Toronto/Lab Protocols/Quantitation

From 2007.igem.org

(Difference between revisions)
(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.
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          * 1 μL plasmid + 4 μL ddH2O + 1 μL Loading Dye
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*1 μL plasmid + 4 μL ddH2O + 1 μL Loading Dye
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           * 1 μL insert + 4 μL ddH2O + 1 μL Loading Dye
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           * *      Gel Lane Fragment
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      Gel Lane Fragment
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       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

  1. 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)
  1. Ensure that the plasmid and inserts are at the correct lengths using the 1 Kbp ladder.
  2. 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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