Purpose
1. Successfully isolate DNA from cheek cells
2. Prepare a PCR reaction for amplification of an ALU insert
2. Prepare a PCR reaction for amplification of an ALU insert
Materials
9% saline solution
micro-pipettes, tips
waste container
micro-centrifuge
micro-centrifuge tubes
PCR tubes
agarose
1XTAE
gel chambers + molds
Load dye
racks
primer mix
master mix
H2O
+ control DNA
Chelex
micro-pipettes, tips
waste container
micro-centrifuge
micro-centrifuge tubes
PCR tubes
agarose
1XTAE
gel chambers + molds
Load dye
racks
primer mix
master mix
H2O
+ control DNA
Chelex
DNA Procedure
1. Swish salt water in mouth for 30 seconds, then spit into cup
2. Transfer to microfuge tube and spin
3.pour of supernatent and leave 100ul and re-suspend
4. add 50ul of cell suspension to Chelex
5. heat for 10 minutes
6 Insert into new tube
2. Transfer to microfuge tube and spin
3.pour of supernatent and leave 100ul and re-suspend
4. add 50ul of cell suspension to Chelex
5. heat for 10 minutes
6 Insert into new tube
PCR procedure
1. Put 20 ul of mastermix into pcr tube
2. add 10ul of dna
3. set up 2 tubes of control
4. place tubes into thermal cycler
2. add 10ul of dna
3. set up 2 tubes of control
4. place tubes into thermal cycler
Gel making
1.50 ml 1xTAE + 1g agarose
2.heat untill dissolved
3.pour in mold when cool
2.heat untill dissolved
3.pour in mold when cool
GEl electrophoresis
1. Spin DNA
2.take 20ul of Dna into 1.5 ml tube
3.put 4ul of loading dye into tube
4.Spin
5. Put 20ul of mixture into gel
6. run gels
2.take 20ul of Dna into 1.5 ml tube
3.put 4ul of loading dye into tube
4.Spin
5. Put 20ul of mixture into gel
6. run gels