Cumbers:protocols/picogreen quantIT: Difference between revisions
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*Make up standard measurements along with your samples. | *Make up standard measurements along with your samples. | ||
*If unsure of your scale (if you have a lot or a little) then do 3 concentrations of your sample to check, one at 100% concentration, one at 1:10 other at 1:100 | *If unsure of your scale (if you have a lot or a little) then do 3 concentrations of your sample to check, one at 100% concentration, one at 1:10 other at 1:100 | ||
*add 1ul DNA to 9ul TE to make 1:10, then | *First you are making a 1:10 (add 1ul DNA to 9ul TE to make 1:10), then you are taking 1 uL of this 1:10 solution and adding it to 99 uL. So that is 1:100 of the 1:10 so the solution where you took 1 uL of this 1:10 solution and added to 99ul TE is 1:1000 of the original DNA concentration (1:10 * 1:100). | ||
*Do 2 of TE or whatever you are resuspending in to subtract this background from your results. | *Do 2 of TE or whatever you are resuspending in to subtract this background from your results. | ||
*set up the plate reader and computer. | *set up the plate reader and computer. |
Revision as of 10:36, 31 January 2007
- Follow the Quant-it protocol for DNA (and RNA) quantification.
- Make up standard measurements along with your samples.
- If unsure of your scale (if you have a lot or a little) then do 3 concentrations of your sample to check, one at 100% concentration, one at 1:10 other at 1:100
- First you are making a 1:10 (add 1ul DNA to 9ul TE to make 1:10), then you are taking 1 uL of this 1:10 solution and adding it to 99 uL. So that is 1:100 of the 1:10 so the solution where you took 1 uL of this 1:10 solution and added to 99ul TE is 1:1000 of the original DNA concentration (1:10 * 1:100).
- Do 2 of TE or whatever you are resuspending in to subtract this background from your results.
- set up the plate reader and computer.
- Add the fluorescent reagent (make up enough: 100 assays = 20 ml HS buffer) - 200ul added to each plate, make sure you have enough and careful when pipetting, keep track of which hole you want to add it to.
- read the plate
1 2 3 4 A 10 10 1C 1S B 8 8 1C(1:10) 1S(1:10) C 6 6 1C(1:10) 1S(1:10) D 4 4 1C(1:10) 1S(1:10) E 2 2 1C(1:100) 1S(1:100) F 1 1 1C(1:100) 1S(1:100) G 0.5 0.5 1C(1:100) 1S(1:100) H 0 0 TE TE