Daniel:Protocols/TBUgel: Difference between revisions
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=Urea Gel Quantification= | =Urea Gel Quantification= | ||
[[Daniel Jacobsen|Back to Main]] | |||
==Protocol 1== | ==Protocol 1== | ||
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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 | ||
===Example Gel Image Using Protocol 1=== | |||
[[Image:LambdaExo-030613.jpg|600px]] | |||
==Protocol 2== | ==Protocol 2== | ||
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5. Load samples and run gel for 20-25 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) | 6. Stain with 3 uL SYBR gold for at least 15 minutes (rec: 20-25) | ||
===Example Gel Image using Protocol 2=== | |||
[[Image:PrimerQuant_050813.jpg|600px]] | |||
==Invitrogen Low Mass Ladder== | ==Invitrogen Low Mass Ladder== |
Latest revision as of 18:58, 29 May 2013
Urea Gel Quantification[edit]
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]
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]
Invitrogen Low Mass Ladder[edit]
Invitrogen, Catalog #: 10068-013, $113.00
Infosheet: File:InvitrogenLowMassLadder.pdf