DNA Lab(Labs 4a, 4b, 4i,4j)
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
-15 mL Capped Tubes
- Tube Racks
- TRIS
- EDTA
- Disodium Salt
- 125 mL Bottle
- 100 mL Graduated Cylinder
- pH Paper
- Hydrochloric Acid
- Sodium Hydroxide
- Glass Rods
- 50 mL Beakers
- Salmon Sperm DNA
- 2 mL Pipette
- P-1000 Micropipette, and tips
- 95% Ethanol
- Permanent Marker Pens
- 1L Tripour Plastic Peaker
- 40X TAE Buffer Concentrate
- 600 mL Beakers
- Agarose
- 250 mL MEdia Bottle
- Microwave Oven
- Hot Hands Protector
- Horizontal Gel Box
- 65 degree Celcius water bath
Purpose:
Lab 4a
To make 10 ml of 5M NaCl, 100 ml of 10 mM Tris, 1mM EDTA.
Lab 4b
Spool DNA to extract it from the liquid.
Lab 4i
Pour the agarose gels into the Tris buffer.
Lab 4j
Find the size and movement of the DNA.
Procedure: (4a and 4b)
1. Make molarity calculations.
[Given: 1 mole = 6.02 x 10^23 ; 5 moles = 5 (6.02 x 10^23 particles) / 1 Litre ; (Molarity) (Volume) (Formula weight) = g substance needed]
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 Solution by combining Solutions Two and Three
3. Dilute DNA with TE solution in a flask.
4. Add 2.92 grams of NaCl.
5. Add 4 mL of alcohol by trickling it down the side of the flask.
6. Spool DNA
7. Put the spooled DNA into a new tube and add 2 mL of fresh TE solution.
Procedure: (4i)
1. Prepare 100 mL solution of 0.8% agarose in 1X TAE buffer solution.
2. Weigh out the required mass of powdered agarose in a weigh boat. Add it to a 250-mL media bottle.
3. Measure out enough TAE buffer to prepare a total of 100 mL of agarose and buffer mixed together. Swirl to mix.
4. Cap the media bottle and swirl the flask to suspend the agarose in the buffer.
5. To dissolve the agarose, microwave for 4 minutes at 50% power. Wait for the solution to boil.
6. Place the hot dissolved agarose solution on a fireproof lab tabletop and let cool to 65 degrees Celsius before pouring into a gel tray.
7. Place a six-well comb into the notches at the end of the gel tray. This will create the necessary wells for the next experiment.
8. Place the gel (on the gel tray) into a gel box.
9. Pour 1X TAE buffer into the gel box and completely submerge the gel.
10. Gently pull the comb out of the gel and make sure the wells are not broken or cracked.
- Tabletop Milligram Balance
- 7.6 x 7.6 cm Weigh Paper
- 3.5 x 3.5 Weigh Boat
- Lab Scoops
- Sodium Chloride
-15 mL Capped Tubes
- Tube Racks
- TRIS
- EDTA
- Disodium Salt
- 125 mL Bottle
- 100 mL Graduated Cylinder
- pH Paper
- Hydrochloric Acid
- Sodium Hydroxide
- Glass Rods
- 50 mL Beakers
- Salmon Sperm DNA
- 2 mL Pipette
- P-1000 Micropipette, and tips
- 95% Ethanol
- Permanent Marker Pens
- 1L Tripour Plastic Peaker
- 40X TAE Buffer Concentrate
- 600 mL Beakers
- Agarose
- 250 mL MEdia Bottle
- Microwave Oven
- Hot Hands Protector
- Horizontal Gel Box
- 65 degree Celcius water bath
Purpose:
Lab 4a
To make 10 ml of 5M NaCl, 100 ml of 10 mM Tris, 1mM EDTA.
Lab 4b
Spool DNA to extract it from the liquid.
Lab 4i
Pour the agarose gels into the Tris buffer.
Lab 4j
Find the size and movement of the DNA.
Procedure: (4a and 4b)
1. Make molarity calculations.
[Given: 1 mole = 6.02 x 10^23 ; 5 moles = 5 (6.02 x 10^23 particles) / 1 Litre ; (Molarity) (Volume) (Formula weight) = g substance needed]
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 Solution by combining Solutions Two and Three
3. Dilute DNA with TE solution in a flask.
4. Add 2.92 grams of NaCl.
5. Add 4 mL of alcohol by trickling it down the side of the flask.
6. Spool DNA
7. Put the spooled DNA into a new tube and add 2 mL of fresh TE solution.
Procedure: (4i)
1. Prepare 100 mL solution of 0.8% agarose in 1X TAE buffer solution.
2. Weigh out the required mass of powdered agarose in a weigh boat. Add it to a 250-mL media bottle.
3. Measure out enough TAE buffer to prepare a total of 100 mL of agarose and buffer mixed together. Swirl to mix.
4. Cap the media bottle and swirl the flask to suspend the agarose in the buffer.
5. To dissolve the agarose, microwave for 4 minutes at 50% power. Wait for the solution to boil.
6. Place the hot dissolved agarose solution on a fireproof lab tabletop and let cool to 65 degrees Celsius before pouring into a gel tray.
7. Place a six-well comb into the notches at the end of the gel tray. This will create the necessary wells for the next experiment.
8. Place the gel (on the gel tray) into a gel box.
9. Pour 1X TAE buffer into the gel box and completely submerge the gel.
10. Gently pull the comb out of the gel and make sure the wells are not broken or cracked.