agarose gel melted after 4h running.. - (Feb/07/2007 )
Hi all,
I am running two 0.8%agarose gel in separate tank after the genomic DNA was digested with RE. The weird thing is one of the agarose gel melted after 4h of running. I run it at 60v and i prepared the gel together, buffer was the same. The only difference is the cast. The melted gel is 12cm x 10cm and the other one is 10 x 11cm. Do you have any idea what is wrong?
strange indeed for an agarose gel to even melt. for this to happen the agarose gel must be heated to more then 50 Celsius.
perhaps the current is too high?
My guess is that it has been the larger one because it creates more resistance to the current flow. Bigger the gel, higher resistance and probably higher agarose percentage higher the resistance (although I'm not sure about this).
However, I think it is strange in any case getting the gel melt. Try to leave it less time or decrease the voltage if you can. Test if something is going wrong with your gel bath.
One more thing easy to test is just to run gels without casting anything and see what happens. I bet that the melting is not due to what the cast is.
HI,
U may probably try casting a fresh gel with the same dimensions of that which melted and try running it in the buffer tank which workd fine. Also i have a doubt on ur power input probably the display on the powr pack is showing less. U can alternatively try low voltag and overnight run, i have done that with 25volts overnight for a 0.7%gel.
Cheers
I am running two 0.8%agarose gel in separate tank after the genomic DNA was digested with RE. The weird thing is one of the agarose gel melted after 4h of running. I run it at 60v and i prepare the gel together, buffer is the same. the only thing different is the cast is different the melted gel is 12cm x 10cm and the other one is 10 x 11cm. Do you have any idea what is wrong?
did you run them with the same power supply at the same time?
power will not proportion evenly if they are not identical. one of them (the melted gel) may have taken the lion's share of power.
how old are the leads?
Just a guess -- are you using 1x buffer, or did you accidentally use 10x buffer in your apparatus. The 10x buffer will have much higher conductivity, and will cause much higher power dissipation and melting.
Thanks for the reply.
I am using 1X TAE, the buffer were the same for the 2 tanks, but only one of the gel melted.
I am running them using same power supply at the same time. i guess that's one of the reason.
I am using old 0.5M EDTA to prepare 50X TAE, will it affect too?
The lead.. u mean the lead of the gel tank? 2-3 years. One of tank is newer. Does that affect too?
well, if one of the leads is faulty or damaged, you may not have the proper electrical connection all the way around
I think a buffer error is more likely, but if that is ruled out there may be a physical cause relating to integrity of the cords/apparatus
I am using old 0.5M EDTA to prepare 50X TAE, will it affect too?
when you run two or more gels from the same power supply they must be the same or you will not get equal distribution of the power. for agarose electrophoresis, not only do the gels have to have the same dimensions but the apparatus for each gel must be the same (doesn't have to be the same age, just the same model or dimensions).
this is the most likely reason that your gel melted.
how old is the edta? was the bottle closed tight to prevent evaporation? was it in a leaded glass bottle? was anything growing in it? was there any crystallization?
it was probably okay and not likely to have caused one gel to melt and not the other. that was caused by unequal distribution of power.