agarose gel electrophoresis - Question (Jul/23/2008 )

In order to find the concentration of DNA after running on an agarose gel... the commercial ladders one can use for this purpose gives the conc providing there is 50microgram of DNA per well... how does one know how much DNA i.e microlitre of DNA u have to load in the well

-Amritha Nair-

i may be wrong here but i thought the ladders gave you the size of the lumps not the conc of dna (more dna = more distinct band)

-Dominic-

See the attaced Image; you have a specified concentration of DNA for every size. This means: if your PCR product is as intense as the 1000 bp of this ladder you have 100 ng of DNA in the amount of PCR product you have loaded on the gel; lets say you loaded 4µL of PCR product; so you have 25 ng per µL PCR product (=100/4). If you are using less or more ladder than recomended by the manufacturer you have to divide/multiply the concentrations.

-gebirgsziege-

QUOTE (gebirgsziege @ Jul 24 2008, 03:57 AM)
See the attaced Image; you have a specified concentration of DNA for every size. This means: if your PCR product is as intense as the 1000 bp of this ladder you have 100 ng of DNA in the amount of PCR product you have loaded on the gel; lets say you loaded 4µL of PCR product; so you have 25 ng per µL PCR product (=100/4). If you are using less or more ladder than recomended by the manufacturer you have to divide/multiply the concentrations.

wow!!! thanks a lot... u just cleared up my confusion

-Amritha Nair-