#Gel Protocol
This is still a work in progress
I'm running and making all gels with 1x SB Buffer
Don't pour bioling hot gel into gel rig. Needs to cool a bit. You can use a bucket of ice to speed this up.
##required reagents
- Ethidium Bromide
- SB Buffer
- agarose
- dna ladder
- loading dye
###20X SB Buffer
- 45g boric acid
- 8g NaOH
- 1L H2O
- to make 1X working buffer
- 1 liter: 50ml 20x into 950ml water
- 5 liters: 250ml 20x into 4.75 liters water
##Quick and dirty check for high molecular weight dna
Use this if you just want to check quality of extracted DNA.
1.5% gel, 200V, 28 min
- note, you could probably use a 1% gel for even less time. I just haven't tested this.
Making small gel:
- 1.1 g agarose into 70 ml 1x SB Buffer
- 3ul etbr after gel is heated, but before it solidifies.
- 2ul dna, 2ul dye.
- currently using xylene cyanol 6x
- Load between 3-4 ul dye dna combo
- Can reuse gel 3-4 times. Add 1.5 ul etbr to melted gel before each run
- Ladder:
- dna ladder : 1ul
- water: 5ul
- combine 2 ul diluted ladder with 2 ul dye
2% gel, 110V, 1.5 hours
Dilute 100 bp ladder 1:8
- 4X recipe for diluted ladder
- 8.5 ul water
- 1.5 ul ladder
- 2 ul 6X loading dye
Load 3 ul ladder
Load 3 ul DNA+dye mix, .5 ul dye, 2.5 ul dna