1. 1 DNA Polymorphisms: DNA markers a useful tool in biotechnology Any section of DNA that varies among individuals in a population, “many forms”. Examples include: SNPs, RFLPs, STRPs, and AFLPs; –RFLPs include VNTRs and STRPs –microsatellites (STRs) = SSLPs = STRPs = SSRs Useful for finding, mapping genes involved in disease, and –Individual identification, epidemiology, anthropology, population/ecology studies, taxonomy.

2. 2 SNPs Single nucleotide polymorphisms: regions of DNA where one base pair is different. Occur evenly spread over all the DNA. 1/ 1000-3000 bp Detected by sequencing. If SNP occurs in a restriction enzyme site, it generates an RFLP. Could be in coding or non-coding regions. Over 300,000 human SNPs known and are being mapped.

3. 3 SNP

4. 4 RFLPs Restriction fragment length polymorphism. –Any difference in the DNA that results in a restriction enzyme producing a different sized piece. Mutation at a restriction site prevents recognition & cutting. –Results in one band of larger DNA instead of 2 smaller ones. Different numbers of repeats between restriction recognition sites also generates an RFLP

5. RFLPs 5 DNA marker may be associated with genetic condition. www.ncbi.nlm.nih.gov/.../doc/TechRFLP.shtml

6. 6 VNTRs and STRPs as RFLPs: Minisatellites and Microsatellites These are RFLPs because they are defined by or visible following restriction enzyme cuts. –Variable Number Tandem Repeats Groups (10-100) of nucleotides repeated 2 – 100 times (depending on individual and locus). Restriction sites on both sides of repeated DNA The more repeats, the longer the fragment. –Simple Tandem Repeat Polymorphisms Shorter, 2-9 nucleotides repeated Small enough number for PCR amplification Also called STRs, SSLPs, etc.

7. 7 Use of VNTRs Restriction sites are on either side; fragment length depends on number of repeats in between sites.

8. 8 STRPs Primers for both sides of repeated region allow PCR amplification of DNA; generates PCR products that differ in length depending on number of repeats. Becoming the standard method for DNA testing in forensics labs. Cheaper, easier, more sensitive.

9. Hardy-Weinberg meets Gil Grissom Simple tandem repeats –13 have been chosen for use in forensic work –The 13 independently assort, meaning they are on different chromosomes or far apart on the same. Product law can be used –Each of the 13 have a number of different alleles Alleles differ by number of repeats –Repeats vary from 3 to 5. vWA is a tetranucleotide. –Allele frequencies: p1, p2, etc. for each allele –Humans are diploid, have 2 alleles for each locus 9

10. 10 STRs in forensics Locus vWA 14 0.081 15 0.107 15.2 0.179 16 0.306 17 0.192 18 0.089 19 0.047 Alleles in different ethnic and racial groups examined, used as database. Panel of 13 different STRs are used. Because the odds of a particular combination of the 13 is product of the frequencies, numbers like 1 in 10 billion can be generated. Hardy-Weinberg: 2pq Bandfrequency

11. 11 THE 13 CODIS STRs and “probabilities of Identity” STR African-AmericanU.S. Caucasian D3S13580.1020.078 vWA0.0580.065 FGA0.0350.036 TH010.1020.094 TPOX0.0810.211 CSF1PO0.0700.122 D5S8180.0970.140 D13S3170.1310.074 D7S8200.0810.061 D8S11790.0750.067 D21S110.0330.045 D18S510.0280.030 D16S5390.0660.103 http://expertpages.com/news/dna.htm

12. 12 Repeat #CaucasianHispanicAfrican American Asian 140000 1500.00100 160.0010.0100.0020.034 170.0020.0090.0280.025 180.2370.2240.0730.152 190.0030.0050.0030.022 200.0180.0130.0320.007 210.0210.0280.1150.034 220.0380.0240.0810.017 Allele frequencies for D1S80 among US population groups Chance of a white person being heterozygous for alleles 19 and 20: 2 x 0.003 x 0.018 (One in 9,259)

13. 13 RAPD: using PCR to find polymorphisms “Random amplified polymorphic DNA” Screen DNA from individuals by doing PCR with random short primers. (about 8-12 nucleotides) By random chance, primers will amplify many different sections of DNA. Look for bands on gel that are not present in each individual tested. avery.rutgers.edu/.../ archives/onions/rapd.html

14. 14 RAPD: using PCR to find polymorphisms-2