2 replies to this topic

[Jen_S]

Hi,

 

This is probably a pretty simple issue. I'm currently separating bands between 200 to 300 bp in length. At present I'm using a 2% gel set at 65V for 40 mins, however, the resolution isn't great. I understand that better resolution can come from decreasing the voltage and increasing the run time, but I can't seem to work out what the relationship between the two is. For instance, if I decreased the voltage to 60 what should the increase in the time be? 50 minutes? 60 minutes?

 

Many thanks in advance

[perneseblue]

In this situation better resolution can be achieved by increase the agarose gel concentration to 2.5% or even 3%(if you are careful and get the agaorse powder to disperse before melting it)

 

Other alternatives would be to use an acrylamide gel. (~10%)

[bob1]

I would also say to run it for as long as possible - without running them off the gel.  If you can, use Orange G loading dye, which runs at about 50-100 bp, rather than using bromophenol blue, so that you don't accidentally run off the gel.