1. The Case of the Missing Strawberries: RFLP Analysis Heidi Sleister Assoc. Professor of Biology Drake University 1344 27 th Street Des Moines, Iowa 50311 Heidi.Sleister@Drake.edu Microsoft Clipart picture of strawberries

2. Problem: – Strawberries are missing from the school garden. – A search led to 5 people with strawberries. – Do any of these strawberries match the strawberries from the garden? Case Background Heidi Sleister

3. Chromosomes are made of DNA and are found in the nucleus of a cell Insert a picture which shows the relationship of DNA, chromosomes, and a cell. Heidi Sleister

4. Insert a picture which shows the structure of DNA (double helix, paired nitrogenous bases (G-C, A-T)). DNA has a double-stranded helical structure Heidi Sleister

5. Experimental Strategy: – Isolate DNA from strawberries – Use RFLP analysis to compare garden strawberry plant DNA to the “suspect” strawberries. Solving the Case Heidi Sleister

6. Disease diagnosis Genetic mapping Forensic science Genetically modified organisms Paternity testing Personal identification Plant breeding Characterization of genetic diversity Species identification Applications of RFLP Analysis Heidi Sleister

7. Restriction endonucleases cut specific sequences of DNA and produce DNA fragments ACCATAAGAATTCAATCC TGGTATTCTTAAGTTAGG EcoRI restriction site TGGTATTCTTAA ACCATAAGAATTCAATCC GTTAGG Fragment 1Fragment 2 Heidi Sleister

8. Different sizes of DNA fragments can be distinguished by gel electrophoresis Insert picture of gel electrophoresis chamber and gel. Heidi Sleister

9. Restriction fragment length polymorphisms (RFLPs) -------GAATTC------------------- -------CTTAAG------------------- -------GACTTC------------------- -------CTGAAG------------------- EcoRI site No EcoRI site Allele 1 Allele 2 Fragment A1-1Fragment A1-2 Fragment A2-1 Marker Allele 1Allele 2 Fragment A2-1 Fragment A1-2 Fragment A1-1 Allele 1 and Allele 2 differ by only a single basepair which is located within the EcoRI restriction site. Gel electrophoresis of allele 1 and allele 2 DNAs digested with EcoRI. Heidi Sleister