hb showed a high correlation. Source: Science, Vol 329, 2 July 2010, p.72, Simonson et.al.
4 RFLPs Restriction Fragment Length Polymorphisms Utilizes restriction enzyme analysis of total genomic DNA to identify regions with SNPs SNPs produce RFLPs
5 SNPs & RFLP (1) A change in a specific nucleotide at a specific location produces a change in the DNA sequence DNA Sequence changes alter Restriction Enzyme recognition sites Add a restriction site Delete a restriction site
6 Person 1 Fragment Fragment 1 Fragment 2 Fragment 4 3 RE1 RE2 RE3 RE4 RE5 Person 2 Fragment 1 Fragment 2 Fragment 3 RE = Restriction Enzyme Site In this example, RE3 no longer exists due to a sequence change in Person 2 Person 1 has a different set of restriction fragments than Person 2 a RFLP
7 SNPs & RFLPs (2) A change in restriction sites will change the resulting pattern of Restriction Fragments RFP: Restriction Fragment Pattern In or across populations, different nucleotide changes, SNPs, occur in different individuals: In or across populations, SNPs produce RFP variations, these RFP variations are termed RFLPs
8 SNPs & HAPLOTYPES Populations: Individuals with a common pool of genetic variations. (SNPs, Indels, rearrangements, etc). HAPLOTYPE: a genotype with a single or a group of variations. HAPLOTYPES can be defined by the presence of a single variation (1 SNP) or a collection of variations.
9 Steps in RFLP analysis: 1. Collect sample (need a fair amount) 2. Purify DNA from the sample 3. Digest with restriction enzymes 4. Run on agarose gel 5. Transfer to nitrocellulose 6. Hybridize with labeled probe 7. Detect bands by autoradiography
10 DNA Fingerprint from Identical Twins No RFLPs present
11 DNA Fingerprints from a Sexual Assault Case Which Suspect can be Ruled Out?
12 Which bands implicate the defendant? Why?
13 Paternity Testing: DNA Evidence
14 PCR (Polymerase Chain Reaction) BASICS OF PCR (Animation)
15 PCR of STRs (Short Tandem Repeats) Another Tool Used for DNA Fingerprinting Usually only 3-4nucleotides in each repeat Advantages Can be done using very small amounts of DNA DNA can be very old or partially degraded Routinely used in forensic analysis FBI has chosen 13 STRs for CODIS database Combined DNA Index System Odds of matching someone else at all 13 STRs loci is greater than one in a billion Main problem is sample contamination
16 STRs
17 Applications for DNA Fingerprinting Forensics (exclude or implicate a suspect) Exoneration of falsely accused (Innocence Project) Paternity cases Evolutionary studies Want to Know More: Genetic Justice
18 Haplotypes & Genotypes for Alu insertion at PV92 Locus Alu insert = 300bp Haplotypes PV92 locus with NO Alu insertion (400bp) PV92 locus WITH Alu insertion (700bp) Genotypes Homozygous NO Alu insertion (Alu -/-) Heterozygous for Alu insertion (Alu +/-) Homozygous for Alu insertion (Alu +/+)
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