1. DNA Laboratory Building a DNA Model DNA Fingerprinting

2. DNA Fingerprinting Detects differences between individuals at the DNA levelDetects differences between individuals at the DNA level Uses restriction enzymes that cut DNA at specific sequencesUses restriction enzymes that cut DNA at specific sequences Differences in position of restriction enzyme cuts yield different DNA fragment sizesDifferences in position of restriction enzyme cuts yield different DNA fragment sizes RFLP = restriction fragment length polymorphismRFLP = restriction fragment length polymorphism

3. Loss of an EcoRI Recognition Site Produces Different Numbers of DNA Fragments

4. Gel Electrophoresis Direction of Migration Larger fragments move more slowly; smaller fragments move more rapidly

5. Pouring the agarose Submerging the gel Loading the gel

6. Using the Micropipettor  Set volume to 35 ul  Attach a clean tip  Push plunger to first stop  Insert tip into liquid  Release plunger SLOWLY  Position over well in gel  Push plunger SLOWLY to second stop to fill well  Discard tip in orange bag

7. DNA RNA Protein Model will show Replication, Transcription and Translation transcription translation replication (S phase) (G1 and G2 phases) phases)

8. Completed Model One DNA double helix, two chains of 9 nucleotides eachOne DNA double helix, two chains of 9 nucleotides each –one chain can have any sequence –second chain must be complementary One chain of mRNA, 9 nucleotides longOne chain of mRNA, 9 nucleotides long –must be complementary to one of the two DNA chains (DNA template is 3’  5’, mRNA is 5’  3’) Three tRNAs with appropriate amino acids attachedThree tRNAs with appropriate amino acids attached –tRNA anticodons must match mRNA codons

9. Structure of a Nucleotide Components Phosphate Sugar Base Phosphate at 5’ position 3’ position

10. Link the phosphate of one nucleotide to the sugar of the next nucleotide to produce a DNA chain.

11. Replication Pair A with T and G with C to produce the second chain. One chain is 5’  3’ Other chain is 3’  5’ 5’ 3’ 5’

12. 3’ DNA polymerase adds nucleotides to the growing chain in a 5’  3’ direction. 3’ 5’

13. 3’ 5’ 3’ Your completed model will have two chains, each with 9 nucleotides.

14. Transcription One DNA chain is used as a template. Template is read 3’  5’ RNA is produced 5’  3’ Base in DNA Base in RNA AU TA GC CG 5’ 3’ 5’

15. RNA polymerase adds nucleotides to the growing chain in a 5’  3’ direction. 5’ 3’ 5’

16. 3’ 5’ 3’ Your completed messenger RNA will have 9 nucleotides.

17. Three Types of RNA DNA rRNAmRNA tRNA transcription Joins with proteins to form ribosome Specifies order of amino acids Carries amino acids

18. Structure of transfer RNA anticodon amino acid attachment site mRNA Anticodon on tRNA recognizes codon on mRNA to bring amino acid into place.

19. 5’3’ 5’ 3’ codon Each group of 3 nucleotides on mRNA is a codon. Codons are read from 5’  3’. tRNA anticodons, from 3’  5’ match the codons. anticodon

20. Read the CODON from 5’  3’ to determine the amino acid carried by the tRNA. Serine

21. Your completed model will have 3 tRNAs, one for each codon in mRNA. Show the name of the amino acid carried by each tRNA. Serine

22. CLEAN UP! After pouring gel, fill flask with water and return to bathAfter pouring gel, fill flask with water and return to bath Put used tips in the orange bagsPut used tips in the orange bags Rinse and dry the gel tray, comb and rubber damsRinse and dry the gel tray, comb and rubber dams Leave buffer in electrophoresis chamberLeave buffer in electrophoresis chamber After analysis, discard staining sheets and agarose gels into trashAfter analysis, discard staining sheets and agarose gels into trash Put all paper clippings into trashPut all paper clippings into trash Make sure the sinks stay cleanMake sure the sinks stay clean

23. Worksheet Attach your completed DNA model inside a legal-sized folderAttach your completed DNA model inside a legal-sized folder Complete pages 91-92 from lab manualComplete pages 91-92 from lab manual Be sure your instructor checks both of these items before you leave the labBe sure your instructor checks both of these items before you leave the lab