Toronto/Lab Protocols/Quantitation
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#Take the extracted DNA and run a gel to find out the concentration (per μL) of your DNA. | #Take the extracted DNA and run a gel to find out the concentration (per μL) of your DNA. | ||
| - | + | *1 μL plasmid + 4 μL ddH2O + 1 μL Loading Dye | |
| - | * | + | * * Gel Lane Fragment |
| - | + | ||
1 HindIII Ladder (5 μL) | 1 HindIII Ladder (5 μL) | ||
2 HindIII Ladder (2 μL) | 2 HindIII Ladder (2 μL) | ||
Revision as of 03:47, 27 October 2007
Quantitation
- Take the extracted DNA and run a gel to find out the concentration (per μL) of your DNA.
- 1 μL plasmid + 4 μL ddH2O + 1 μL Loading Dye
* * Gel Lane Fragment
1 HindIII Ladder (5 μL)
2 HindIII Ladder (2 μL)
3 HindIII Ladder (1 μL)
4 Blank
5 Plasmid (All of it)
6 Blank
7 Insert (All of it)
8 Blank
9 1 Kbp Ladder (3 μL)
- Ensure that the plasmid and inserts are at the correct lengths using the 1 Kbp ladder.
- Note that 2 μL of HindIII will twice as bright as 1 μL of HindIII, and 5 μL of HindIII will be five times as bright. So to determine concentration, estimate the intensity of the bands compared with the 3 HindIII ladders and assign it a relative intensity factor. Read off the concentration value on the Ladder card, and multiply by the relative intensity factor to determine your sample concentration in ng/μL.
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