Materials:
- Analytical Balance - Tabletop Milligram Balance - 7.6 x 7.6 cm Weigh Paper - 3.5 x 3.5 Weigh Boat - Lab Scoops - Sodium Chloride - 2 mL Pipette - P-1000 Micropipette, and tips - 95% Ethanol - Permanent Marker Pens - 1L Tripour Plastic Beaker - Tube Racks - TRIS - EDTA - Horizontal Gel Box |
- 40X TAE Buffer Concentrate
- 600 mL Beakers - Agarose - 250 mL Media Bottle -15 mL Capped Tubes - Disodium Salt - 125 mL Bottle - 100 mL Graduated Cylinder - pH Paper - Hydrochloric Acid - Sodium Hydroxide - Glass Rods - 50 mL Beakers - Microwave Oven - Hot Hands Protector - 65 degree Celcius water bath |
1. Make molarity calculations.
Solution One: NaCl: 5M x 0.010 L x 58.44 g/mole = 2.92 grams
Solution Two: TRIS: 0.01 M x 0.1 L x 372.24 g/mole = 0.158 grams
Solution Three: 0.001 M x 0.1 L x 372.24 g/mole = 0.037 grams
2. Create TE soulution by combining Solutions 2 and 3
3. Dilute DNA with TE solution in a flask.
4. Add 2.92 grams of NaCl.
5. Add 4 mL of alcohol
6. Spool DNA
7. Put the spooled DNA into a tube and add 2 mL of TE solution. 1x TAE Buffer
Solution One: NaCl: 5M x 0.010 L x 58.44 g/mole = 2.92 grams
Solution Two: TRIS: 0.01 M x 0.1 L x 372.24 g/mole = 0.158 grams
Solution Three: 0.001 M x 0.1 L x 372.24 g/mole = 0.037 grams
2. Create TE soulution by combining Solutions 2 and 3
3. Dilute DNA with TE solution in a flask.
4. Add 2.92 grams of NaCl.
5. Add 4 mL of alcohol
6. Spool DNA
7. Put the spooled DNA into a tube and add 2 mL of TE solution. 1x TAE Buffer
- Make 500ml 1x TAE Buffer and 50ml 8% agarose
- Add agarose to 100ml 1x TAE in 250ml flask
- Heat and swirl to boil and dissolve
- Let flask cool until you can touch it
- Pour in gel mold and let cool until solidified
- Remove tape from gel, place in gel tank
- Pour TAE over gel until covered, gently remove combs
- Prepare Samples: 20 ml DNA + 4ml 6x loading dye; spin for 2 seconds together in mini centrifuge
- Load samples onto gel
- Cover gel tank, plug into power supply
- Run at 110V for 45 minutes
- Stain several hours with EtBr
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