Daniel:Protocols/TBUgel
Jump to navigation
Jump to search
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)
Example Gel Image using Protocol 2
Invitrogen Low Mass Ladder
Invitrogen, Catalog #: 10068-013, $113.00
Infosheet: File:InvitrogenLowMassLadder.pdf