Daniel:Protocols/TBUgel: Difference between revisions
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>Djacobse (Created page with "=Urea Gel Quantification= ==Protocol 1== 1. Prerun gel for at least 20 minutes at 250 Volts 2. Prepare samples, typical setup: {| class="wikitable" <hiddentext>generated ...") |
>Djacobse No edit summary |
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5. Load samples onto gel and run for 20-25 minutes at 250V | 5. Load samples onto gel and run for 20-25 minutes at 250V | ||
6. Stain with 3 uL SYBR gold for at least 15 minutes | 6. Stain with 3 uL SYBR gold for at least 15 minutes | ||
==Protocol 2== | |||
1. Preheat TBE buffer by incubating it in 60C incubator | |||
Microwave to do it quickly, but make sure not to get the buffer too hot (ideal: 45-60C) | |||
2. Prerun gel in heated buffer at 250V for ~15-20 minutes | |||
3. Prepare samples according to following table: | |||
{| class="wikitable" <hiddentext>generated with [[:de:Wikipedia:Helferlein/VBA-Macro for EXCEL tableconversion]] V1.8</hiddentext> | |||
|- style="font-size:12pt" align="center" | |||
| align="center" width="65" height="35" | | |||
| width="65" | uL sample | |||
| width="65" | uL ladder | |||
| width="65" | uL TBE | |||
| width="65" | uL 2X urea buffer | |||
| width="65" | uL loaded | |||
|- style="font-size:12pt" align="center" | |||
| height="15" | samples | |||
| align="center" | 0.5 | |||
| align="center" | 0 | |||
| align="center" | 5.5 | |||
| align="center" | 6 | |||
| align="center" | 12 | |||
|- style="font-size:12pt" align="center" | |||
| height="15" | ladder 1 | |||
| align="center" | 0 | |||
| align="center" | 0.5 | |||
| align="center" | 5.5 | |||
| align="center" | 6 | |||
| align="center" | 12 | |||
|- style="font-size:12pt" align="center" | |||
| height="15" | ladder 2 | |||
| align="center" | 0 | |||
| align="center" | 1 | |||
| align="center" | 5 | |||
| align="center" | 6 | |||
| align="center" | 12 | |||
|} | |||
4. Denature samples for 8 minutes at 65C | |||
5. Load samples and run gel for 20-25 minutes at 65C | |||
6. Stain with 3 uL SYBR gold for at least 15 minutes (rec: 20-25) |
Revision as of 17:03, 29 April 2013
Urea Gel Quantification
Protocol 1
1. Prerun gel for at least 20 minutes at 250 Volts 2. Prepare samples, typical setup:
uL sample | uL ladder | uL TBE | uL 2X urea buffer | uL loaded | |
samples | 0.5 | 0 | 5.5 | 6 | 12 |
ladder 1 | 0 | 0.5 | 5.5 | 6 | 12 |
ladder 2 | 0 | 1 | 5 | 6 | 12 |
3. Denature samples for 8 minutes at 65C 4. Snap cool using freezer box 5. Load samples onto gel and run for 20-25 minutes at 250V 6. Stain with 3 uL SYBR gold for at least 15 minutes
Protocol 2
1. Preheat TBE buffer by incubating it in 60C incubator Microwave to do it quickly, but make sure not to get the buffer too hot (ideal: 45-60C) 2. Prerun gel in heated buffer at 250V for ~15-20 minutes 3. Prepare samples according to following table:
uL sample | uL ladder | uL TBE | uL 2X urea buffer | uL loaded | |
samples | 0.5 | 0 | 5.5 | 6 | 12 |
ladder 1 | 0 | 0.5 | 5.5 | 6 | 12 |
ladder 2 | 0 | 1 | 5 | 6 | 12 |
4. Denature samples for 8 minutes at 65C 5. Load samples and run gel for 20-25 minutes at 65C 6. Stain with 3 uL SYBR gold for at least 15 minutes (rec: 20-25)