Daniel:Protocols/TBUgel: Difference between revisions

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(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 ...")
 
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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" | &nbsp;
| 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)