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| META TOPICPARENT |
name="ProtocolList" |
Reagent and Buffer Recipes |
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General calculation resources |
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50x TAE
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< | Prepare by filling bottle with 900 ml of ddH20 and adding the above chemicals. Adjust volume to 1 L with ddH20. |
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> | Prepare by filling bottle with 700 ml of ddH20 and adding the above chemicals. Adjust volume to 1 L with ddH20. |
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Working concentration is 1x, so measure 400ml of 50x solution in graduated cylinder and then pour into 20 L carboy and fill to 20L with ddH20; if filling a 10 L carboy use 200 ml of stock. Unused or left over acetic acid must be disposed of in a chemical waste bottle located in the fume hoods. To avoid having left over Acetic acid use serological pipettes to measure out the Acetic acid (use 1 50 ml and 1 10 ml). |
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< | 5x TBE Tris•Boric Acid•EDTA |
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> | 5x TBE Tris•Boric Acid•EDTA |
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| 54 g |
Tris base |
| 27.5g |
Boric Acid |
| 20 mL |
0.5 M EDTA (pH 8.0) |
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| | RNase A, 5 mg/ml
| Dissolve 100 mg of RNase A in 20 ml of 0.05% glacial acetic acid, and transfer to a 50-ml conical tube |
| Place the tube in a boiling-water bath for 15 minutes |
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| Cool the solution, and neutralize by adding 120 μl of 1 M Tris (pH 8.0) |
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| Cool the solution, and neutralize by adding 120 μl of 1 M Tris (pH 8.0) |
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| Distribute 1 ml aliquote into 1.5 ml MFT, and store at -20 C |
rNTP, 100 mM |
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- Dissolve 1 g of desired NTP in 10 ml ddH2O:
- ATP - Adenosine 5′-triphosphate disodium salt (MW 551.14)
- GTP - Guanosine 5′-triphosphate sodium salt hydrate (MW 523.18)
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- pH to 8.0 with 1 M NaOH. It takes approximately 1.0–1.5 ml.
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- pH to 8.0 with 1 M NaOH. It takes approximately 1.0–1.5 ml.
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- Add ddH2O to final volume of:
- GTP - 18.14 ml
- GTP - 19.11 ml
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- Store at –20°C in 1.0–1.5 ml aliquots.
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- Store at –20°C in 1.0–1.5 ml aliquots.
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< | HEPES•NaOH, 1M, pH 7.0 |
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> | HEPES•NaOH, 1M, pH 7.0 |
| | pH buffer with less temperature dependence than Tris.
To make 100 ml: |
