Urea Gel Quantification[edit]
Back to Main
Protocol 1[edit]
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
Example Gel Image Using Protocol 1[edit]
File:LambdaExo-030613.jpg
Protocol 2[edit]
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)
Example Gel Image using Protocol 2[edit]
File:PrimerQuant 050813.jpg
Invitrogen Low Mass Ladder[edit]
Invitrogen, Catalog #: 10068-013, $113.00
Infosheet: File:InvitrogenLowMassLadder.pdf
File:LowMassLadder Invitrogen.png