Purpose: To successfully isolate DNA from cheek cells and to prepare a PCR reaction for amplification of an Alu insert
Materials: 9% saline solution, micropipettes, tips, microcentrifuge, PCR tubes, agarose, 1XTAE, gel chambers and molds, load dye, racks, primer mix, master mix, H2O, positive control DNA, chelex
Procedure:
1. swish 10mL of saline solution in your mouth for 30 seconds
2. spit into cup and swirl to mic cells
3. label a 1.5mL microfuge tupe with PIN number
4. transfer 1mL to 1.5mL of the saline/cell suspension into the labled tube
5. spin suspension for 1 minute to pellet cells
6. pour supernatant off of the pelleted cells but leave 100 micro liters on top of cells
7. flick to resuspend cells
8. withdraw 50 micro liters of cell suspension and put in Chelex
9. place of heat block for 10 minutes
10. afterwards open tube to release pressure then centrifuge for 1 minute
11. label another tube with PIN and "DNA"
12. withdraw 50 micro liters of Chelex/DNA and put into new labeled tube. Note: do not take chelex beads
13. label tiny PCR tube with PIN
14. pipet 20 micro liters of master mix into PCR tube
15. add 20 micro liters of primer mix
16. add 10 micro liters of DNA
17. place reaction into the thermal cycler
18. afterward to a quick spin
19. 20 mL sample into new tube
20. add 4 mL of load dye
21. quick spin
22. load 20 mL in your well
23. record gel number and lane
Materials: 9% saline solution, micropipettes, tips, microcentrifuge, PCR tubes, agarose, 1XTAE, gel chambers and molds, load dye, racks, primer mix, master mix, H2O, positive control DNA, chelex
Procedure:
1. swish 10mL of saline solution in your mouth for 30 seconds
2. spit into cup and swirl to mic cells
3. label a 1.5mL microfuge tupe with PIN number
4. transfer 1mL to 1.5mL of the saline/cell suspension into the labled tube
5. spin suspension for 1 minute to pellet cells
6. pour supernatant off of the pelleted cells but leave 100 micro liters on top of cells
7. flick to resuspend cells
8. withdraw 50 micro liters of cell suspension and put in Chelex
9. place of heat block for 10 minutes
10. afterwards open tube to release pressure then centrifuge for 1 minute
11. label another tube with PIN and "DNA"
12. withdraw 50 micro liters of Chelex/DNA and put into new labeled tube. Note: do not take chelex beads
13. label tiny PCR tube with PIN
14. pipet 20 micro liters of master mix into PCR tube
15. add 20 micro liters of primer mix
16. add 10 micro liters of DNA
17. place reaction into the thermal cycler
18. afterward to a quick spin
19. 20 mL sample into new tube
20. add 4 mL of load dye
21. quick spin
22. load 20 mL in your well
23. record gel number and lane