1. DNA Fingerprinting Class instructions

2. In this lesson your students will learn DNA fingerprinting uses STR repeats STRs are repeats of short sequences on DNA Number of STR copies varies between individuals Primers must match only one region in the template DNA Primers must match each end of the target Complementary base pairing is critical for DNA fingerprinting How gender is determined from DNA

3. The devil is in the detail! The 5’ prime and 3’ prime ends of the bases must be round the right way! IMPORTANT: Do not take bases apart!!!

4. Correct base pairing is critical! Green (Guanine) pairs with yellow (Cytosine) Blue (Adenine) pairs with orange (Thymine)

5. Using primers to determine PCR product size Primer set 1 Primer set 2

6. Lesson Part One – STR identification Group 1 and 2 will work out the number of repeats of one STR located on chromosome 16 (see manual for full details). At this loci people have between 5-16 copies of the STR giving differenet sized products. The length of the PCR product allows you to count how many copies of the STR repeat are present.

7. Group 1 – STR analysis 1Make the double stranded DNA template using bases with black sugars. Remember to get the 5' to 3' orientation correct! 2Denature the two strands by separating them. 3Anneal single stranded primers to the complementary bases - one primer per strand. 4Work out the length of the final PCR product. How many copies of the STR repeat do you have? 5 Optional extension – use parts from PCR Puzzle to make a model of the double stranded PCR product using bases with red sugars.

8. DNA Sequence Denature Anneal primers PCR product

9. Group 2 – STR analysis 1Make the double stranded DNA template using bases with black sugars: 2Denature the two strands by separating them. 3Anneal single stranded primers to the complementary bases - one primer per strand. 4Work out the length of the final PCR product. How many copies of the STR repeat do you have? 5Optional extension – use parts from PCR Puzzle to make a model of the double stranded PCR product using bases with red sugars.

10. DNA Sequence Denature Anneal primers PCR product

11. Lesson Part Two - Gender determination Group 3 and 4 look at gender X chromosome has a short deletion (see manual for details) Y chromosome does not have the deletion Male & female give different size PCR products You can visualise these on an electrophoresis gel

12. How to determine gender from these results A female will have XX chromosomes so you get one band on a gel as both give the same size PCR product A male will have XY chromosomes so you get two bands of different sizes on a gel Think about how male and female DNA will appear on an electrophoresis gel

13. Group 3 – Gender chromosome 1Make the double stranded DNA template using bases with black sugars. 2Denature the template by separating the strands. 3Anneal single stranded primers to the complementary bases - one primer per strand. 4Work out length of the final PCR product. 5Optional extension – use parts from PCR Puzzle to make a model of the double stranded PCR product using bases with red sugars.

14. DNA Sequence Denature Anneal primers PCR product

15. Group 4 – Gender chromosome 1Make the double stranded DNA template using bases with black sugars. 2Denature the template 3Anneal single stranded primers to the complementary bases - one primer per strand. 4Work out length of the final PCR product. 5Optional extension – use parts from PCR Puzzle to make a model of the double stranded PCR product using bases with red sugars.

16. DNA Sequence Denature Anneal primers PCR product