too large bands on Agarose gel

[Silvia_84]

Hi all,

 

im am new in doing electrophoresis on agarose gel. I tried to run a couple of gels and my markers are not separated appropriately. I have to say that the bands corresponding to both the marker and to my DNA samples are so large that I am not able to interpret the results. Even if I run the gel for more time, I obtain always the same result. On what it could depend? on the amount of sample I load? I usually load 5 ul of sample.

 

Thank you in advance.

 

 

[CharonY]

You have to be more precise in your description. If by too large you mean non sufficient separation of high MW bands you are likely to have a too high percentage gel. If the bands are too thick but otherwise resolved, your sample is too concentrated. The volume does not matter as long as it fits into the pockets.

[Silvia_84]

Yes, sorry , I did not explain appropriately. The bands are so thick that they overlap and the separation is not visible..the ladder appears as a continue vertical line in certain points. I prepared a 1.5 % of agarose gel in 1XTAE buffer.

[CharonY]

Are they thick bands or appear like a smear? In the former case you need to dilute your sample. In the latter you may have degradation.