1 DNA Polymorphisms: DNA markers a useful tool in biotechnology Any section of DNA that varies among individuals in a population, “many forms”. Examples.

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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 SNPs Single nucleotide polymorphisms: regions of DNA where one base pair is different. Occur evenly spread over all the DNA. 1/ 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 SNP

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

RFLPs 5 DNA marker may be associated with genetic condition.

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

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.

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 STRs in forensics Locus vWA 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 THE 13 CODIS STRs and “probabilities of Identity” STR African-AmericanU.S. Caucasian D3S vWA FGA TH TPOX CSF1PO D5S D13S D7S D8S D21S D18S D16S

12 Repeat #CaucasianHispanicAfrican American Asian Allele frequencies for D1S80 among US population groups Chance of a white person being heterozygous for alleles 19 and 20: 2 x x (One in 9,259)

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 RAPD: using PCR to find polymorphisms-2