Estimating size of PCR products in a agarose gel - (Mar/08/2012 )
Greetings! I want to determine the size of bands in a PCR. I have downloaded ImageJ but after reading and reading I still dont know how to measure the size of the bands. In my gel, I will have the ladder and then some samples, I just want to know exactly the size of the products. I have read that with ImageJ you can determine the density (amount) of sample, but I will love if someone can explain whow to measure the size of the products.
There should be a key for the ladder you use. It will have each band size marked in base pairs. Also, it will say how many nanograms each band is assuming you loaded the proper amount (usually 5uL). Then you simply compare your experimental bands to the marker and you can estimate size and concentration. If imageJ is a densitometer, then can actually compare band intensities. Just make sure you load too much or you will saturate the signal!
Thanks, just to clarify, so in ImageJ is a way in which I can tell it the size of each band of the ladder and the concentration? Can you please just direct me to which command I have to look for that? Thanks!!
here is a link to imagej documentation on gel analysis.
Especially if you probe more than 1 or 2 samples, it is nice to have a ladder on each site of your gel in order to connect the corresponding ladder-bands to make a kind of grid. this should be appropriate to estimate your band size, you don't need a special program for that